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)

Epidemiological results suggest that the etiological agent of the acquired immune deficiency syndrome (AIDS) is transmitted primarily through blood products, semen, and saliva. There is evidence that the human T-cell leukemia (lymphotropic) virus type III (HTLV-III) is this agent. HTLV-III has been isolated repeatedly from T cells obtained from peripheral blood or lymph node tissue of AIDS and pre-AIDS patients and of healthy people believed to have been exposed to the virus. In the present study, HTLV-III was detected in and isolated from T cells present in the seminal fluid of AIDS patients. Mononuclear cells from the semen of AIDS patients and normal individuals were cultured in the presence of T-cell growth factor (interleukin-2). After 6 to 8 days, HTLV-III antigens were transiently expressed by the cells from the AIDS patients but not by those from the normal individuals. When the mononuclear cells from the semen of AIDS patients were cocultured with a permissive human T-cell line, cell cultures were produced that expressed high levels of reverse transcriptase activity, showed retroviral particles by electron microscopy, and were positive for HTLV-III-specific antigens when tested by fixed-cell indirect immunofluorescence with the use of monoclonal antibodies to the p24 and p15 antigens of HTLV-III.
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PMID:HTLV-III in cells cultured from semen of two patients with AIDS. 620 7

We have developed a non-radioactive method to quantitate precisely levels of gene expression. This method is based on RT-PCR (reverse transcriptase-polymerase chain reaction) with an RNA competitor, followed by the covalent capture of the amplified DNA onto the wells of microtiter plates, and the quantitation of the PCR product by oligonucleotide hybridization and ELISA (enzyme-linked immunosorbent assay). The assay can reproducibly detect 1 zeptomole mRNA. The assay was successfully used to quantitate mRNA levels of the T cell derived cytokines interleukin-2, interleukin-4 and interferon-gamma in resting and stimulated human lymphocytes. Because it is performed in a microtiter ELISA format, this rapid, sensitive and non-radioactive method should facilitate measurements of gene expression, particularly in large clinical studies.
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PMID:Competitive RT-PCR ELISA: a rapid, sensitive and non-radioactive method to quantitate cytokine mRNA. 751 Jul 52

We previously identified an immunodominant CD4+ T cell determinant in the carboxy-terminal region of HIV-1 reverse transcriptase (RT528-543). The present study aimed at enumerating all the potential sites of HIV-1 RT recognized by Th cells in the BALB/c (H-2d) mouse model. To achieve this we used a panel of 62 overlapping 15-mer synthetic peptides covering the whole RT sequence to assay the following parameters: (i) immunogenicity in naive BALB/c mice injected either with peptides pools or individual peptides; (ii) antigenicity, as detected by their ability to restimulate in vitro T cells from BALB/c mice primed with native RT; (iii) MHC class II (Ad)-binding capacity as measured by the inhibition of the antigen-specific, Ad-restricted presentation of unfolded apamin (4-Acm) by fixed antigen-presenting cells to Ad/4-Acm-specific, interleukin-2-producing T hybridoma cells; and (iv) the presence of typical or degenerate consensus Ad-binding motifs. The results in this study permitted identification of three novel immunodominant RT mouse CD4+ T cell sites (RT276-290, RT375-389 and RT411-425) located in regions of limited polymorphism among RT from several HIV isolates. Some of these RT segments were found to be in the vicinity of B cell or H-2Kk- or HLA-A2-restricted cytotoxic T lymphocyte epitopes. Finally, the approach used in this study was found to be very efficient for enumerating most T cell recognition sites in a complex protein, a result that would have not been achieved by a single parameter-based analysis.
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PMID:Human immunodeficiency virus-1 reverse transcriptase immunodominant CD4+ T cell epitopes: a peptide-based multiparametric assessment in the mouse. 751 71

The outer surface lipoproteins of Borrelia burgdorferi, OspA and OspB, stimulate the production of nitric oxide (NO) by murine bone marrow-derived macrophages from BALB/c, C3H/HeN, and C3H/HeJ mice. Gamma interferon (IFN-gamma) caused a three- to fivefold enhancement of this production of NO, and the L-arginine analog N-guanidino-monomethyl L-arginine inhibited it. Activation of transcription of the inducible NO synthase gene in stimulated macrophages was demonstrated by reverse transcriptase rapid PCR. Although IFN-gamma increased the amount of NO produced in macrophage cultures, it did not cause transcription of the inducible NO synthase gene greater than that seen with the Borrelia proteins. OspA and OspB also induced the production of high levels (40 to 150 ng/ml) of IFN-gamma in cultures of macrophages incubated with interleukin-2 (IL-2)-elicited cells from normal (T and NK cells) and scid (NK cells) mice but not in macrophages or IL-2-elicited cells cultured individually. This suggests that OspA stimulated macrophage production of cytokines, which, in turn, stimulated the production of IFN-gamma by NK and T cells. Reverse transcriptase rapid PCR demonstrated that OspA and sonicated B. burgdorferi stimulated production of several inflammatory cytokines in macrophage cultures, including IL-1, IL-6, IL-12, IFN-beta, and tumor necrosis factor alpha. As tumor necrosis factor alpha, IFN-beta, and IL-12 are potent activators of IFN-gamma production by T and NK cells, their presence in these cocultures could be responsible for the IFN-gamma production. Lymphocytes from infected C3H mice also produced IFN-gamma when stimulated with B. burgdorferi; thus, immune cells may also modulate NO responses. The generation of NO during infection with B. burgdorferi may be important, as NO has potent antimicrobial properties. NO can also be involved in pathological inflammatory processes in which its generation is detrimental to the host. Thus, the colocalization of B. burgdorferi lipoproteins, NO-producing cells, and regulatory cytokines may determine the outcome of infection.
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PMID:Outer surface lipoproteins of Borrelia burgdorferi stimulate nitric oxide production by the cytokine-inducible pathway. 752 Apr 17

Immunosuppressive drugs used in clinical transplantation block cytokine mRNA transcription in vitro, but clinical rejection episodes are common. An understanding of what cytokine message is transcribed would be helpful in determining what contributes to the success of immunosuppression and provide directions for further research aimed at targeting specific cytokines. Previous studies have examined cytokine mRNA in rejecting solid organ biopsies by the reverse transcriptase polymerase chain reaction (RT-PCR) with variable results. We used nonradioactive in situ hybridization with cytokine-specific riboprobes to determine the frequency of cells expressing cytokine mRNA in the allograft infiltrate. Kidney biopsies were obtained from patients receiving protocol biopsies and with clinical evidence of rejection. Fourteen biopsies with a pathologic diagnosis of rejection were studied. Eight showed no cytokine staining, 2 expressed IL-2, and 3 expressed IL-4 and IFN-gamma. The positive cells were present at a low frequency (mean 2, range 1-5 per 10 high-power fields). The proportion of kidney biopsies expressing detectable message for interleukin-2 (IL-2), interleukin-4 (IL-4), and interferon-gamma (IFN-gamma) by in situ hybridization were similar to those reported using RT-PCR. The novel finding is that these cytokines are expressed in a few strongly positive cells in the allograft infiltrate. The vast majority of infiltrating cells are negative. This suggests that either the biopsies were performed when cytokine message was not expressed at a high level or that in human allograft recipients the sustained expression of the cytokines IL-2, IL-4, and IFN-gamma may not be necessary for graft rejection.
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PMID:Low frequency of infiltrating cells intensely expressing T cell cytokine mRNA in human renal allograft rejection. 753 48

Cytotoxic T lymphocytes (CTL) specific for human immunodeficiency virus type 1 (HIV-1) are thought to play an important role in controlling HIV-1 infection. HIV-1-specific CTL are readily demonstrated in unstimulated peripheral blood mononuclear cells (PBMC) of HIV-infected adults but less frequently in PBMC from vertically infected children. HIV-1-specific CTL lines were derived from a long-term survivor of vertical HIV-1 infection using PBMC stimulated with a CD3-specific monoclonal antibody and interleukin-2; these lines had Gag- or reverse transcriptase (RT)-specific cytotoxicity. Cytotoxicity was restricted by major histocompatibility complex class I antigen and blocked by antibody to the T cell receptor complex. Fluorescence-activated cell sorting analysis demonstrated their phenotype to be CD3+CD4-CD8+. Unstimulated PBMC from this patient had no detectable HIV-1-specific cytotoxicity when tested against autologous HIV-1 envelope-, Gag-, or RT-expressing target cells. Thus, this child with vertically acquired HIV-1 infection likely has HIV-1-specific CTL precursors despite the absence of circulating, activated HIV-1-specific CTL.
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PMID:Cytotoxic T lymphocyte lines specific for human immunodeficiency virus type 1 Gag and reverse transcriptase derived from a vertically infected child. 768 62

Feline immunodeficiency virus (FIV) infection in the cat is similar to human immunodeficiency virus type 1 infection in causing a selective reduction in CD4+ cell numbers, leading to inversion of the CD4+/CD8+ ratio. To determine whether FIV, similar to human immunodeficiency virus type 1, has a tropism for CD4+ cells, we examined the in vitro and in vivo susceptibilities of feline lymphocyte subpopulations to FIV infection. Infection of interleukin-2-dependent CD4+ or CD8+ lymphocyte cultures with the NCSU1 isolate of FIV (FIV-NCSU1) resulted in syncytium formation, cell death, and Mg(2+)-dependent reverse transcriptase (RT) activity in both cases. Monoclonal antibodies to feline lymphocyte subsets were used to sort peripheral blood mononuclear cells from FIV-infected cats into highly (> 95%) purified CD4+ cell, CD8+ cell, immunoglobulin-positive (Ig+) cell, and monocyte subpopulations. The mononuclear cell subpopulations were analyzed for FIV provirus by polymerase chain reaction and Southern blot analysis and for virus expression by RT activity. All 16 cats infected with FIV-NCSU1 demonstrated FIV provirus in CD4+ cell-, CD8+ cell-, and Ig+ cell-enriched lymphocyte populations. Southern blot detection of amplified gag gene sequences and limiting-cell-dilution polymerase chain reaction analysis indicated that Ig+ cells carried a higher FIV provirus burden in chronically (> or = 1-year) infected cats than either CD4+ or CD8+ cells. In contrast, CD4+ cells carried the greatest provirus burden in acutely (2- to 4-week) infected cats. FIV provirus was detected in monocytes from only 1 of 10 cats with asymptomatic infection. Addition of culture supernatants from enriched CD4+, CD8+, and Ig+ cells from FIV-infected cats to an FIV-susceptible CD4+ lymphocyte culture resulted in syncytium formation, cell death, and RT activity. Infection of Ig+ cells is not unique to FIV-NCSU1, as lymphocyte subpopulations from other cats with natural infections and cats infected with the Petaluma or Mount Airy isolate of FIV demonstrated a similar distribution of FIV provirus and RT activity. These data suggest that FIV possesses a broad tropism for peripheral blood mononuclear cells and that an Ig+ cell may serve as a major reservoir for the virus in chronically infected cats.
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PMID:In vivo lymphocyte tropism of feline immunodeficiency virus. 768 19

Two new myeloid cell lines (K051 and K052) were established from a patient with multilineage CD7-positive acute leukemia. The K051 and K052 were established from the patient's bone marrow cells at diagnosis and at relapse, respectively. The K051 cell expressed myeloid-associated antigens (CD13 and CD33), a platelet-associated antigen (CD41), and an erythroid antigen (glycophorin A). The K052 cell expressed myeloid-associated antigens (CD13, CD14, and CD33), lymphoid markers (CD2, CD5, and CD7), and HLA-DR. Chromosome analysis of both cell lines showed a 17p- chromosome. Both cell lines were investigated for aberrations of the p53 gene and the N-ras gene. A p53 mutation detected in both cell lines consisted of a C-->T substitution in codon 248. An N-ras mutation detected only in the K052 cell consisted of a G-->C substitution in codon 13. Expression of the multidrug resistance gene (MDR1) was also investigated by the semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR). MDR1-mRNA was more highly expressed by the K052 cell than the K051 cell, being equivalent to that in HEL cells. The functional MDR1-protein against vincristine was also observed, and its function was inhibited by verapamile and Cyclosporin A. The K052 cells were capable of phenotypic or morphologic differentiation after being incubated with granulocyte colony-stimulating factor, interleukin-2, phorbol 12-myristate 13-acetate, or 1,25-dihydroxy-vitamin D3. In contrast, the K051 cells responded phenotypically to retinoic acid. Thus, the K051 and K052 cell lines will be useful for investigating the cellular and molecular events in leukemogenesis and differentiation, and the mechanism of expression of the MDR1 gene.
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PMID:p53 and N-ras mutations in two new leukemia cell lines established from a patient with multilineage CD7-positive acute leukemia. 769 50

An involvement of cellular immunity in alveolar echinococcosis is strongly suggested by the intense granulomatous infiltrations observed around the hepatic parasite lesions. However, the basis of cellular immunoregulation in patient with alveolar echinococcosis is poorly understood. The present report shows a comparative analysis of lymphoid cell function in peripheral blood mononuclear cells (PBMC) of 16 patients with alveolar echinococcosis and of healthy individuals. Our in vitro restimulation studies with crude Echinococcus multilocularis antigen demonstrated that PBMC from patients with alveolar echinococcosis were responsive to challenge with parasitic antigen as measured by lymphoid cell proliferation. In this system, we also evaluated cytokine expression at the gene and protein levels after stimulation with E. multilocularis antigen. Analysis of cytokine mRNA expression revealed distinct patterns of cytokine expression in patients and normal donors. By using reverse transcriptase PCR, we could demonstrate that the TH1 cytokine transcripts interleukin-2 (IL-2) and gamma interferon (IFN-gamma) are present in PBMC from patients with alveolar echinococcosis. Moreover, it was found that stimulation with E. multilocularis antigen induced or enhanced the expression of the TH2 cytokine IL-3, IL-4, IL-10, and especially IL-5 mRNAs in PBMC from 13 of 16 patients with alveolar echinococcosis. Two patients who were examined after radical surgery, as well as another patient with a stable course of the disease under continuous chemotherapy, were not able to generate the same pattern of cytokine response and had no evidence of IL-5 mRNA synthesis. In contrast to the frequent expression of TH2 cytokine mRNAs observed in patients with alveolar echinococcosis, PBMC cultures from normal donors showed prominent IL-2 and IFN-gamma mRNA expression but weak IL-3, IL-4, and IL-10 mRNA expression. Most interestingly, IL-5 mRNA was substantially absent in PBMC from healthy individuals. In accordance with the mRNA studies, it was found that E. multilocularis antigen induced the secretion of large amounts of IL-5 and intermediate amounts of IFN-gamma in patients with alveolar echinococcosis, whereas large amounts of IFN-gamma and no or threshold amounts of IL-5 were detected in supernatants from healthy individuals. Collectively, the present study provides the first evidence that a TH2 immune response is gradually activated during the course of E. multilocularis infection, indicating a critical role for IL-5 in the manifestation of human alveolar echinococcosis.
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PMID:Interleukin-5 is the predominant cytokine produced by peripheral blood mononuclear cells in alveolar echinococcosis. 772 73

Twenty-one cytotoxic T lymphocyte (CTL) clones or lines that killed autologous tumor cells, but not allogeneic tumor, K562, or Daudi cells, were established from fresh tumor-infiltrating lymphocytes of two individuals (HP-1 and HP-2) with head and neck cancer by limiting dilution in the presence of recombinant interleukin-2. Sixteen (76%) of these 21 clones or lines comprised CD4+ CTLs and the other five comprised CD8+ CTLs. These observations suggest that autologous tumor cell-specific CD4+ CD8- and CD4- CD8+ CTLs are present in vivo at the tumor site in head and neck cancer. Analysis of T cell receptor (TCR) gene arrangements in 20 of the 21 CTL isolates with reverse transcriptase and the polymerase chain reaction revealed that five of 12 and five of eight isolates from HP-1 and HP-2, respectively, were clones, the other isolates being lines comprised of two or more clones. Each CTL clone showed a different combination of V alpha and V beta gene expression, suggesting that more than five different tumor-associated antigens may be expressed on head and neck cancer cells. In spite of the diversity of TCR alpha beta combinations, TCR V alpha 1, V alpha 3, V alpha 8, V alpha 10, V beta 8, V beta 9, and V beta 17 were also frequently expressed in both patients. These data suggest that specific CTLs proliferate oligoclonally and contribute to the specific immune response against head and neck cancer in vivo.
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PMID:Functional and T cell receptor gene usage analysis of cytotoxic T lymphocytes in fresh tumor-infiltrating lymphocytes from human head and neck cancer. 779 Mar 20


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