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)

The Fas/FasL system mediates apoptosis in several different cell types, including T lymphocytes. Fas ligand (FasL), a 40-kD type II membrane protein also expressed in activated T cells, belongs to the tumor necrosis factor ligand family. We describe a new alternative splicing of mouse FasL, named FasL short (FasLs), cloned by reverse transcriptase-polymerase chain reaction. FasLs is encoded by part of exon 1 and part of exon 4 of FasL gene. The protein encoded by FasLs mRNA has a putative initiation code at position 756 and preserves the same reading frame as FasL, resulting in a short molecule lacking the intracellular, the transmembrane, and part of the extracellular domains. RNase protection and immunoprecipitation analysis showed that FasLs is expressed in nonactivated normal spleen cells and in hybridoma T cells and that it is upregulated upon activation by anti-CD3 monoclonal antibody (MoAb). Moreover, FasLs-transfected cells expressed soluble FasLs in the supernatant and became resistant to apoptosis induced by agonist anti-Fas MoAb. Thus, FasLs, a new alternative splicing of FasL, is involved in the regulation of Fas/FasL-mediated cell death.
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PMID:Cloning and expression of a short Fas ligand: A new alternatively spliced product of the mouse Fas ligand gene. 1055 56

Immunologic injury to heart allografts is an initial and essential event in the pathogenesis of graft coronary artery disease (GAD). A variety of cytokines expressed in heart allografts modify both acute rejection and chronic inflammation, and could contribute to the development of GAD. The present study investigated the gene expression of interleukin (IL)-1beta, IL-2, IL-4, IL-6, IL-10, tumor necrosis factor (TNF)-alpha, interferon (IFN)-gamma, and Fas ligand in chronically rejecting DBA/2-to-B 10.D2 mouse heart allografts at defined intervals of 7, 14, 28, or 70 days after transplantation by semiquantitative reverse transcriptase-polymerase chain reaction. GAD developed gradually, showing the highest value for mean intima/media ratio at day 70. Fas ligand, and the Th1 cytokines IL-2 and IFN-gamma, were vigorously induced in allografts at day 7, when histology showed pronounced parenchymal rejection, and rapidly decreased by day 28. However, the level of mRNA expression of Th2 cytokines, IL-6 and IL-10, and other inflammatory cytokines, TNF-alpha and IL-1beta, were still elevated on day 28. The persistent expression of specific cytokines suggests an important role in chronic inflammation. Thus, a persistently high level expression of inflammatory cytokines could be associated with chronic inflammation in the allografts, which promotes the development of GAD.
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PMID:Cytokine gene expression during the development of graft coronary artery disease in mice. 1055 20

The functional properties, regarding parasite growth inhibition in vitro, the cytotoxic potential and cytokine profiles of human gammadelta+ and alphabeta+ T cells, T-cell lines and clones stimulated with Plasmodium falciparum-antigen-or T-cell mitogen in vitro were investigated. Using reverse transcriptase-polymerase chain reaction (RT-PCR) and specific primers, mRNA for the cytolytic molecules perforin, granzyme A and B, Fas and Fas ligand (FasL) were detected in both the gammadelta- and the alphabetaT cells. Despite this fact, only gammadeltaT cells inhibited, both Vdelta1+ and Vdelta2+, the in vitro growth of the asexual blood stages in a dose dependent manner. The inhibition required cell-to-cell contact and was not observed until the second parasite replication implied that the likely gammadeltaT-cell target was the extracellular merozoite or schizont. The failure of alphabetaT cells to inhibit the growth of the parasite suggests requirement of additional cytolytic molecules/signals or different receptor specificities exhibited by the gammadeltaT cells. Both the gammadelta- and alphabetaT cells expressed mRNA for a large number of cytokines. Interferon (IFN)-gamma, interleukin (IL) IL-5, IL-6, IL-8, tumour necrosis factor alpha (TNFalpha), tumour necrosis factor beta (TNF-beta)/lymphotoxin (LT) and T-cell growth factor beta-1 (TGF-beta1) were observed in all activated clones tested. No IL-3 was detected, while IL-1beta, IL-2, IL-4, IL-10 and GM-CSF were variably expressed. In conclusion, our data show that gammadeltaT cells in malaria nonimmune individuals inhibit the asexual blood stages of P. falciparum malaria, while similarly activated alphabetaT cells do not. Thus, it is likely that the gammadeltaT cells could play a mandatory role in the elimination of parasites and/or the regulation of the early immune response to malaria infection.
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PMID:Human gamma delta T cells that inhibit the in vitro growth of the asexual blood stages of the Plasmodium falciparum parasite express cytolytic and proinflammatory molecules. 1060 13

Dysregulation of apoptosis, particularly in the Fas/Fas ligand (FasL) pathway, is considered to be involved in the pathogenesis of autoimmune diseases such as systemic lupus erythematosus (SLE). Recently, a soluble decoy receptor, termed decoy receptor 3 (DcR3), that binds FasL and inhibits FasL-induced apoptosis, has been identified. Silicosis is clinically characterized not only by respiratory disorders but by immunological abnormalities. We have found that serum soluble Fas (sFas) levels are elevated in silicosis patients and that sFas message is dominantly expressed in PBMC derived from these patients. This study examined DcR3 gene expression in PBMC derived from patients with silicosis, SLE, or progressive systemic sclerosis (PSS), and compared it with that in healthy volunteers (HV). The relative expression level of the DcR3 gene was examined in PBMC derived from 37 patients with silicosis without clinical symptoms of autoimmune disease, nine patients with SLE, 12 patients with PSS, and 28 HV using the semiquantitative multiplex-reverse transcriptase-polymerase chain reaction (MP-RT-PCR). The correlation between the relative expression level of the DcR3 gene and multiple clinical parameters for respiratory disorders and immunological abnormalities in individuals with silicosis was analysed. The DcR3 gene was significantly over-expressed in cases of silicosis or SLE when compared with HV. In addition, the DcR3 relative expression level was positively correlated with the serum sFas level in silicosis patients. It is unclear, however, whether over-expression of the DcR3 gene in silicosis is caused by chronic silica exposure, merely accompanies the alteration in Fas-related molecules, or precedes the clinical onset of autoimmune abnormalities. It will be necessary to study these patients further, establish an in vitro model of human T cells exposed recurrently to silica compounds, and resolve whether the increase in DcR3 mRNA expression is a cause or consequence of disease.
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PMID:Over-expression of the decoy receptor 3 (DcR3) gene in peripheral blood mononuclear cells (PBMC) derived from silicosis patients. 1063 70

We investigated the cytotoxicity mechanisms of alloantigen-specific human CD4(+) and CD8(+) cytotoxic T lymphocytes (CTLs) using cells from family members with the Fas gene mutation. Alloantigen-specific CD4(+) and CD8(+) CTL bulk lines and clones were generated from 2 individuals by stimulation of their peripheral blood lymphocytes with allogeneic Fas(-/-) or Fas(+/-) cell lines that were established from B-lymphocytes of a patient with Fas deficiency and her mother, respectively. Both CD4(+) and CD8(+) CTL bulk lines and clones directed against allogeneic HLA antigens exerted cytotoxicity against Fas(-/-) and Fas(+/-) cells to almost the same degree. The cytotoxicity of CD4(+) and CD8(+) CTLs appeared to be Ca(2+)-dependent and was completely inhibited by concanamycin A, an inhibitor of perforin-mediated cytotoxicity. Messenger RNAs for the major mediators of CTL cytotoxicity, Fas ligand, perforin, and granzyme B were all detected in these CD4(+) CTLs with the use of the reverse transcriptase polymerase chain reaction. The majority of CD4(+) CTL clones that showed Fas-independent cytotoxicity were T(H)0, as determined by their cytokine production profile. These data, obtained with the use of a novel experimental system, clearly show that the main pathway of cytotoxicity mediated by alloantigen-specific human CD4(+) as well as by CD8(+) CTLs is granule exocytosis, and not the Fas/Fas ligand system.
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PMID:Granule exocytosis, and not the fas/fas ligand system, is the main pathway of cytotoxicity mediated by alloantigen-specific CD4(+) as well as CD8(+) cytotoxic T lymphocytes in humans. 1073 6

Fas receptor is a member of a superfamily of receptors characterized by cysteine-rich motifs in the extracellular domain of the molecule. Binding of Fas ligand to the receptor leads to receptor activation and the induction of intracellular signals that result in apoptotic cell death. In the present study, the expression of mRNA and proteins of Fas receptor and Fas ligand were examined in human submandibular gland ductal (HSG) cells treated with okadaic acid by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunoblot analysis. Six hundred and eighty-two bp of the PCR product of Fas receptor mRNA was detected in HSG cells and a protein with an estimated molecular weight of 58,000 was expressed in HSG cells. Treatment of HSG cells with an agonistic anti-Fas monoclonal antibody resulted in death of HSG cells, indicating that the functional Fas receptor protein is expressed in HSG cells. Fas receptor protein expression stimulated by okadaic acid was elevated in a dose- and time-dependent manner, with maximal expression at 20 nM and 48 h treatment. Fas ligand mRNA was also detected constitutively in HSG cells by RT-PCR. Okadaic acid stimulated the expression of Fas ligand protein in HSG cells in a time-dependent manner, while the expression of the ligand was low in untreated HSG cells. The molecular weight of Fas ligand was estimated as 68,000. An antagonistic anti-Fas ligand monoclonal antibody prevented okadaic acid-induced death in HSG cells in a dose-dependent fashion as determined by WST-1 assay. The results indicate that the expression of Fas receptor and ligand is regulated by protein phosphatase(s) sensitive to okadaic acid and is involved in okadaic acid-induced apoptosis in HSG cells. The results also suggest that the Fas receptor-ligand system might regulate apoptosis in HSG cells.
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PMID:Upregulation of the expression of Fas antigen and Fas ligand in a human submandibular gland ductal cell line by okadaic acid. 1086 77

In the present study, we demonstrated that a close relationship exists between hepatitis C virus (HCV) infection of peripheral blood mononuclear cells (PBMCs) and cell-surface Fas expression in patients with hepatitis C, and showed the possibility of PBMCs apoptosis via a Fas-mediated system. The expression of Fas on PBMCs was found by flowcytometric analysis to be significantly increased in these patients. In addition, the treatment of patients' PBMCs with anti-Fas antibody induced cell death, with nuclear condensation and fragmentation and cellular DNA fragmentation. These data indicate that the patients' PBMCs expressed a large amount of functional Fas on the cell surface and were susceptible to stimulation against Fas, causing apoptotic cell death. We then quantified the serum-soluble Fas ligand (sFasL), which was known to bind to Fas and induce the apoptotic signals into the sensitized cells. The patients' serum sFasL levels were significantly higher than those of normal subjects and showed a good negative correlation with their PBMC number. To demonstrate the correlation between Fas expression and HCV infection, nested reverse transcriptase polymerase chain reaction (RT-PCR) was performed to detect HCV RNA. Interestingly, HCV RNA was preferentially detected from Fas-positive cells but not from Fas-negative cells, which had been isolated from PBMCs by magnetic beads. These results suggest that HCV infection of PBMCs might induce Fas expression and additional stimulation such as sFasL might induce apoptosis in these Fas-expressing cells. These mechanisms, in addition to hypersplenism, may explain the decrease in the number of PBMCs observed in patients with chronic hepatitis C.
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PMID:Fas-mediated apoptosis of peripheral blood mononuclear cells in patients with hepatitis C. 1093 Sep 83

Activated cytolytic effector cells like lymphokine-activated killer (LAK) and the recently described bacillus-Calmette-Guerin-activated killer (BAK) cells are thought to mediate antitumor effects against metastatic renal cell carcinoma (RCC) and superficial bladder cancer respectively. Perforin and Fas ligand (FasL) have been described as the major lytic principles in cellular cytotoxicity. Using a radioactive-release assay and specific inhibitors, we investigated the molecular mechanisms used by LAK and BAK cells in the lysis of renal carcinoma cells. In addition, we evaluated the susceptibility of RCC cells to FasL-mediated cytotoxicity. LAK and BAK cells effectively lysed the renal cancer cell line SK-RC-35 upon cell-cell contact. Both effector cell populations were shown to produce perforin and FasL as determined by reverse transcriptase/polymerase chain reaction (RT-PCR). Using fluorescence-activated cell sorting analyses and RT-PCR, we detected a marked Fas receptor (Fas, CD95) expression on RCC cells. However, RCC cells were shown to be resistant to killing by recombinant FasL and lysis by BAK and LAK cells was not inhibited in the presence of anti-FasL antibody. In contrast, the cytotoxicity exerted by LAK and BAK cells was drastically reduced in the presence of the Ca2+-chelating agent EGTA as well as concanamycin A, a specific inhibitor of perforin-mediated lysis. These results demonstrate that cytolysis of FasL-resistant RCC cells by activated immune cells is mediated via perforin. Our findings give further insights into the molecular mechanisms involved in the elimination of RCC by cytotoxic lymphocytes activated with biological response modifiers.
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PMID:Killing of Fas ligand-resistant renal carcioma cells by interleukin-2- and BCG-activated effector cells. 1099 63

The present study investigates the molecular apoptotic pathway in germ cells following acute ischemia of the rat testis. Rats were subjected to ischemia-inducing torsion and testes were harvested after reperfusion. Apoptotic cells were identified with an antibody to single-stranded DNA. Seminiferous tubule RNA was examined by RNase protection assay or by reverse transcriptase-polymerase chain reaction (RT-PCR) for the presence and regulation of apoptotic molecules. Proteins from seminiferous tubules were used for Western blot analysis of cytochrome c. Germ cell apoptosis was maximal at 24 h after repair of torsion. Germ cells in stages II-III of the seminiferous epithelium cycle were the predominant early responders. The RNase protection assays revealed that Bcl-X(L) was the prominent mRNA species. Caspases 1, 2, 3, and Bax mRNA were consistently upregulated; however, the time of upregulation after torsion was variable. The Bcl-X(L) and Bcl-X(S) mRNAs were less consistently upregulated and there was no evidence for upregulation of Fas or Bcl-2. Fas ligand (FasL) was not detected by RNase protection assay, but RT-PCR revealed a significant increase in FasL expression 4 h after the repair of torsion. Western blot analysis for cytochrome c release demonstrated a significant increase 4 h after the repair of torsion. Results suggest that germ cell apoptosis following ischemia/reperfusion of the rat testis is initiated through the mitochondria-associated molecule Bax as well as Fas-FasL interactions.
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PMID:Molecular pathway of germ cell apoptosis following ischemia/reperfusion of the rat testis. 1105 53

Ovarian follicular atresia occurs by apoptosis of granulosa and theca cells. The Fas antigen (Fas), a cell surface receptor that triggers apoptosis when activated by Fas ligand (FasL), may be involved in this process. A possible role of the Fas pathway in mediating serum withdrawal-induced apoptosis of granulosa cells was examined. Granulosa cells collected from 5- to 10-mm bovine follicles were cultured in DMEM-F12 containing serum for 3 days, deprived of serum, and live cells were counted at various times after serum withdrawal. Cell death increased significantly 6 h after serum withdrawal (21% +/- 7%; P: < 0.05 vs. 0 h) and continued to increase until 24 h (43% +/- 6%). No further increases in cell death were observed through 72 h. Detection of the translocation of phosphatidylserine to the outer surface of the cell membrane by annexin V binding indicated that cells died by apoptosis. Quantitative reverse transcriptase-polymerase chain reaction assays showed no changes in Fas mRNA levels but a 4.7-fold increase in FasL mRNA 3 h after serum withdrawal (P: < 0.05 vs. 0 h). FasL mRNA remained elevated through 24 h and returned to basal levels at 48 h. Immunohistochemical staining showed that both Fas and FasL protein increased on the cell surface within 3 h and remained elevated through 12 h (the last time point tested). Binding of FasL to Fas was blocked with two reagents that bind to the extracellular domain of FasL: an anti-FasL antibody and Fas:Fc, a chimeric protein consisting of the Fc portion of human immunoglobulin G and the extracellular domain of human Fas. Cell death 24 h after serum withdrawal was reduced 55% +/- 10% and 34% +/- 12% by anti-FasL antibody and Fas:Fc, respectively (P: < 0.05 vs. no blocking protein). In conclusion, serum withdrawal-induced apoptosis of bovine granulosa cells is mediated at least partially by Fas/FasL interactions. These results are consistent with a potential role of Fas in an autocrine or paracrine pathway to trigger ovarian follicular atresia.
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PMID:Apoptosis of bovine granulosa cells after serum withdrawal is mediated by Fas antigen (CD95) and Fas ligand. 1115 54


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