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)

We investigated the role of endogenous or exogenous nitric oxide (NO) on human lymphocyte function. We used sodium nitroprusside, nitroglycerine, S-nitroso-N-acetylpenicillamine, sodium nitrite and S-nitroso-L-glutathione as NO-generating compounds. All agents were used at doses that do not produce direct cytotoxicity as measured by trypan blue exclusion as well as chromium-51 release assay. The immune responses examined were peripheral blood lymphocytes (PBL) proliferation and IL-2 production after activation with OKT3 and PHA; allogeneic mediated proliferation and cell mediated cytotoxicity (CML) in MLR; IgG and IgM production after PBL activation with Con-A; proliferation and expression of IFN-gamma and IL-4 mRNA after activation of allogeneic CD4+T cell clones. Cytokine mRNA expression was measured by reverse transcriptase PCR. Our results show that proliferating lymphocytes do not produce a detectable amount of NO as measured by the Griess reaction. In separate experiments, the addition of NG-monomethyl-L-arginine (L-NMMA) did not affect lymphocyte proliferation. Sodium nitroprusside and nitroglycerine exerted a dose dependent antimitogenic effect, inhibited cytokine production and expression, CML generation and antibody production. DNA gel electrophoresis showed no evidence for enhanced programmed cell death. The antimitogenic effect could not be blocked by the NO scavengers, hemoglobin or methylene blue. In contrast, the other nitric oxide generating compounds did not inhibit lymphocyte mitogenesis. The results suggest that human lymphocytes do not produce appreciable amounts of NO to affect lymphocyte mitogenesis. Sodium nitroprusside and nitroglycerine have a potent but nonspecific immunoinhibitory effect on human lymphocyte function by a mechanism other than NO production. In addition, pharmacological levels of NO do not inhibit human lymphocyte mitogenesis.
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PMID:Analysis of the in vitro effect of exogenous nitric oxide on human lymphocytes. 920 99

Culture of rat pancreatic islets with interleukin-1 (IL-1) results in up-regulation of the inducible isoform of nitric oxide synthase and overproduction of nitric oxide (NO). This is associated with reversible inhibition of both glucose-induced insulin secretion and islet glucose oxidation, and these effects are prevented by the inducible nitric oxide synthase inhibitor NG-monomethylarginine. IL-1 also induces accumulation of nonesterified arachidonic acid in islets by an NO-dependent mechanism, and one potential explanation for that effect would involve an IL-1-induced enhancement of islet glycolytic flux. We have therefore examined effects of IL-1 on islet glycolytic utilization of glucose and find that culture of islets with IL-1 in medium containing 5.5 mM glucose results in suppression of islet glucose utilization subsequently measured at glucose concentrations between 6 and 18 mM. The IL-1-induced suppression of islet glucose utilization is associated with a decline in islet glucokinase mRNA content, as determined by competitive reverse transcriptase-polymerase chain reaction, and in glucokinase protein synthesis, as determined by immuoprecipitation experiments, and all of these effects are prevented by NG-monomethylarginine. These findings suggest that IL-1 can down-regulate islet glucokinase, which is the primary component of the islet glucose-sensor apparatus, by an NO-dependent mechanism. Because reductions in islet glucokinase levels are known to cause a form of type II diabetes mellitus, these observations raise the possibility that factors which increase islet NO levels might contribute to development of glucose intolerance.
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PMID:Interleukin-1 reduces the glycolytic utilization of glucose by pancreatic islets and reduces glucokinase mRNA content and protein synthesis by a nitric oxide-dependent mechanism. 921 38

Expression and androgen regulation of the gene for neuronal nitric oxide synthase (NOS I) were examined in neurons of the major pelvic ganglia in male rats. Some of these postganglionic neurons innervate the penis and produce nitric oxide, which is believed to play a major role in penile erection. Rats were either castrated or sham operated and implanted with SILASTIC brand capsules filled with powdered testosterone (T) or 5alpha-dihydrotestosterone (5alphaDHT) or left empty. After 4 days, the number of neurons intensely stained for NADPH-diaphorase as well as those giving a NOS I signal in in situ hybridization experiments increased in castrated rats treated with testosterone by 31% and 42%, respectively, relative to those in untreated castrated rats. This suggests that the increase in NADPH-diaphorase activity resulted from enzyme synthesis and was due to a modification of NOS I messenger RNA (mRNA) accumulation. After 7 days, Northern blot analysis showed that castration produced a decrease in the amount of NOS I mRNA relative to that of ribosomal RNA. This decrease was almost prevented by T treatment. No significant differences were observed by reverse transcriptase-PCR between 7-day and 28-day treatments. However, in 7-day castrated rats treated with 5alphaDHT, NOS I signals relative to those of hypoxanthine phosphoribosyltransferase, taken as reference, were significantly higher than those in castrated rats and resembled those in sham-castrated rats, suggesting that 5alphaDHT was probably more potent than testosterone in preventing the decrease in NOS I mRNA levels elicited by castration. These results show that NOS I can be positively regulated by androgens and are consistent with the suggestion that these steroids play a role in the physiological processes of penile erection.
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PMID:Androgens modulate nitric oxide synthase messenger ribonucleic acid expression in neurons of the major pelvic ganglion in the rat. 923 55

The hemodynamic effects of sepsis have been attributed in part to increased nitric oxide (NO) production and activation of guanylate cyclase, resulting in increased cGMP and relaxation of vascular smooth muscle. Heme oxygenase-1 (HO-1), a heat shock protein, has been shown to increase intracellular cGMP levels by formation of carbon monoxide (CO). We hypothesized that HO may be an important mediator of the hepatic response to infection. Male Swiss Webster mice underwent standard cecal ligation and puncture (CLP, 18 gauge 2X) or sham operation, and received either normal saline (NS) or Zn protoporphyrin IX (ZN PP IX), a competitive HO inhibitor (n = 6-8/group). Hepatic tissue samples were collected at 3, 6, 12, and 24 hr from separate mice. Serum was collected at 3 and 24 hr. A semiquantitative reverse transcriptase polymerase chain reaction method was used to measure HO-1 mRNA levels. Hepatic cGMP levels were measured by ELISA. Groups were repeated (n = 10/group) to assess mortality. Serum was collected at 3 and 24 hr to measure serum aspartate aminotransferase (AST) levels. HO-1 mRNA expression increased significantly by 3 hr after CLP and with HO inhibition alone (P < 0.05 vs sham + NS). HO-1 mRNA remained elevated through 24 hr. CLP animals with HO inhibition showed a significant reduction of hepatic cGMP following CLP compared with CLP + saline at 24 hr (P < 0.05). Mortality was significantly increased in the CLP + ZN PP group at 24 hr (P < 0.05 CLP NS vs CLP ZN PP). CLP caused a marked increase in AST activity, which was increased further with HO inhibition. HO-1 mRNA expression was induced by CLP. AST levels following CLP were markedly increased with HO inhibition. HO-1 function appeared to contribute to elevation of hepatic cGMP during peritonitis and may be an important hepatic adaptive response to infection.
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PMID:Heme oxygenase-dependent carbon monoxide production is a hepatic adaptive response to sepsis. 927 Dec 71

Nitric oxide (NO), a potent and versatile free radical, is synthesized in leukocytes by the inducible form of NO synthase (iNOS). In this study, leukocytes in pregnant mouse uterus were investigated for expression of the iNOS gene. Inducible NOS mRNA, which was identified by reverse transcriptase polymerase chain reaction, was high relative to an invariant mRNA, glyceraldehyde-3-phosphate dehydrogenase, in midgestation uteri (gestation days [g.d.] 10, 12, and 14) but was low in late-gestation uteri (g.d. 16 and 18). Inducible NOS protein, identified immunohistochemically in paraformaldehyde-fixed uteri taken from g.d. 6 through 18 using rabbit antibodies generated to mouse carboxyl terminus iNOS peptides, was prominent in a few myometrial mast cells at early stages and was strongly expressed from g.d. 6 through g.d. 14 in myometrial macrophage-like cells. Inducible NOS protein was first detected in uterine (u) natural killer (NK) cells at g.d. 8. Signals peaked in this lineage at g.d. 10 and declined thereafter. Uterine leukocytes cultured in vitro expressed the iNOS gene; a hybridoma cell line derived from mouse uNK cells (GWM1-2) contained iNOS mRNA, and cells migrating from mouse metrial gland explants included iNOS/ leukocytes. Large, granular iNOS + uNK cells were absent from the uteri of homologously mated pregnant TgE26 mice, an NK cell-deficient transgenic mouse strain, but immunoreactive iNOS was detectable in trophoblast, a cell lineage that did not contain immunoreactive iNOS in NK cell-competent Swiss-Webster mice. In TgE26 mothers gestating normal embryos, the same pattern was observed. Collectively, the results of this study demonstrate that iNOS is present in mouse uterine leukocytes including mast cells, macrophage-like cells, and uNK cells, and suggest that in the absence of uNK cells, the placenta synthesizes iNOS. These findings are consistent with the postulate that leukocyte NO contributes importantly to events associated with successful pregnancy that are likely to include relaxation of vascular smooth muscle.
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PMID:Expression of the inducible nitric oxide synthase gene in mouse uterine leukocytes and potential relationships with uterine function during pregnancy. 931 87

Endotoxin-mediated macrophage synthesis of nitric oxide (NO) is associated with immune effector function, intercellular communication, leukocyte adhesion, vascular integrity, and neurotransmission. However, little is known of the cellular receptor and signal transduction pathway by which endotoxin induces NO production. With the use of a model of ANA-1 murine macrophages, we stimulated NO production by incubation with increasing concentrations of endotoxin and 5% fetal calf serum. In selected instances, the anti-CD14 antibody, ED9, was added. Endotoxin-mediated NO synthesis was dependent on CD14 function and the presence of an additional serum factor. Endotoxin treatment increased plasma membrane GTPase activity and 35S-labeled guanosine 5'-O-(3-thiotriphosphate) ([35S]GTP gamma S) binding. Conversely, coincubation of cells with endotoxin and the heterotrimeric G protein inhibitors, suramin and guanosine 5'-O-(2-thiodiphosphate) trilithium salt, was associated with decreased NO synthesis, plasma membrane GTPase activity, and [35S]GTP gamma S binding. Blockade of CD14 or G protein function was associated with ablation of endotoxin-mediated inducible NO synthase (iNOS) protein expression, iNOS mRNA levels, and iNOS gene transcription, as determined by immunoblot, reverse transcriptase-polymerase chain reaction, and nuclear run-on analyses, respectively. These results indicate that endotoxin-mediated NO synthesis is a CD14-heterotrimeric G protein-dependent process.
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PMID:CD14-dependent mechanism for endotoxin-mediated nitric oxide synthesis in murine macrophages. 931 24

The cytokine interleukin-1beta (IL-1beta) has been shown to inhibit insulin secretion and destroy pancreatic islets by a mechanism that involves the expression of inducible nitric oxide synthase (iNOS), and the production of nitric oxide (NO). Insulin containing beta-cells, selectively destroyed during the development of autoimmune diabetes, appear to be the islet cellular source of iNOS following treatment with IL-1beta. In this study we have evaluated the presence of type I IL-1 signaling receptors on purified pancreatic beta-cells. We show that the interleukin-1 receptor antagonist protein (IRAP) prevents IL-1beta-induced nitrite formation and IL-1beta-induced inhibition of insulin secretion by isolated islets and primary beta-cells purified by fluorescence-activated cell sorting (FACS). The protective effects of IRAP correlate with an inhibition of IL-1beta-induced iNOS expression by islets and FACS purified beta-cells. To provide direct evidence to support beta-cell expression of IL-1 type I signaling receptors, we show that antiserum specific for the type I IL-1 receptor neutralizes IL-1beta-induced nitrite formation by RINm5F cells, and that RINm5F cells express the type I IL-1 receptor at the protein level. Using reverse transcriptase-polymerase chain reaction (RT-PCR), the expression of type I IL-1 signaling receptors by FACS purified beta-cells and not alpha-cells is demonstrated. These results provide direct support for the expression of type I IL-1 receptors by primary pancreatic beta-cells, the cell type selectively destroyed during the development of autoimmune diabetes.
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PMID:Evidence for the presence of type I IL-1 receptors on beta-cells of islets of Langerhans. 937 6

Nitric oxide (NO) is suggested to play a role in mediating pulmonary injury. However, interspecies differences appear to exist in the ability of alveolar macrophages (AM) to express the inducible nitric oxide synthase (iNOS) and to generate NO. The purpose of this study was to compare iNOS expression and NO production by rat, hamster, monkey, and human AM using the identical experimental conditions in vitro. As AM donors, CD rats, Syrian golden hamsters, cynomolgus monkeys, and nonsmoking, healthy human volunteers were used. The AM were obtained by bronchoalveolar lavage and stimulated in vitro with various concentrations and combinations of lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). The oxidation product of NO, nitrite, was measured in the AM supernatant by the Griess reaction. The expression of iNOS in AM was detected using immunocytochemistry and immunoblotting. The expression of iNOS mRNA was assessed by reverse transcriptase-polymerase chain reaction (RT-PCR). Rat AM, stimulated with either LPS or IFN-gamma, produced nitrite in a time- and dose-dependent manner. Combination of LPS and IFN-gamma resulted in a significantly enhanced nitrite formation. However, none of the treatments was able to induce hamster, monkey, or human AM to release measurable amounts of nitrite. Whereas expression of iNOS protein was only detected in stimulated rat AM, expression of iNOS mRNA was found in unstimulated and stimulated rat AM, slightly in stimulated hamster AM, but not in monkey and human AM. In conclusion, our findings point to distinct regulatory mechanisms of the NO pathway in AM from these four different species.
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PMID:Expression of inducible nitric oxide synthase and formation of nitric oxide by alveolar macrophages: an interspecies comparison. 940 Jul 41

Cardiac hypertrophy and heart failure are frequently accompanied by elevated plasma levels of tumor necrosis factor alpha (TNF alpha), the pathogenetic relevance of this finding being a matter of debate. In human acute septic cardiomyopathy, on the other hand, the negative inotropic impact of TNF alpha on the heart is well documented and frequently ascribed to the induction of inducible nitric oxide (NO) synthase (iNOS) and an enhanced production of NO in the heart. Yet the present study presents evidence that in cardiomyocytes TNF alpha in non-toxic concentrations specifically depresses contractile performance independent of NO. In spontaneously beating neonatal rat cardiomyocytes, TNF alpha in a low, pathophysiologically relevant concentration (10 U/ml, 1-3 days) does not alter basal pulsation amplitude, but blocks alpha- and beta-adrenoceptor-stimulated increase in contractility and beating irregularity and impairs the impact of high extracellular calcium on contractile performance. However, this low TNF alpha-concentration does not suffice to induce iNOS - documented by reverse transcriptase polymerase chain reaction - or enhance nitrite concentrations in the cell culture supernatants as a measure of cellular NO production, neither in the presence nor absence of dexamethasone (0.1 micro M). Only in high concentration - the specific proinflammatory action being documented by an enhanced release of interleukin-6 from cardiomyocytes - TNF alpha (1000 U/mol; 6, 24 h) weakly induces the mRNA for iNOS, with a consecutive moderate rise in cellular nitrite production. TNF alpha-incubation (10-1000 U/ml) does not alter the morphological appearance of the cells displayed by phase contrast microscopy or evoke gross cytotoxicity.
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PMID:Tumor necrosis factor alpha (TNF alpha) is cardiodepressant in pathophysiologically relevant concentrations without inducing inducible nitric oxide-(NO)-synthase (iNOS) or triggering serious cytotoxicity. 940 66

Nitric oxide synthase (NOS) has been characterized in different tissues, and its localization has been suggested in different neuronal tissues, including the myenteric neurons and other nonneuronal cells. The present study examined the distribution of the neuronal NOS (nNOS) mRNA in different tissues of the opossum gastrointestinal tract, internal anal sphincter (IAS) smooth muscle cells, and myenteric neurons using slot-blot and Northern blot hybridization techniques with a specific rat brain nNOS cDNA probe. Significant levels of nNOS gene expression were found in both smooth muscle cells and myenteric neurons of the opossum IAS. This finding was confirmed by reverse transcriptase-polymerase chain reaction analysis of the RNA obtained from cultured opossum IAS smooth muscle cells and myenteric neurons and also from human intestinal smooth muscle and neuroblastoma cell lines. Pyloric sphincter had the highest level of nNOS gene expression compared with other gastrointestinal tissues. There was no significant difference in the nNOS gene expression between other sphincteric and nonsphincteric tissues examined. The present study shows the presence of nNOS gene expression in both neurons and smooth muscle cells. The higher levels of nNOS gene expression in the pyloric sphincter compared with other tissues may have pathophysiological significance in some disease conditions.
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PMID:Neuronal NOS gene expression in gastrointestinal myenteric neurons and smooth muscle cells. 943 91


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