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Biomedical subjects

J Silver

Publications and source records attributed to J Silver.

At least 163 records · Page 9Linked to original sources
PubMed1995

V beta-specific activation of T cells by the HIV glycoprotein gp 160.

Studies by several groups have suggested that HIV infection in vivo results in a V beta-specific alteration of the TCR repertoire and that this might play a role in the pathogenesis of AIDS. However, there is very little agreement as to which V beta segments are affected. In order to circumvent the confounding factors present in vivo we have examined the abilities of both a crude protein extract of HIV and purified gp160 to alter the V beta repertoire of normal T cells in vitro. We find that both a crude extract of HIV as well as gp160 specifically activate T cells expressing a common set of V beta segments (V beta 3, 12, 14, 15, and sometimes V beta 17 and 20) in individuals of disparate HLA type. This set of V beta segments is remarkably similar to those recognized by staphlococcal enterotoxin B and supports the hypothesis that bacterial superantigens produced by opportunistically acquired micro-organisms could have an exacerbating effect in AIDS.

Antibodies, Monoclonal↗
PubMed1995

The influence of non-HLA genes on the human T-cell receptor repertoire.

We previously demonstrated a central role for HLA genes in determining the T-cell receptor (TCR) repertoire. However, these studies also suggested that other genetic factors might also play a role in the development of this repertoire. In order to assess the role of non-HLA genes in the development of the TCR repertoire, we have analysed and compared the TCR repertoires of individuals in three families consisting of both monozygotic twins as well as an HLA-identical sib. TCR repertoire analysis was performed with both V-segment-specific MoAb and the polymerase chain reaction using TCRBV-segment-specific oligonucleotide primers. We observed that in every case the TCR repertoires of identical twins were more similar to each other than to their HLA-identical sib. Furthermore, in one family we were able to show by genotype analysis that most of the differences in repertoire between the identical twins and their HLA-identical sib were caused by polymorphisms in the TCR genes that influence expression levels. These studies document an important role for non-HLA genes in determining the TCR repertoire in man and raise the possibility that such TCR polymorphisms may play a significant role in determining disease susceptibility.

Base Sequence↗
PubMed1995

Differential patterns of T-cell receptor BV-specific activation of T cells by gp120 from different HIV strains.

Studies by several groups have suggested that HIV infection in vivo results in a BV-specific alteration of the TCR repertoire and that this might play a role in the pathogenesis of AIDS. Our earlier studies demonstrated that both a crude extract of HIV451 as well as purified gp 160 from HIV451 could specifically activate, in vitro, T cells expressing a common set of TCRBV segments (TCRBV3, 12, 14, 15, and sometimes BV17 and 20) in individuals of disparate HLA type. Furthermore, purified gp120 from HIV451 was shown to have a similar ability to activate T cells, although with a slightly different TCRBV-specific pattern. In order to determine whether gp120 from other HIV strains could similarly activate T cells in a TCRBV-specific pattern, PBMC from HIV seronegative individuals of disparate HLA type were stimulated with gp120 from three strains of HIV (451, IIIB, and MN). The authors found that gp120 from all three strains activate T cells bearing TCRBV2 and BV3 in nearly every individual. T cells expressing other BV segments are also activated, but this is more variable and appears to be unique to each individual. Furthermore, gp120(451) and gp120 from HIVIIIB and HIVMN differ in their ability to activate T cells expressing these other TCRBV segments. These observations suggest that variation in the structure of gp120 and in the genetic and/or environmental background of the individual play an important role in determining which TCRBV segments are 'triggered' by gp120. Furthermore, these observations may have important implications for the rate of disease progression in HIV-infected individuals.

Cells, Cultured↗
PubMed1995

Replacement of the DR alpha chain with the E alpha chain enhances presentation of Mycoplasma arthritidis superantigen by the human class II DR molecule.

Mycoplasma arthritidis mitogen (MAM) is produced by an organism which can cause chronic proliferative arthritis in rodents. MAM possesses a typical superantigenic activity; it has the ability to activate a large panel of T cells which express specific V beta segments of the T-cell receptor. The presentation of MAM to T cells by antigen-presenting cells is mediated primarily through its binding to the major histocompatibility complex (MHC) class II E alpha chain in mice and the DR alpha chain in humans. However, MAM is much less active for human peripheral blood lymphocytes than for mouse splenocytes. It was suggested that a difference in MAM binding affinity between human and mouse class II molecules may account for their different MAM activities. To examine this possibility, we generated a panel of B-cell transfectants whose DR molecule is composed of either the DR alpha or the E alpha chain paired with a DR3 beta chain. The ability of these transfectants to present MAM to human peripheral T cells was analyzed. Our data show that transfectants expressing E alpha DR beta chimeric molecules have higher MAM-presenting activity than transfectants expressing wild-type DR alpha DR beta molecules, while the latter have higher activity in stimulating DR3-alloreactive T cells. Since both types of transfectants present MAM to T cells expressing the same T-cell receptor V beta gene families, the higher MAM-presenting activity of the E alpha transfectant is not due to its ability to interact with a different set of T cells. Furthermore, both the E alpha 1 and E alpha 2 domains contribute to this increased affinity for MAM binding. Taken together, our data suggest that there may be multiple MAM binding sites on the E alpha and DR alpha chains and residues unique to the E alpha chain may provide additional affinity for MAM.

Antigen Presentation↗
PubMed1995

Low vision in east African blind school students: need for optical low vision services.

AIMS: There is increasing awareness of the needs of children with low vision, particularly in developing countries where programmes of integrated education are being developed. However, appropriate low vision services are usually not available or affordable. The aims of this study were, firstly, to assess the need for spectacles and optical low vision devices in students with low vision in schools for the blind in Kenya and Uganda; secondly, to evaluate inexpensive locally produced low vision devices; and, finally, to evaluate simple methods of identifying those low vision students who could read N5 to N8 print after low vision assessment. METHODS: A total of 230 students were examined (51 school and 16 university students in Uganda and 163 students in Kenya, aged 5-22 years), 147 of whom had a visual acuity of less than 6/18 to perception of light in the better eye at presentation. After refraction seven of the 147 achieved 6/18 or better. Eighty two (58.6%) of the 140 students with low vision (corrected visual acuity in the better eye of less than 6/18 to light perception) had refractive errors of more than 2 dioptres in the better eye, and 38 (27.1%) had more than 2 dioptres of astigmatism. RESULTS: Forty six per cent of students with low vision (n = 64) could read N5-N8 print unaided or with spectacles, as could a further 33% (n = 46) with low vision devices. Low vision devices were indicated in a total of 50 students (35.7%). The locally manufactured devices could meet two thirds of the need. CONCLUSION: A corrected distance acuity of 1/60 or better had a sensitivity of 99.1% and a specificity of 56.7% in predicting the ability to discern N8 print or better. The ability to perform at least two of the three simple tests of functional vision had a sensitivity of 95.5% and a specificity of 63.3% in identifying the students able to discern N8 or better.

Adolescent↗
PubMed1995

Coordinate regulation of rat renal parathyroid hormone receptor mRNA and Na-Pi cotransporter mRNA and protein.

Parathyroid hormone (PTH) acts on the kidney by binding to the PTH receptor, leading to a decrease in the active renal reabsorption of phosphate by the Na-Pi cotransporter, which is also independently activated by hypophosphatemia. We have studied the effects of hypo- and hyperparathyroidism and hypophosphatemia on PTH receptor mRNA and Na-Pi cotransporter mRNA and protein. Both surgical parathyroidectomy and hypophosphatemia, which itself leads to hypoparathyroidism, led to an upregulation of the PTH receptor mRNA and Na-Pi cotransporter mRNA and protein. Parathyroidectomized rats fed a low-Pi diet had an increase in PTH receptor and Na-Pi cotransporter mRNAs. Diet-induced hyperparathyroidism had no effect on PTH receptor mRNA and Na-Pi cotransporter mRNA and protein. The effect of hypoparathyroidism and hypophosphatemia to increase both PTH receptor mRNA and Na-Pi cotransporter mRNA and protein shows that there is a tight coordinate regulation of these factors, which are both involved in PTH action.

Animals↗
PubMed1995

Parathyroid hormone gene expression in hypophosphatemic rats.

Phosphate is central to bone metabolism and we have therefore studied whether parathyroid hormone (PTH) is regulated by dietary phosphate in vivo. Weanling rats were fed diets with different phosphate contents for 3 wk: low phosphate (0.02%), normal calcium (0.6%), normal phosphate (0.3%), and calcium (0.6%); high phosphate (1.2%), high calcium (1.2%). The low phosphate diet led to hypophosphatemia, hypercalcemia, and increased serum 1,25(OH)2D3 together with decreased PTH mRNA levels (25 +/- 8% of controls, P < 0.01) and serum immunoreactive PTH (4.7 +/- 0.8: 22.1 +/- 3.7 pg/ml; low phosphate: control, P < 0.05). A high phosphate diet led to increased PTH mRNA levels. In situ hybridization showed that hypophosphatemia decreased PTH mRNA in all the parathyroid cells. To separate the effect of low phosphate from changes in calcium and vitamin D rats were fed diets to maintain them as vitamin D-deficient and normocalcemic despite the hypophosphatemia. Hypophosphatemic, normocalemic rats with normal serum 1,25(OH)2D3 levels still had decreased PTH mRNAs. Nuclear transcript run-ons showed that the effect of low phosphate was posttranscriptional. Calcium and 1,25(OH)2D3 regulate the parathyroid and we now show that dietary phosphate also regulates the parathyroid by a mechanism which remains to be defined.

Animals↗
PubMed1995

Parathyroid cell proliferation in normal and chronic renal failure rats. The effects of calcium, phosphate, and vitamin D.

Secondary hyperparathyroidism is characterized by an increase in parathyroid (PT) cell number, and parathyroid hormone (PTH) synthesis and secretion. It is still unknown as to what stimuli regulate PT cell proliferation and how they do this. We have studied rats with dietary-induced secondary hyper- and hypoparathyroidism, rats given 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) and rats after 5/6 nephrectomy for the presence of PT cell proliferation and apoptosis. PT cell proliferation has been measured by staining for proliferating cell nuclear antigen (PCNA) and apoptosis by in situ detection of nuclear DNA fragmentation and correlated with serum biochemistry and PTH mRNA levels. A low calcium diet led to increased levels of PTH mRNA and a 10-fold increase in PT cell proliferation. A low phosphate diet led to decreased levels of PTH mRNA and the complete absence of PT cell proliferation. 1,25 (OH)2D3 (25 pmol/d x 3) led to a decrease in PTH mRNA levels and unlike the hypophosphatemic rats there was no decrease in cell proliferation. There were no cells undergoing apoptosis in any of the experimental conditions. The secondary hyperparathyroidism of 5/6 nephrectomized rats was characterized by an increase in PTH mRNA levels and PT cell proliferation which were both markedly decreased by a low phosphate diet. The number of PCNA positive cells was increased by a high phosphate diet. Therefore hypocalcemia, hyperphosphatemia and uremia lead to PT cell proliferation, and hypophosphatemia completely abolishes this effect. Injected 1,25 (OH)2D3 had no effect. These findings emphasize the importance of a normal phosphate and calcium in the prevention of PT cell hyperplasia.

Animals↗
PubMed1995

Oligoclonality in the human CD8+ T cell repertoire in normal subjects and monozygotic twins: implications for studies of infectious and autoimmune diseases.

BACKGROUND: We have previously demonstrated CD8+ T cell clonal dominance using a PCR assay for the CDR3 length of T cell receptors belonging to a limited number of TCRBV segments/families. In this study, we have modified this approach in order to analyze more comprehensively the frequency of oligoclonality in the CD8+ T cell subset in 25 known TCRBV segments/families. In order to assess the relative roles of genes and environment in the shaping of a clonally restricted CD8+ T cell repertoire, we have analyzed clonal dominance in the CD8+ T cell population of monozygotic twins, related siblings, and adoptees. MATERIALS AND METHODS: Oligoclonality was assessed in the CD8+ T cell subsets using a multiplex PCR approach to assay for CDR3 length variation across 25 different TCRBV segments/families. Specific criteria for oligoclonality were established, and confirmed by direct sequence analysis of the PCR products. This assay was used to investigate the CD8+ T cell repertoire of 56 normal subjects, as well as six sets of monozygotic (MZ) twins. RESULTS: Seventy-two percent of normal subjects (n = 56) had evidence of oligoclonality in the CD8+ T cell subset, using well-defined criteria. Although MZ twins frequently displayed CD8+ T cell clonal dominance, the overall pattern of oligoclonality was very diverse within each twin pair. However, we occasionally observed dominant CD8+ T cell clones that were highly similar in sequence in both members of some twin pairs. Not a single example of such similarity was observed in normal controls or siblings. CONCLUSIONS: Oligoclonality of circulating CD8+ T cells is a characteristic feature of the human immune system; both host genetic factors and environment shape the pattern of oligoclonality in this T cell subset. The high frequency of this phenomenon in normal subjects provides a background with which to evaluate CD8+ T cell oligoclonality in the setting of infection or autoimmune disease. Further phenotypic and functional characterization of these clonally expanded T cells should provide insight into normal immune homeostasis.

Adolescent↗
PubMed1994

Chondroitin sulfate proteoglycans are a common component of neuronal inclusions and astrocytic reaction in neurodegenerative diseases.

Previously, we showed three differentially sulfated forms of chondroitin sulfate proteoglycans (CSPG) associated with senile plaques, astrocytes and neurofibrillary tangles in Alzheimer's disease. Here, monoclonal antibodies were used to demonstrate CSPGs in other neurodegenerative diseases. CSPGs were found associated with inclusions of Parkinson's, diffuse Lewy body, Pick's diseases, and progressive supranuclear palsy. Reacting astrocytes in each of these neurodegenerative diseases and Huntington's disease showed immunoreactivity for CSPG. CSPG distribution in a variety of neurodegenerative diseases suggests that similar mechanisms may be involved in the accumulation of proteoglycans in a number of filamentous inclusions.

Aged↗
PubMed1994

Immortalization of immature and mature mouse astrocytes with SV40 T antigen.

The ability of neonatal astrocytes to promote neurite outgrowth in vitro and in vivo diminishes as astrocytes mature. This property correlates with the developmental loss of the central nervous system's ability to regenerate after injury. Cell lines representative of immature and mature astrocytes would be useful for studies to determine differences between these two populations. Previous work on immortalization of bipotential neural/glial precursors and fully differentiated glial cells suggests that immortalization of astrocytes at timed intervals of culture may yield cell lines trapped in different maturation states. To test this, neonatal mouse cortical astrocytes were immortalized by retrovirus-mediated transfer of the SV40 T antigen (Tag) gene at 2, 6 and 17 days of culture. The clonal cell lines express Tag and are contact-inhibited. Three phenotypes that change as a function of astrocyte maturation were examined to determine the fidelity with which the cell lines represent immature and mature astrocytes. These were: (1) cell morphology, growth pattern and size, (2) level of glial fibrillary acidic protein (GFAP) expression, and (3) neurite outgrowth promotion. First, immature and mature lines resemble mortal type 1 astrocytes of corresponding ages with respect to morphology and growth pattern, and retain a quantitative difference in cell size (mature cells are larger). Second, the pattern of GFAP expression is preserved, with immature lines expressing lower levels than mature cell lines, but the overall GFAP levels are significantly lower in immortalized cell lines compared to mortal cells. Finally, promotion of neurite outgrowth from embryonic chick retinal ganglion cells on monolayers of the cell lines was examined. While all neurite outgrowth measures are significantly greater for the immortalized lines than for control 3T3 cells, they are attenuated relative to mortal astrocytes. The age-related pattern of stronger outgrowth support on immature astrocytes is retained for neurite initiation, but not retained for mean neurite length. Thus, SV40 Tag-immortalized astrocytes have a complex phenotype characterized by retention of age-related differences in morphology, growth pattern and cell size, and by a marked attenuation of some astrocyte-specific characteristics but retention of age-related differences in the expression level of these same characteristics, and finally, loss of the ability to support neurite extension at level characteristic of immature astrocytes.

3T3 Cells↗
PubMed1994

Exogenous glycosaminoglycans induce complete inversion of retinal ganglion cell bodies and their axons within the retinal neuroepithelium.

Prior to forming an axon, retinal ganglion cells retain a primitive radial configuration while maintaining ventricular and vitreal endfeet attachments. During their subsequent differentiation, ganglion cells polarize their cell body and axon only along the vitreal surface. When the ventricular surfaces of intact retinas in organ culture were exposed to free chondroitin sulfate (CS) in solution, both the cell body and nerve fiber layers were repolarized to the opposite side of the neuroepithelium. However, the basal lamina remained in its usual position. Thus, the ability to initiate an axon is not restricted to the vitreal endfoot region of differentiating neurons, and in addition, the radial position at which the axon emerges can be mediated by the location and concentration of the extracellular CS milieu.

Animals↗
PubMed1994

Recombinant soluble CD14 inhibits LPS-induced tumor necrosis factor-alpha production by cells in whole blood.

CD14 functions as a cell surface receptor for LPS in the activation of monocytes/macrophages and neutrophils by endotoxin. To assess the utility of soluble forms of the CD14 receptor as a possible therapeutic for endotoxin shock, we have produced recombinant human soluble CD14 using a baculovirus expression system. We find that the recombinant protein is not only expressed on the surface of the insect cells as a glycosyl phosphatidylinositol (GPI)-anchored protein, but is also released into the culture medium as a soluble form that lacks the GPI anchor. Functional analyses of recombinant human soluble CD14 show that it binds specifically to LPS and can inhibit the LPS-induced release of TNF-alpha by macrophages and mononuclear cells as well as by cells in whole human blood when used at concentrations of approximately 70 micrograms/ml. Thus, soluble CD14 may be useful as an adjunct in the treatment of endotoxin shock.

Animals↗
PubMed1994

Erythrocytosis after renal transplantation. The response to theophylline treatment.

Erythrocytosis was found in 20 out of 127 patients who underwent renal transplantation (15.7%). Four were found to have elevated erythropoietin concentrations (139.7 +/- 22.0 microns/ml) and 7 had normal levels (52.3 +/- 84 microns/ml (normal -60). The growth of erythroid CFUs was found to be normal in all patients. Theophylline treatment was given to 11 patients. The patients were divided into 3 groups according to their response: (1) patients with high erythropoietin levels whose hematocrit decreased from 59.6% to 46.3% within 2 weeks of treatment in parallel with a fall in erythropoietin; (2) patients with normal erythropoietin levels in whom theophylline therapy reduced the hematocrit from 58.7% to 47.2%; and (3) patients in whom theophylline had no effect on their erythrocytosis. We conclude that erythrocytosis after renal transplantation is a heterogenous condition and that several mechanisms are involved. In some patients, high erythropoietin levels or high sensitivity to erythropoietin are associated with the erythrocytosis. Theophylline therapy is beneficial in more than 50% of the cases.

Adult↗
PubMed1994

Unique phenotype and distinct TCR V beta repertoire in human peripheral blood alpha beta TCR+, CD4-, and CD8- double negative T cells.

TCR-alpha beta+ CD4- CD8- double-negative (DN) T cells represent a small, poorly defined T cell subset in human peripheral blood that has been postulated to be potentially autoreactive. To define some of the characteristics of this subset of T cells, DN cells and CD4+ and CD8+ single-positive (SP) cells were purified from the peripheral blood of six unrelated individuals by a combination of positive selection and depletion using mAb conjugated to immunomagnetic beads. Purified DN cells were found to be enriched in cells expressing HLA-DR and the NK cell marker, CD56, when compared to the SP population. Furthermore, in contrast to SP cells that express the adhesion marker CD44, DN cells were found to express very little, if any, CD44. When the V beta TCR repertoires of DN and SP (CD4+ and CD8+) cells, determined by quantitative (q) PCR, were compared all three populations were found to be considerably different. Furthermore, several V beta segments (V beta 11 and V beta 19) were consistently expressed at higher levels on DN cells than on SP cells. The TCR repertoires of both DN and SP cells were frequently characterized by dominance of one or more V beta segments that could in some instances be shown to be restricted to the CD45RO+ ("memory") population. However, differences in TCR repertoire between DN and SP cells were observed even when CD45RO+ cells were removed before qPCR analysis. These studies suggest that the TCR repertoires of DN and SP cells are determined by different selection mechanisms and that DN and SP cells are directed against different Ag.

CD4 Antigens↗
PubMed1994

Regional differences in reactive gliosis induced by substrate-bound beta-amyloid.

Regions of gliosis surround deposits of beta-amyloid peptide (beta AP) in senile plaques of Alzheimer's disease (AD). The association between reactive astrocytes and beta AP in senile plaques is most pronounced in cortex and hippocampus but not at other anatomical sites of beta AP deposition. We hypothesized that this region-specific pathology in AD could be attributed to differences in glial reactivity in different parts of the central nervous system (CNS). To test this hypothesis, we assayed astrocytes from cerebral cortex, hippocampus, cerebellum, and spinal cord for cellular responsiveness to substrate-bound beta AP in vitro. Astrocyte reactivity was monitored by morphological changes, increased deposition of chondroitin sulfate proteoglycan-containing matrix, and alterations in proteoglycan metabolism. Based on these criteria, only cortical and hippocampal astrocytes showed marked reactivity to immobilized beta AP. In cortical and hippocampal cultures only, immobilized beta AP resulted in increased total radiosulfate incorporation into proteoglycans which was mainly found in the cell/matrix rather than in the media-associated compartment. There were also differences in the proteoglycan synthesis patterns of astrocyte cultures isolated from these CNS regions. These findings suggest that (1) astrocytes are regionally heterogenous in their reactive response to beta AP and (2) that specific molecules, in addition to beta AP, may exist following trauma or disease which trigger reactive states in astroglia in the cerebellum or spinal cord. These local differences in the interaction between beta AP and surrounding astrocytes may play a role in the region-specific pathogenesis of Alzheimer's disease.

Amyloid beta-Peptides↗
PubMed1994

Mössbauer spectra of the heme peptide (HP) 1-50 and the heme peptide:non-heme peptide (NHP) non-covalent complex 1-50:51-104 derived from cytochrome c: evidence for cytochrome c iron site solvation in aqueous solution.

Mössbauer spectroscopic studies on a heme peptide (HP) derived from cytochrome c and on the HP recombined non-covalently with the remaining cleaved section are reported. The results suggest that the environment of the heme site in the known crystal structure of cytochrome c may differ in detail from the environment of the heme in the working protein.

Cytochrome c Group↗