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)

An altered peptide ligand (analog) of the encephalitogenic epitope of proteolipid protein residues 139-151 (p139-151) in which residues 144 and 147 are substituted with leucine and arginine, respectively (LR), protects from clinical but not histological experimental allergic encephalomyelitis (EAE). To understand in situ events associated with this protection, T cells from brains of mice immunized with either native p139-151, the analog LR or a combination of the two were isolated and characterized. High proportions of cells from co-immunized mice (38%) and LR-immunized mice (58%) reacted to both p139-151 and LR, whereas fewer cells from p139-151 immunized mice (7%) were cross-reactive. T cell clones derived from brains of LR- and co-immunized mice were also cross-reactive in vitro. By reverse transcriptase-based polymerase chain reaction, higher levels of TGF-beta mRNA, and lower levels of TNF-alpha and IFN-gamma mRNA were found in the central nervous system (CNS) tissue of LR and co-immunized mice. Immunohistochemistry demonstrated greater TGF-beta immunoreactivity in CNS inflammatory foci in co-immunized and LR-immunized mice. There were no significant differences in CD4+ or CD8+ cell infiltrates among the groups and differences in other cytokines were not identified by immunocytochemistry. Protection from clinical EAE in LR and co-immunized mice was partially abolished by anti-TGF-beta antibody treatment. Thus, protection from clinical disease following immunization with the analog LR is associated with infiltration into the CNS of a T cell population that could potentially recognize the native PLP peptide and with enhanced TGF-beta production by cells within CNS inflammatory foci.
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PMID:Altered peptide ligand modulation of experimental allergic encephalomyelitis: immune responses within the CNS. 952

High concentrations of oxygen, indispensable for the treatment of severe hypoxemia from neonatal as well as adult respiratory distress syndrome, increase the risk of oxygen toxicity. Biochemical mechanisms are lipid peroxidation, protein sulfhydryl oxidation, enzyme inactivation, and DNA damage. Recent reports suggest that cytokines might be involved in free radical injury as well as in adaptive response to hyperoxic injury. However, actual signal transduction pathways involving cytokines have not yet been clarified. In this study we exposed cultured human umbilical vein endothelial cells (HUVECs) to either ambient air or 100% oxygen, and compared for the rate of DNA synthesis ([3H]thymidine uptake) at different time points up to 72 h. After exposing the cells to each treatment condition, we extracted RNA, constructed complementary DNA using reverse transcriptase, amplified the specific DNA segments of cytokines by polymerase chain reaction (PCR), and used the PCR products for gel electrophoresis to examine the bands which signified mRNA levels of corresponding cytokines. There was a significant decrease in the rate of DNA synthesis as early as 24 h. The mRNA expression of IL-1 beta and TNFa seemed less influenced by hyperoxia, while IL-8 and TGF beta showed marked increase in mRNA levels at 6 h of 100% oxygen exposure.
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PMID:Hyperoxia influences mRNA expression of cytokines in cultured human umbilical vein endothelial cells. 952 79

A common feature of some parasitic infections and allergic and atopic skin diseases is the involvement of Th2 lymphocytes and the dermal appearance of eosinophils (Eos). Because Th2 lymphocytes apparently do not release Eo attractants, we addressed the question of whether the Th2 cytokine IL-4 induces its production in dermal fibroblasts. We therefore stimulated fibroblasts with IL-4. HPLC investigation of supernatants revealed a single Eo chemotactic protein, which was purified to homogeneity giving a single 13-kDa band upon SDS-PAGE analyses. Peptide mapping with subsequent amino acid sequencing revealed an Eo-selective chemotaxin, which consists of a mixture of N-terminally truncated and O-glycosylated forms of the chemokine eotaxin. Other chemokines such as RANTES, MCP-3, MCP-4, or MIP-1alpha were not detected as Eo chemotaxins under these conditions. Using reverse transcriptase-PCR techniques, we found that IL-4 dose and time dependently induces eotaxin mRNA in dermal fibroblasts. Stimulation with IL-4 and TNF-alpha caused a 10- to 20-fold increase of the release of three biochemically different eotaxin forms, each consisting of a mixture of N-terminally truncated and O-glycosylated variants having the same backbone amino acid sequence but different specific activities. Our findings support the hypothesis that eosinophil recruitment seen in IL-4-mediated skin reactions, at least in part, may be due to Th2 cytokine-mediated induction of eotaxin in dermal fibroblasts.
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PMID:IL-4 induces eotaxin: a possible mechanism of selective eosinophil recruitment in helminth infection and atopy. 955 56

The aim of the present study was to further characterize the role of alveolar macrophages (AM) in acute human lung inflammation by evaluating their capacity to produce pro-inflammatory cytokines such as tumour necrosis factor (TNF)-alpha, interleukin (IL)-6 and IL-8. Patients with severe community-acquired pneumonia (CAP; n=12) and healthy volunteers (n=10) underwent bronchoalveolar lavage (BAL). AM were separated to high purity (>96%) using fluorescence-activated cell sorting. We determined the TNF-alpha, IL-6 and IL-8 cytokine gene expression in AM ex vivo using semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR). Moreover, we measured in vitro unstimulated, lipopolysaccharide (LPS)- and LPS/interferon-gamma inducible TNF-alpha, IL-6 and IL-8 cytokine release and evaluated samples of BAL fluids for the same pro-inflammatory cytokines using an enzyme-linked immunosorbent assay (ELISA). We found increased TNF-alpha, IL-6 and IL-8 messenger ribonucleic acid (mRNA) levels in AM from CAP patients that were significantly elevated only for IL-8. When challenged with endotoxin in vitro, AM obtained from CAP patients showed a strongly reduced potential to release TNF-alpha and IL-6 compared to healthy controls, whereas IL-8 secretion did not differ significantly between groups. Moreover, stimulation of AM from CAP patients with LPS plus IFN-gamma augmented TNF-alpha and IL-6 cytokine release to near normal levels. Interestingly, no TNF-alpha protein was measured in BAL samples from CAP patients, whereas IL-6 and IL-8 protein levels were found to be significantly increased. Together, highly purified alveolar macrophages from community-acquired pneumonia patients show relatively low ex vivo tumour necrosis factor-alpha and interleukin-6 but not interleukin-8 messenger ribonucleic acid levels that are associated with a decreased pro-inflammatory cytokine release in vitro which, however, can be restored by concurrent interferon-gamma stimulation.
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PMID:Expression of pro-inflammatory cytokines by flow-sorted alveolar macrophages in severe pneumonia. 959 98

Cystic fibrosis (CF) is an inherited disorder associated with severe inflammation and repeated bacterial infection and colonization in the lung. Airway epithelium is involved in defence against bacteria, but this system may be defective in CF. Pro-inflammatory cytokines can stimulate the expression of inducible nitric oxide synthase (iNOS), an enzyme generating nitric oxide, which functions as an important mediator in host defence mechanisms. To understand better the poor resistance to infections in the CF lung, the expression of the iNOS gene was investigated in explanted lungs from patients with cystic fibrosis (n = 13), bronchiectasis (n = 3), emphysema (n = 14), and in normal lungs (n = 8). In addition, bronchial epithelial cell lines were examined to study iNOS gene expression in vitro. Strong immunoreactivity for iNOS was seen in inflammatory cells and bronchial epithelium in all the diseased lungs, except for bronchial epithelium in CF. Quantitative analysis showed a significant reduction in the area of epithelium immunostained in CF [CF 6.8 +/- 1.6 (% +/- SEM); emphysema 18.2 +/- 2.8; normal 9.6 +/- 0.8, P < 0.01], regardless of steroid treatment. These results were supported by in situ hybridization of iNOS mRNA, which showed a pattern of gene expression in CF, emphysema, and normal lung which paralleled that of protein immunoreactivity. Stimulation with cytokines (IL-1 beta, TNF-alpha, and IFN-gamma) increased the expression of iNOS mRNA detected by reverse transcriptase-polymerase chain reaction (RT-PCR) in cultures of normal (16HBE14o-), but not CF (CFBE41o-, with delta F508 CFTR mutation) epithelial cells. Expression of iNOS in inflammatory cells suggests that the gene is normal in CF. Absence of iNOS from bronchial epithelium may be due to low expression of the gene resulting from abnormalities in the signalling system that normally causes induction, such as cytokine receptors, second messengers or transcription factors. The resulting deficiency of the nitric oxide defence system may be relevant to the susceptibility of CF patients to pulmonary bacterial colonization.
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PMID:Lack of inducible nitric oxide synthase in bronchial epithelium: a possible mechanism of susceptibility to infection in cystic fibrosis. 961 86

Thyroid-associated ophthalmopathy (TAO) has a major effect on the two compartments of the retro-orbital (RO) space, leading to enlargement of the extraocular muscles and other RO tissues. T lymphocyte infiltration of RO tissue is a characteristic feature of TAO and there is current interest in whether these T cells are specifically and selectively reactive to RO tissue itself. We recently established 18 T cell lines (TCL) from RO adipose/connective tissue of six patients with severe TAO by using IL-2, anti-CD3 antibodies and irradiated autologous peripheral blood mononuclear cells (PBMC) to maintain the growth of T cells reactive to autologous RO tissue protein fractions. Here we report on the phenotype characteristics and cytokine gene expression profiles of these orbital TCL and on their immunoreactivity to the organ-specific thyroid antigens thyrotropin receptor (TSH-R), thyroidal peroxidase (TPO) and thyroglobulin (TG). Flow cytometry revealed that 10 TCL were predominantly of CD4+ phenotype, three being mostly CD8+ and five neither CD4+ nor CD8+. Analysis with reverse transcriptase-polymerase chain reaction (RT-PCR) of cytokine gene expression revealed both Th1- and Th2-like products in all TCL: IL-2 product (in 17 TCL), interferon-gamma (IFN-gamma) (n = 10), tumour necrosis factor-beta (TNF-beta) (n = 15), IL-4 (n = 12), IL-5 (n = 17), IL-6 (n = 13), TNF-alpha (n = 12) and IL-10 (n = 4). Reactivity to thyroid antigens was observed only in two TCL, the other 16 being uniformly unreactive. Although 10 out of 18 RO tissue-reactive TCL were predominantly CD4+ there were no significant relationships between TCL phenotype, cytokine gene profile, magnitude of reactivity to RO tissue protein or the (rare) occurrence of thyroid reactivity. The findings of both Th1- and Th2-like cytokine gene expression in all RO tissue-reactive TCL support the concept that TAO is a tissue-specific autoimmune disease, distinct immunologically from the thyroid, and involving both T cell and B cell autoimmune mechanisms in disease pathogenesis.
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PMID:Analysis of orbital T cells in thyroid-associated ophthalmopathy. 964 11

An experimental oral infection of neonatal (< 2 weeks old) lambs with a cervine isolate of Mycobacterium avium subspecies paratuberculosis (M.a. paratuberculosis), the causal agent of ruminant paratuberculosis (Johne's disease) was used to investigate bacteriological, histopathological and immunological changes during the early (up to 8 weeks) post-infection phase. In vitro culture for mycobacteria was positive in one faecal and three mesenteric lymph node (MLN) samples from the eight infected lambs. All mycobacterial isolates from MLN were identified as M.a. paratuberculosis by polymerase chain reaction (PCR). Small-to-medium sized focal granulomata were observed in jejunal (JPP) and ileal Peyer's patches (IPP) from four of the eight infected lambs. Compared with controls, JPP from all infected lambs had significantly (p < 0.05) higher proportions of CD8+ and CD2+ lymphocytes, and there were significantly (p < 0.05) fewer cells expressing B lymphocyte-associated markers in IPP and MLN. The T/B cell ratio was significantly (p < 0.05) increased in both JPP and MLN from infected lambs. The expression of a range of genes for cytokines was examined using specific reverse transcriptase PCR (RT-PCR) amplification of messenger RNA (mRNA) template isolated from MLN, JPP and IPP from both groups of animals. Densitometric analyses indicated that, in infected animals, MLN expressed significantly (p < 0.05) more mRNA for TNF-alpha: JPP had significantly increased (p < 0.05) mRNA for GM-CSF and significantly decreased (p < 0.05) mRNA for IL-4 and IFN-gamma. Infected lambs had significantly (p < 0.05) decreased titres of both circulating IgG and gut mycobacteria-associated IgG antibody. Infection was not associated with any consistent changes in lymphocyte reactivity to specific mycobacterial antigens, IFN-gamma release into supernatants from in vitro intestinal lymphocyte cultures or gut IgA antibody levels.
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PMID:Early immunopathological events in experimental ovine paratuberculosis. 965 60

Epidermal cells produce various kinds of cytokines and express cell adhesion molecules. To analyze early events which induced in human epidermis by stimulation with various chemicals, we analyzed mRNA of cytokines expressed in epidermis in a human skin organ culture system. After painting haptens, primary irritants or vehicle control on human skin specimens sliced to 1 mm thickness and cut into approximately 5 x 5 mm blocks, the pieces were cultured in serum-free medium. After separating epidermis from dermis, total RNA was extracted and mRNA of cytokines was assessed by the reverse transcriptase-poly-merase chain reaction. Only haptens induced IL-1 beta mRNA at 1-3 hours. TNF-alpha mRNA was induced 9 hours after application of haptens and 1 hour after application of primary irritants. IL-1 alpha mRNA was not induced by either haptens or primary irritants. Thus, cytokine mRNA expression induced by haptens in epidermis differs from that induced by primary irritants.
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PMID:Epidermal cytokine mRNA expression induced by hapten differs from that induced by primary irritant in human skin organ culture system. 971 73

Helicobacter pylori urease is absorbed into the gastric mucosa at sites of inflammation, but whether the enzyme activates mucosal macrophages is not known. Because mucosal macrophages differ phenotypically and functionally from blood monocytes, whether recombinant H. pylori urease (rUrease) activated purified lamina propria macrophages in vitro was investigated. rUrease (1-10 microgram/mL) induced primary mucosal macrophages to produce interleukin (IL)-1beta, IL-6, and tumor necrosis factor (TNF)-alpha but not IL-8 proteins in a dose-dependent manner (P<.05 to P<.001). Quantitative reverse transcriptase-polymerase chain reaction using capillary electrophoresis laser-induced fluorescence showed that rUrease (0.1-10 microgram/mL) also induced dose-dependent expression of IL-1beta, IL-6, and TNF-alpha but not IL-8 mRNA (P<.05), suggesting that rUrease-induced production of certain cytokines is regulated at the level of gene transcription. These findings indicate that the ability of H. pylori urease to activate mucosal macrophages, resulting in production of proinflammatory cytokines, may be involved in the pathogenesis of H. pylori-associated mucosal inflammation.
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PMID:Recombinant Helicobacter pylori urease activates primary mucosal macrophages. 978 Feb 78

GM-CSF is a cytokine with pleiotropic biological activities and is increasingly used in clinical trials. The present study demonstrates the ability of recombinant human granulocyte-macrophage colony-stimulating factor (rGM-CSF) to induce elevation of interleukin-10 (IL-10) mRNA and protein production in the monocytic cell line U937. As shown by a semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR), IL-10 mRNA increases up to 10 times when stimulated with rGM-CSF (100 U/ml) compared to nonstimulated control cells. Maximal IL-10 mRNA expression occurs at 6 h and remains high for 2 h. Thereafter IL-10 mRNA is downregulated and reaches basal level at approximately 24 h. IL-10 protein was measured by ELISA. The protein yield is dose-dependent on the rGM-CSF concentration. Combined stimulation of U937 cells with both GM-CSF and TNF-alpha results in an additive elevation of the IL-10 protein yield. Application of a neutralising antibody against TNF-alpha revealed that GM-CSF induces IL-10 expression independently from TNF-alpha. By using a luciferase reporter gene it was shown that rGM-CSF enhances IL-10 promoter activity 2-3-fold in a transient transfection assay.
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PMID:Recombinant human granulocyte-macrophage colony-stimulating factor triggers interleukin-10 expression in the monocytic cell line U937. 979 52


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