ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

ObjectiveTo elucidate the tissue specificity of the expression of HIV-1 genes in an animal and its pathological effects on these tissues.Design and methodsTransgenic mice carrying a defective HIV-1 genome were bred in order to overcome the host–range barrier of this virus.ResultsmRNA specific to the transgene was detected in the eyes and the spleen, and, in smaller quantities, in the thymus and the brain. Interestingly, many of the transgenic mice developed cataracts at 3–6 months of age. Swelling and vacuolation of the lens fiber cells were marked, but the epithelial cells of the lens were less affected. HIV antigens were detected in the lens fiber cells and the retina by immunological staining. Accumulation of large amounts of p24 Gag antigen was demonstrated in the affected lens by immunoblot analysis, while negligible Env or other viral proteins was detected. Although accumulation of the Gag protein was also detected in the skin and the brain, no apparent abnormality was observed in these tissues.ConclusionsPreferential expression of the HIV genes in the eyes, skin, brain and lymphoid tissues was demonstrated. The accumulation of the Gag protein is suggested to have detrimental effects on lens fiber cells, causing cataracts.

ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

ObjectiveSpecific cytotoxic T-lymphocytes (CTL) are induced in humans or monkeys after infection with HIV-1 or SIVmac, respectively. Since, like HIV-1, HIV-2 causes AIDS, our objective was to determine the characteristics of the HIV-2-specific CTL response.DesignSince it is rarely possible to study cellular immunity in individuals, because of the small number of HIV-2-infected patients available in Europe and the necessity for co-operation in the performance of sequential CTL assays, cynomolgus macaques were infected with HIV-2. Autologous transformed B-lymphoblastoid cell lines infected with recombinant vaccinia viruses were used as target cells for cytotoxicity assays.MethodsRecombinant vaccinia viruses expressing HIV-2 genes were constructed to infect B-lymphoblastoid cell lines from macaques. These cells were used as target cells for cytotoxicity assays with peripheral blood mononuclear cells from HIV-2BEN-infected cynomolgus macaques. To characterize the effector cells, CD8+ cells were separated with immunomagnetic beads. Major histocompatibility complex (MHC) restriction of the cytotoxic cells was determined by incubation with matched or mismatched target cells.ResultsHIV-2BEN-infected cynomolgus macaques raised CTL against proteins of the three major viral structural genes, gag,polandenv.The cytotoxic cells were CD8+ and their activity was MHC class I-restricted. In contrast to SIVmac-infected macaques,env-specific lysis was mediated exclusively by CD8+ cells. CTL from individual animals recognized different viral proteins and the recognition pattern varied over time.ConclusionsLike HIV-1 and SIVmac, HIV-2 induces virus-specific CTL. The variation of antigen recognition between individual animals and over time indicates that sequential experiments are necessary to determine the complete spectrum of the CTL response of infected animals. HIV-2-infected macaques represent a suitable model for investigations into the cellular immune response against HIV.

ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

ObjectiveTo evaluate specific anti-HIV cytotoxic T-lymphocyte (CTL) activity in relation to basic clinical and laboratory parameters used to follow HIV infection.MethodsLymphocytes from HIV-1-infected subjects with different clinical and immunologic features of HIV infection were tested for circulating and inducible anti-HIV CTL activity using autologous B-lymphoblastoid cells infected with recombinant vaccinia viruses expressing the HIVgag, polandenvgenes as targets. Anti-HIV CTL were induced by stimulation with HIV-infected autologous lymphoblastsin vitro.ResultsWe detected circulating anti-HIV CTL in asymptomatic subjects exclusively and found a significant association (PP<0.001), but there was no significant difference in mean CD4+ lymphocyte count. CD8+ human histocompatibility leukocyte antigen (HLA) class l-restricted anti-HIV CTL were induced in all HIV-infected subjects tested following stimulation with HIV-infected autologous lymphoblastsin vitro.In subjects without detectable circulating anti-HIV CTL, circulating HLA-DR+ cells contributed to anti-HIV CTL activity induced by stimulation with HIV or concanavalin Ain vitro.ConclusionsCirculating anti-HIV CTL activity is associated with CD8+ lymphocyte counts ≥900/μl. Anti-HIV CTL retain proliferative and functional capacity followingin vitrostimulation with HIV and interleukin-2 through all stages of HIV infection. Persistent inducible anti-HIV CTL activity in subjects with advanced HIV disease and without circulating CTL suggests impaired activation and/or proliferation of the CTLin vivo.

ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

ObjectiveTo characterize an HIV-1 quasispeciesin vivoat high resolution (1%) in order to determine its genetic structure.MethodsThe first coding exon oftatwas amplified by polymerase chain reaction from uncultured peripheral blood mononuclear cells of an HIV-1-infected patient. The products were cloned into M13mp18 RF, and 106 clones were sequenced.ResultsThirty-one different Tat protein variants were found. Amongst these, five major forms with frequencies of 44, 11, 8 and 5% were identified. All of the remaining 26 sequences were unique, 15 of which were defective. Within the variant spectrum a small number of genomes encoded novel open reading frames, for example, atat–vpufusion product.ConclusionSome of the myriad proviruses present in an individual harbour novel coding sequences. While these are probably of little importance for AIDS pathogenesis they emphasize the ability of HIV to explore a huge range of genetic configurations.

ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

ObjectiveTo directly infect human fetal intestine with HIVin vitro.DesignHuman fetal intestinal explant cultures were exposed to HIV-1 and monitored for evidence of infection by biochemical assay, immunohistochemistry andin situhybridization.MethodsHuman fetal intestinal explants (14–21 weeks) were established in culture and exposed to HIV-1. Tissue culture fluid was assayed for p24 antigen and reverse transcriptase activity over a 14-day period. Explants were removed from culture on days 4, 7, 10 and 14 postinoculation and subjected (1) to immunohistochemistry to detect p24 and gp160/41 antigens, and (2) toin situhybridization to detect HIV-1 RNA. Explant tissue culture fluid was cocultured with Jurkat T-cells to detect infectious viral particles.ResultsReverse transcriptase activity and p24 antigen levels in fetal explant culture fluid rose between 7 and 14 days after viral inoculation. Jurkat T-cell cocultures confirmed the presence of infectious virus. Cells in the lamina propria resembling lymphocytes and macrophages of both small intestine and colon stained positively for the viral proteins p24 and gp41. The same type of cells also stained positively for HIV-1 RNA usingin situhybridization. Dual-label immunohistochemistry, combined immunohistochemistry andin situhybridization confirmed the presence of viral protein and RNA in cells bearing the CD3, CD4 (lymphocyte) or CD68 (macrophage) surface markers. There was no evidence at any time of HIV-1 infection of epithelial cells.ConclusionsCells of the lamina propria of the small intestine and colon, bearing lymphocyte or macrophage markers, can be directly infected by and support the replication of HIV-1. Such infection may be implicated in the pathogenesis of HIV enteropathy.

ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

ObjectiveTo study cell surface molecules and HIV-1 proteins on H9 cells 2 days after infection by immunogold electron microscopy, either in single or in double labelling using combinations of host cell-derived molecules and HIV-1 proteins.Design and methodsThe presence of host cell antigens CD3, CD4 and human leukocyte antigen-DR (HLA-DR) and HIV-1 antigens gag p15, p17, p24 andenvgp41 was evaluated using immunocytochemistry at the light microscopic level. H9 cells 2 days after infection were processed for conventional transmission electron microscopy and cryo-ultramicrotomy. Leukocyte antigens investigated were CD2, CD3, CD4 (two antibodies), CD5, CD8, CD25, CD30, CD63 antigens and HLA-DR; HIV-1-encoded antigens were gag p24,polreverse transcriptase, andenvgp41 and gp120. Double immunogold labelling was performed using reagents with different sized gold particles. For leukocyte markers, the labelling density of the cell membrane was assessed quantitatively on uninfected and infected H9 cells.ResultsInfected cells revealed the presence ofgagp24,pol, andenvgp41 and gp120 antigens on HIV-1 virions. Uninfected H9 cells showed a random distribution of cell surface molecules, including CD4 antigen, along the plasma membrane. The CD63 antigen, a lysosomal membrane glycoprotein, was located mainly in the cytoplasm of uninfected cells. Cells 2 days after infection showed CD4 labelling on sites where virions were budding from or attached to the cell surface and on free virions. Virions also showed labelling by CD3, CD5, CD25, CD30 and CD63 antibodies and anti-HLA-DR. Compared with uninfected cells, a significantly lower density was found on infected cells in labelling for CD4, CD5 and anti-HLA-DR. A significantly higher density on cells 2 days after infection was seen in CD63 labelling.ConclusionDuring the first phase of infection host cell molecules concentrate on budding structures and newly generated HIV-1 virions. This phenomenon might contribute to the disappearance of these molecules (like the CD4 molecule) from the cell membrane after infection.

ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

ObjectiveTo evaluate the time-course of HIV-1 detection in peripheral blood mononuclear cells (PBMC) from newborns at risk of vertically acquired infection.Design and methodForty-six infants born to HIV-1-infected mothers were enrolled at birth and examined virologically and clinically in the perinatal period and every 30 days for the first 3 months of life. Follow-up was conducted at intervals of 2–3 months. HIV-1 detection in PBMC was performed using virus culture and polymerase chain reaction (PCR) assay.ResultsOnly one out of 24 newborns tested within 48 h of delivery and two out of 22 infants tested between 3 and 15 days of age were found to be HIV-1-positive by both PCR and virus culture. Further testing performed between 30 and 60 days of life identified an additional eight HIV-1-positive children. Subsequent viral, immunological and clinical follow-up confirmed PCR and virus culture results obtained in 30–60-day-old children.ConclusionsInfected infants had detectable levels of HIV-1 in their PBMC at 1 month of age. The negative PCR and virus culture findings in PBMC of newborns indicate strongly that HIV-1 cannot be diagnosed at birth in the majority of cases, and suggests that viral transmission could occur during late pregnancy and/or delivery.

ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

ObjectivesTo determine the amount of circulating CD4+ cells positive for intracellular p24 antigen during HIV infection, and to correlate the results with clinical, virological and therapeutic parameters.MethodsData were obtained from 24 anti-HIV-negative subjects (controls) and 47 anti-HIV-positive patients classified according to clinical diagnosis, serum p24-antigen assay results, and antiretroviral treatment with zidovudine, using a modified flow cytometric assay for the detection of intracellular HIV p24 antigen (p24-FCA) in circulating CD4+ lymphocytes.ResultsThe proportion of CD4+ lymphocytes positive for p24-FCA correlated well with HIV infection (1.685 ± 1.902 versus 0.160 ± 0.152 in controls;PPPConclusionsOur results suggest that CD4+ p24 Ag-FCA is a rapid and easy test for the identification of the proportion of CD4+ lymphocytes with intracellular p24 Ag, and that it could be more appropriate than serum p24 Ag assay in evaluating disease progression and efficacy of antiretroviral treatment.

ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID

摘要:

ObjectiveTo evaluate the role of liver cells in and the effect of tumor necrosis factor-α (TNF-α) on HIV-1 replication.MethodsHuman hepatoblastoma HepG2 cells were infected with various strains of HIV-1 and the effect of TNF-α treatment, either before or after infection, was monitored by p24 antigen assays. Northern blot analysis and gel retardation assays were performed to determine the expression of CD4 and HIV-1trans-acting region (TAR)-binding proteins in these cells, respectively.ResultsHepG2 cells are CD4+ and support active HIV-1 replication, producing infectious virions, as measured by both p24 production and ability to infect T-cell lines with the virus produced by HepG2 cells. In contrast to the stimulatory effect of TNF-α on HIV-1 replication in T-cells and monocytes, up to 200 U/ml TNF-α treatment, at various times, either before or after HIV-1 infection, substantially inhibited p24 antigen production in HepG2 cells without causing any remarkable cytotoxicity. Gel-retardation assay revealed enhancement of a DNA-binding protein in TNF-α-treated HepG2 cells that binds to a specific sequence of the HIV-1 TAR, compared with the untreated control.ConclusionsThese results indicate the importance of cellular factor(s) in HIV-1 infection and suggest that cytokines in different tissues can induce opposite effects. TAR-binding protein may act as an inhibitory factor for HIV-1 replication in the HepG2 cell line.

ISSN:0269-9370    

出版商:OVID    

年代:1992

数据来源: OVID