Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A multicenter study was undertaken to determine the sensitivity and reproducibility of markers for human immunodeficiency virus type 1 (HIV-1) viral growth and the effect of various preparations of lymphocytes on the sensitivity of standard and routinely used procedures for HIV-1 isolation. In phase 1, cocultivated culture supernatants obtained from 10 HIV-1 cultures were transported to three Multicenter AIDS Cohort Study (MACS) Virology Laboratories. Three commercial HIV-p24 antigen capture (AC) tests and two reverse transcriptase (RT) assays were used to ascertain the replication of HIV-1. The Du Pont and Abbott AC assays were found to be most sensitive (85-100%), and the RT assay with 24-h incubation period had comparable sensitivity (75-100%). In phase II, the sensitivity of standard cocultivation procedure for HIV-1 isolation was compared using freshly phytohemagglutinin-P (PHA-P)-stimulated, stimulated-frozen, and frozen-thawed and then stimulated normal human peripheral blood mononuclear cells (PBMCs) as cocultivating cells. Blood samples from 13 HIV-1 infected individuals with various CD4+ cell counts were cocultivated in each of the three MACS laboratories using one of the aforementioned normal PBMCs. The PHA-P-stimulated fresh normal PBMC showed a maximum isolation rate of 100% (13 of 13) with an average of 8 days to positivity. This rate of isolation was significantly greater than other rates using any one of the other PBMC preparations. These findings demonstrated that the use of freshly PHA-P stimulated PBMCs maximized HIV-1 isolation from blood when a sensitive HIV-1 p24 AC assay or RT assay with overnight incubation is employed for the detection of HIV in culture supernatant.
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PMID:The effect of fresh lymphocytes on increased sensitivity of HIV-1 isolation: a multicenter study. 211 52

Human recombinant interferon-alpha (IFN alpha) restricted viral replication in human immunodeficiency virus- (HIV) infected T cells and monocytes. With T cells, reverse transcriptase (RT) activity in culture fluids was reduced threefold from that of control infected cells by IFN treatment, but HIV p24 antigen levels were unchanged. In contrast, levels of p24 antigen and RT activity in lysates of IFN-treated infected cells were threefold greater than those of controls. These differences suggest that the mechanism for IFN-induced antiviral effects in HIV-infected T cells resides in the terminal events (assembly and release) of the virus replication cycle. Monocytes treated with IFN at the time of virus challenge showed no p24 antigen or RT activity, no HIV-specific mRNA, and no proviral DNA in cells for up to 3 weeks after infection. IFN treatment of chronically infected monocytes also decreased virus replication, as assessed by p24 antigen, mRNA and RT detection assays. However, levels of proviral DNA in the IFN-treated and control HIV-infected cells were indistinguishable. The presence of large quantities of proviral DNA in cells with little or no evidence for active transcription documents a situation approaching true microbiological latency.
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PMID:Restriction of HIV replication in infected T cells and monocytes by interferon-alpha. 212 Nov 92

Kupffer cells (liver macrophages) represent the largest reservoir of fixed macrophages in the body. Accordingly, we have undertaken a study to evaluate their susceptibility to human immunodeficiency virus type 1 (HIV-1). Five-day-old primary cultures of Kupffer cells (KC) were infected with HIV-1, and as the infection progressed, syncytia appeared. Within the cells, viral proteins were detected by immunofluorescence using monoclonal antibodies directed against gp120 and p24. Electron microscopic examinations revealed the presence of typical Lentivirinae particles. The particles released from KC in the extracellular medium showed reverse transcriptase activity and p24 antigen; they could infect lymphocytic cells and were neutralized by a HIV+ patient's serum or an anti-gp120 monoclonal antibody. Our results thus demonstrate that the interaction of HIV-1 with KC in vitro leads to a productive infection. They suggest that the KC may be involved in the pathogenesis of HIV-1 infection and may (i) participate in the transmission of the infection to the peripheral blood cells, (ii) play a role in the depletion of uninfected CD4+ cells.
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PMID:Permissivity of primary cultures of human Kupffer cells for HIV-1. 212 Nov 93

Although knowledge has accumulated about GP110-CD4 interaction, viral penetration into human CD4+ lymphocytes remains unclear, in spite of the fact that all studies on HIV infection were performed on cell-transformed lineages, or on human polyclonal CD4+ cells. In order to investigate this viral entrance into susceptible cells, we studied the permissivity of 13 human monoclonal CD4+ lymphocytes by means of reverse transcriptase (RT) assay and immunocapture. We demonstrated a differential susceptibility to HIV of these CD4+ clones. In a second experiment, HIV infection was studied: (1) sequentially by RT assay and P24 immunocapture on several clones; (2) by cocultivation of infected clones with umbilical cord lymphocytes. These experiments suggested existence of permissive and "nonpermissive" CD4+ monoclonal lymphocytes. Slot blot, then PCR, revealed that proviral DNA sequences were detectable in all clones, but were present at lower levels in nonpermissive clones.
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PMID:Lack of permissivity of human monoclonal CD4+ lymphocytes to HIV. 212 40

Phase I and II clinical studies have been conducted to test the safety and potential activity of the reverse transcriptase inhibitor, dideoxycytidine (ddC), in treating human immunodeficiency virus-1-infected patients. Although ddC appears to be active in combating viral infection, as judged by its ability to decrease human immunodeficiency virus-1 p24 antigen titers and increase the number of CD4+ lymphocytes, it is also capable of causing severe peripheral neuropathy in a dose-dependent manner. The studies discussed here indicate that low-dose ddC treatment regimens substantially reduce the toxic side effects of this drug, and yet retain the ability to affect p24 antigen and CD4+ lymphocyte levels. These studies also define the window of therapeutic usefulness for ddC, and suggest that both safety and activity can be maintained during long-term, low-dose use of ddC.
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PMID:Safety and tolerance of dideoxycytidine as a single agent. Results of early-phase studies in patients with acquired immunodeficiency syndrome (AIDS) or advanced AIDS-related complex. Study Group of the AIDS Clinical Trials Group of the National Institute of Allergy and Infectious Diseases. 215 3

The emergence of human immunodeficiency virus resistant to 3'-azido-3'-deoxythymidine (zidovudine, AZT) in patients with acquired immunodeficiency syndrome (AIDS) or AIDS-related complex has been documented. Isolates from non-AZT-treated persons or those who had received AZT for less than six months showed a narrow range of susceptibility to the drug; on the other hand, isolates from those who had received AZT for six months or more consistently showed reduced susceptibility. Five highly AZT-resistant isolates were also insensitive to other compounds containing a 3'-azido group. No cross-resistance was found to other nucleoside analogues, including 2',3'-dideoxycytidine and 2',3'-dideoxyinosine. That cross-resistance occurred only in compounds containing a 3'-azido group suggests that mutations in the reverse transcriptase gene prohibit the enzyme from using nucleoside triphosphate containing a 3'-azido group. Progressive, stepwise increases in resistance have been associated with the sequential accumulation of specific amino acid changes in the reverse transcriptase gene. It is not yet known whether the resistant phenotype as determined in vitro results in clinical resistance to AZT. The gradual appearance of resistant isolates, the variable course of human immunodeficiency virus infections, and the absence of a consistent pattern of resurgent p24 antigen will make the emergence of AZT resistance difficult to correlate with clinical status or other markers. The combination of AZT with other drugs that do not share cross-resistance is a promising area for investigation to identify regimens that are more active and less likely to induce resistance.
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PMID:Susceptibility to nucleoside analogues of zidovudine-resistant isolates of human immunodeficiency virus. 218 29

High levels of unintegrated viral DNA accumulate during human immunodeficiency virus type 1 (HIV-1) infection of CEM T cells. Reinfection of already infected cells is required to attain these levels and reinfection also promotes the development of HIV-induced cytopathology. Rates of virus production, however, are independent of the accumulation of unintegrated viral DNA. Neutralizing antibody added soon after infection reduced viral DNA levels without appreciably affecting the production of cell-free viral p24 antigen or reverse transcriptase activity. Only 50 pM AZT were required to reduce the accumulation of unintegrated viral DNA by 50% in contrast to the 25 nM required to inhibit virus production by 50%. Cytopathology, as measured by number of syncytia in infected cell cultures, was correlated with highly elevated levels of unintegrated viral DNA. The minimal levels of unintegrated viral DNA present constitutively in the persistently infected HCEM cell line were consonant with the absence of cytopathic effects in these cells. These data demonstrate that inhibiting the reinfection of already infected cells modulates cytopathic HIV-1 infection to a form that is persistent and noncytopathic.
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PMID:Reinfection results in accumulation of unintegrated viral DNA in cytopathic and persistent human immunodeficiency virus type 1 infection of CEM cells. 221 39

Sensitivity of the cell-free human immundeficiency virus type 1 (HIV-1) and its producer cells was Studied in acidic media between pH 7.4 and 4.9 vitro. The cytopathic effect, reverse transcriptase activity and p24 antigen production by survived viruses were monitored in indicator cell cultures. It was established that, the cell-free HIV-1 particles are very sensitive to acidity. Between pH 7.4 and 6.0 they loose infectivity gradually, but this process is irreversible under pH 6.0 and subsequent neutralization cannot restore lost infectivity. However, viability, of virus producer cells is hardly affected between pH 7.4 and 4.9, but their ability to release infectious particles is lost gradually, similarly to the case of cell-free viruses. Neutralization of the media after treatment results in gradual restoration of releasing infectious viruses. These data explain that, cell-free HIV-1 looses infectivity in the acidic vagina or does on the skin, but infectivity is preserved in the blood, semen, rectum and breast milk being neutral or slightly alcalic. Virus carrier or producer lymphocytes by any route of infection can survive such protective mechanism of the body.
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PMID:[Different sensitivity to acid reaction of the AIDS virus and virus-producing cells: clinical conclusions]. 221 19

We have investigated the replicative capacity of 14 primary HIV-1 isolates in cultures of normal blood macrophages and PHA-stimulated peripheral blood mononuclear cells (PBMC). All viruses could infect normal macrophages as demonstrated by either reverse transcriptase (RT) activity, p24 antigen assay, or cocultivation with PBMC. One month after infection of macrophages virus could no longer be detected in the culture medium. The cells remained in this nonproductive state for another month. Virus could, however, be recovered by cocultivation with PHA-stimulated PBMC. Such macrophage-passaged virus induced pronounced cell killing with fragmentation and pyknosis and often replicated poorly in PBMC, in contrast to the original isolate. The results indicate that all primary HIV-1 isolates contain virus variants that can infect cells of the monocyte/macrophage lineage. Persistently infected, seemingly nonproducing cells, may serve as infectious reservoirs in the infected individual and spread infection to other susceptible cells over a long period of time. Moreover, the pronounced killing of PBMC by the macrophage-harbored virus may contribute to the deterioration of the immune system.
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PMID:HIV-1 infection of normal human macrophage cultures: implication for silent infection. 237 82

Human epithelial cells (L132) derived from embryonic lung and human lung fibroblasts (MRC5) were infected by human immunodeficiency virus type 1 (HIV-1) or type 2 (HIV-2). Surface CD4 protein was detected on these cells, and recombinant soluble CD4 (sCD4) blocked infection, indicating that HIV infection was mediated by the cell surface CD4 protein. In contrast, infection of human primary chondrocyte cells (C23), synovial cells (HSA), and foreskin fibroblasts (F13) was apparently independent of cell CD4-mediated mechanisms. Surface CD4 protein could not be detected on these cells, and sCD4 did not block the infection. F13 cells could be infected only by HIV-2, not by HIV-1, under our experimental conditions. In cells of mesenchymal orgin, viral production could be detected only after cocultivation with the human T-lymphoid H9 cells but not by conventional viral assays, including reverse transcriptase and p24 antigen assays in cell culture supernatant and immunofluorescence of host cells. Our DNA transfection studies indicated that this lack of detectable viral production was not due to the inefficient use of the HIV long terminal repeat or the Tat protein in these cells. These mesenchymal and epithelial cells were susceptible to HIV infection but differed in mechanism of virus entry compared with hematopoietic cells such as T lymphocytes. These observations may provide insights into clinical syndromes such as lung dysfunction in HIV-infected newborns and connective tissue disorders in HIV-infected adults.
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PMID:Infection of nonlymphoid cells by human immunodeficiency virus type 1 or type 2. 238 19


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