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)

Patients undergoing hemodialysis, present platelets (PLTs) with physiological dysfunction. Aggregated PLTs stimulate endothelin-1 (ET-1) secretion from endothelial cells. In turn, ET-1 abolishes PLT aggregation. The aim of this study was to determine any presence of ET-1 in the PLTs of hemodialysed patients and to compare its levels with normal subjects. Platelets, isolated from hemodialysed patients, revealed lower aggregation (76 +/- 13%) compared with healthy subjects (96 +/- 2%). Plasma ET-1 was increased in hemodialysed patients 23.5 +/- 3.2 fmol/ml, vs. 10.9 +/- 1.6 fmol/ml in normal subjects. Immunoreactive endothelin-1 was detected in the platelets of both groups. The intraplatelet ET-1 of hemodialysed patients was 13.8 +/- 3.1 fmol/mg protein, vs. 7.9 +/- 1.3 fmol/mg protein in normal individuals. Total RNA was isolated from platelets and reverse transcriptase-mediated polymerase chain reaction (RT-PCR) revealed no presence of the preproET-1 mRNA in either normal or hemodialysed platelets. This suggests that ET-1 is internalized from the plasma. In summary, platelets contain but do not express ET-1. The increased levels of ET-1 in the platelets of the hemodialysed patients may be partly responsible for their lower aggregation.
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PMID:Increased endothelin-1 content in the platelets of hemodialysed patients. 934 91

The aim of this investigation was to determine whether the endothelin receptor subtype of a megakaryoblastic cell line (MEG-01) changes during culture passages as cells undergo maturation and differentiation. On early-passage cells, binding of [125I]endothelin-1 was completely inhibited by 1 microM BQ 123 (cyclo-[D-tryptophanyl-D-aspartyl-prolyl-D-valyl-leucyl]), but not by sarafotoxin 6C. Also the endothelin-1-enhancing effect on [Ca2+]i was prevented by BQ 123, whereas sarafotoxin 6C had no effect on [Ca2+]i. In late-passage cells, endothelin ET(B) analogs, unlike endothelin ET(A) analogs, competed with binding of [125I]endothelin-1. Endothelin ET(B) receptor agonists increased [Ca2+]i while the endothelin-1-induced response was inhibited by BQ 788 ([N-[(2R,6S)-2,6-dimethyl-piperidinocarbonyl]-4-methyl-D-leucyl]-[ N(omega)-(methoxycarbonyl)-D-tryptophanyl]-D-norleucine), but not by BQ 123, although both endothelin ET(A) and ET(B) receptor mRNAs were expressed, as shown by reverse transcriptase-polymerase chain reaction. These results demonstrate that in MEG-01 cells switch from expression of endothelin ET(A) to expression of ET(B) receptors during culture. The data also suggest that late-passage MEG-01 cells look like platelets, in terms of endothelin receptor subtype.
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PMID:Change of endothelin receptor subtype in the MEG-01 human megakaryoblastic cell line. 960 Jun 67

The vasoconstrictive peptide endothelin-1 (ET-1) is an autocrine/paracrine peptide of putative pathophysiological importance in renal transplant medicine. The aim of the present study was to develop a method for analysis of gene expression of the renal endothelin system in humans. Only small amounts of tissue are available from renal cortical needle biopsies. Thus, in the present study we developed a quantitative assay based on the competitive reverse transcriptase polymerase chain reaction (RT-PCR) technology. We quantified endothelin A (ET(A)) and B (ET(B)) receptor subtype mRNAs and preproET-1 mRNA levels in renal cortex biopsies obtained before nephrectomy of healthy kidney donors. Mean (+/- SEM) mRNA levels of the ET(A) and ET(B) receptor subtypes in 26 living donors were 212 +/- 23 and 368 +/- 56 amol/microg total RNA, respectively. The preproET-1 mRNA level in 19 living donors was 213 +/- 28 amol/microg total RNA. The inter-assay coefficient of variation (CV) for the assay was 10%; the intra-assay CV was 6-13%. The competitive RT-PCR assay described provides an accurate tool for gene expression investigation of the human endothelin system in renal cortical needle biopsies.
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PMID:Quantification of mRNA levels of endothelin receptor subtypes and preproEndothelin-1 in renal needle biopsies by competitive reverse transcriptase polymerase chain reaction. 974 17

The biochemical and molecular endothelin-1 (ET-1) responses to high dose morphine sulfate infusion were studied in conscious newborn piglets (n = 6) that received a loading dose of 100 micrograms/kg over 5 min followed by a continuous i.v. infusion dose of 100 micrograms.kg-1.h-1 for 4 h. The control group (n = 6) received equivalent volume loading and infusion doses of 5% dextrose. Blood samples were drawn serially from the femoral artery and sagittal sinus vein before (0), during (30 min, 1, 2, 3, and 4 h), and post (1 and 2 h) infusion. Five micrograms of total RNA obtained from brainstem tissue homogenates was analyzed by reverse transcriptase--polymerase chain reaction (RT-PCR). The amounts of mRNA encoding ET-1, and endothelin receptor subtypes ETA and ETB, were semiquantitated using densitometric scanning. Morphine infusion resulted in elevated respiratory rate and mean arterial blood pressure, with no effect on arterial pH, Po2, and O2 saturation. Compared with the control group, morphine induced significant elevations in plasma ET-1 levels following the bolus dose (systemic: 13.2 +/- 3.6 vs. 8.6 +/- 2.2 pg/mL, p < 0.05; sagittal sinus vein: 13.7 +/- 3.4 vs. 8.2 +/- 0.9 pg/mL, p < 0.01). These effects lasted up to 2 h after discontinuation of morphine infusion (systemic: 14.5 +/- 3.4 to 18.7 +/- 5.7 pg/mL vs. 7.5 +/- 0.8 to 9.4 +/- 3.2 pg/mL, p < 0.05 to p < 0.01; sagittal sinus vein: 14.8 +/- 2.7 to 17.6 +/- 2.8 pg/mL vs. 7.5 +/- 1.4 to 9.4 +/- 3.4 pg/mL, p < 0.05 to p < 0.01). The RT-PCR assay showed a twofold (p < 0.02) upregulation in ET-1 and a threefold (p < 0.007) upregulation in ETA receptor mRNA expression in the brainstem of morphine-treated animals. In contrast, there was a threefold (p < 0.0001) downregulation of the ETB receptor mRNA expression. The rapid and sustained elevations in systemic arterial and sagittal sinus venous ET-1 levels suggest a role for ET-1 in the morphine-induced excitatory responses observed in newborn piglets. Upregulation of ETA receptors and downregulation of ETB receptors in the brainstem with high doses of morphine may indicate possible effects on cerebral vascular tone.
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PMID:Biochemical and molecular endothelin responses to morphine sulfate infusion in conscious newborn piglets. 979 54

Osteoclast activity is inhibited by elevated [Ca2+]o; however, the underlying molecular mechanism is unknown. We used the human osteoclast-like cells GCT23 to elucidate their cation-sensing properties. Cells responded to elevated [Ca2+]o with rapid concentration-dependent [Ca2+]i transients (EC50 = 7.8 mm, time to peak 44 +/- 4 sec) that were due to release from intracellular stores, followed by Ca2+ influx across the plasma membrane. Ca2+ store depletion by thapsigargin, endothelin-1, or bradykinin activated calcium entry pathways. Cells responded similarly to Ni2+ and Cd2+ with albeit slower kinetics (EC50 <10 microm and <100 microm, times to peak 140 +/- 25 sec and 150 +/- 24 sec, respectively). The three cations stimulated inositol phosphate production (two-fold, p <.02) similar to bradykinin (2.5-fold, p <. 002), which activates a phospholipase C (PLC)-coupled receptor in GCT23 cells. The cells did not respond to 0.1-1 mM Gd3+ or neomycin B, indicating that the parathyroid calcium receptor (PCaR) is not functionally expressed. In confirmation, PCaR could not be detected by reverse transcriptase polymerase chain reaction in GCT23 cells and in mouse osteoclasts, and the calcimimetic compound NPS R-568 failed to produce the left shift of the concentration-response curve characteristic for PCaR. Our data demonstrate for the first time that cation sensing by osteoclast-like GCT23 cells is mediated by a PLC-coupled receptor that is not identical to PCaR.
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PMID:A novel calcium sensor stimulating inositol phosphate formation and [Ca2+]i signaling expressed by GCT23 osteoclast-like cells. 989 59

1. We confirmed that endothelium-independent contraction of the rabbit pulmonary artery (RPA) is mediated through both an endothelin A (ET(A)R) and endothelin B (ET(B2)R) receptor. 2. The response of endothelium-denuded RPA rings to endothelin-1 (ET-1, pD2 = 7.84 +/- 0.03) was only partially inhibited by BQ123 (10 microM), an ET(A)R antagonist. 3. Pretreatment with 1 nM sarafotoxin S6c (S6c), an ET(B)R agonist, desensitized the ET(B2)R and significantly attenuated the response to ET-3 (pD2 = 7.40 +/- 0.02 before, <6.50 after S6c). 4. Pretreatment with S6c had little effect on the response to ET-1, but BQ123 (10 microM) caused a parallel shift to the right of the residual ETAR-mediated response to ET-1 (pD2 = 7.84 +/- 0.03 before S6c, 7.93 +/- 0.03 after S6c, 6.81 +/- 0.05 after BQ123). 5. Binding of radiolabelled ET-1 to early passage cultures of RPA vascular smooth muscle cells (VSMC) displayed two patterns of competitive displacement characteristic of the ET(A)R (BQ123 pIC50 = 8.73 +/- 0.05) or ET(B2)R (S6c pIC50 = 10.15). 6. Competitive displacement experiments using membranes from late passage VSMC confirmed only the presence of the ET(A)R (ET-1 pIC50 = 9.3, BQ123 pIC50 = 8.0, S6c pIC50 < 6.0). 7. The ET(A)R was functionally active and coupled to rises in intracellular calcium which exhibited prolonged homologous desensitization. 8. Using a reverse transcriptase polymerase chain reaction for the rabbit ET(B2)R, we demonstrated the absence of mRNA expression in phenotypically modified VSMC. 9. We conclude that the ET(B2)R expressed by VSMC which mediates contraction of RPA is rapidly down-regulated at the transcriptional level during phenotypic modulation in vitro.
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PMID:Transcriptional down-regulation of the rabbit pulmonary artery endothelin B receptor during phenotypic modulation. 1005 Nov 26

Oxidative stress and inflammatory reactions associated with stresses that may lead to shock promote hepatic microcirculatory dysfunction, which may lead to hepatic injury. Because altered liver microcirculation may result from an imbalance in the expression of stress-induced vasoactive mediators, our study was conducted to investigate changes in the expression of genes encoding endothelin-1 (ET-1), its receptors, ET(A) and ET(B), heme-oxygenase 1 (HO-1), and inducible nitric oxide synthase (iNOS), using two different rat models of liver stress: ischemia/reperfusion of the liver and lipopolysaccharide (LPS)-induced endotoxemia. In ischemia/reperfusion experiments, rats were subjected to 1 h hepatic ischemia, followed by 6 h of reperfusion. Endotoxemia was induced by i.p. injection of LPS (1 mg/mL/kg body weight); rats were studied after 6 h. mRNA levels were estimated using semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) on total RNA samples prepared from experimental and sham control rat livers. In the ischemic reperfused livers the levels of mRNA for ET-1, ET(B), HO-1, and iNOS were significantly elevated. The fold increase versus sham was 2.5+/-1.1 (ET-1), 2.1+/-1.3 (ET(B)), 2.1+/-.8 (HO-1), and 6.4+/-3.9 (iNOS). In contrast, the expression of ET(A) receptor gene was reduced after ischemia/reperfusion (to 73+/-1% of sham). In the separate experiments we analyzed the same mRNAs levels after 1 h of ischemia (no reperfusion), and did not detect any changes. During endotoxemia we observed a marked increase in iNOS mRNA level (>24-fold), as well as a marked elevation of the other four mRNAs. The fold increase versus sham was 6.1+/-1.7, ET-1); 1.5+/-.3 (ET(A)); 1.6+/-.4 (ET(B)); and 2.4+/-.34 (HO-1). These results show that liver stress, induced by ischemia/reperfusion or LPS injection have characteristic patterns of vasoregulatory genes expression indicating that, although both stresses result in an increase in specific vascular reactivity, different pathways are involved in inducing the hepatic vascular stress response.
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PMID:Patterns of vasoregulatory gene expression in the liver response to ischemia/reperfusion and endotoxemia. 1018 69

Normal human melanocytes have been shown to respond to the signal peptide endothelin by increased proliferation and melanin formation. Contradictory findings, however, have been reported about which of the two endothelin receptors (EDNRA or EDNRB) is expressed in normal melanocytes and melanoma cells. Moreover it was not clear whether malignant cells differ from their normal precursors in this respect. Screening a melanocyte cDNA library for genes downregulated in melanomas identified clones specific for EDNRB. Northern blots proved that the corresponding mRNA is generally expressed in cultures of human cutaneous melanocytes and congenital melanocytic nevus cells. In 16 of 17 melanoma cell lines, however, the expression of EDNRB mRNA was strongly downregulated. EDNRA was only weakly expressed and detectable by northern blotting in 12 of 17 cultures of benign melanocytic cells and four of 17 melanoma cell lines. Nested reverse transcriptase-polymerase chain reaction proved several melanoma cell lines to be completely negative for EDNRA expression. Gene deletion as the cause of missing endothelin receptor expression was ruled out by genomic Southern blots. Receptor binding assays confirmed RNA data revealing 1.6 x 105 endothelin-1 binding sites per cell for a melanocyte culture and between 8.7 x 104 and 400 sites per cell for melanoma cell lines. Expression of pigmentation genes coding for tyrosinase, TRP-1 and TRP-2 correlated positively with that of EDNRB but negatively with EDNRA expression. EDNRB but not EDNRA expression is therefore typical for melanocytic cells, and downregulation of EDNRB seems to be an important characteristic of melanoma cells possibly related to malignancy or apoptosis.
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PMID:Downregulation of endothelin B receptor in human melanoma cell lines parallel to differentiation genes. 1038 40

Tail blood flow (TBF) in the rat markedly increases during sympathetic withdrawal such as hyperthermia or lumbar sympathetic blockade. However, a long-term alteration of TBF after chronic sympathetic denervation is not well understood. In the present study, TBF following lumbar sympathectomy (LSX) was observed to ascertain whether subsequent changes in TBF occur in the absence of the sympathetic nervous activity in the rat tail. Assessed by recording tail and rectal temperature, the LSX immediately caused an increase in TBF. TBF was gradually decreased along with time and returned to the sham operated (SO) control level within 4 days. About a week after the surgery, a rapid increase in TBF in response to whole body heating was almost abolished in denervated animals. Neither hexamethonium (20 mg/kg, i.v.) for ganglion blockade nor intra-arterial infusion of alpha-receptor antagonist, phentolamine (10, 100 microg) produced vasodilation in LSX animals. Nitroprusside, a donor of nitric oxide, produced an increase in TBF in both LSX and SO animals. These results indicate that the tail vasculature after LSX constricts with capability to be vasodilated independent of sympathetic reinnervation. Quantification of the tail vascular mRNA expression by reverse transcriptase-polymerase chain reaction showed less endothelial nitric oxide synthetase in LSX group than that in SO group whereas endothelin-1 was not significantly different in both groups. It is suggested that functional changes in tail vascular endothelium takes at least a part in the reduction in TBF after LSX.
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PMID:The influence of chronic sympathectomy on cutaneous blood flow in the rat tail. 1045 3

Smoking is associated with endothelial dysfunction and increased plasma levels of endothelin-1. The component of tobacco smoke inducing these effects is unknown. Carbon monoxide induces hypoxia, and there is evidence of carbon monoxide acting as a local mediator in both endothelial and smooth muscle cells. The purpose of this study was to determine whether chronic carbon monoxide exposure similar to that experienced by smokers affects myocardial endothelin-1 expression. Sprague-Dawley female rats were exposed to carbon monoxide 100 ppm for one week or to 100 ppm for one week and 200 ppm for a second week. Carboxyhaemoglobin was 12+/-0.9% in the low and 23+/-1.1% in the high carbon monoxide exposure group. Endothelin-1 expression was measured by competitive reverse transcriptase polymerase chain reaction. High carbon monoxide exposure increased endothelin-1 mRNA by 54+/-12% (P<0.001) in the left ventricle and by 53+/-12% (P<0.001) in the right ventricle. In the low carbon monoxide exposure group corresponding changes were 43+/-14% (P=0.06) and 12+/-16%(P=0.29). Right ventricular weight increased by 18+/-7% (P=0.02) after high and by 16+/-5% (P=0.02) after low exposure. Left ventricular weight was elevated by 5+/-2% (P=0.05) when both exposure groups were compared to controls. We conclude that chronic carbon monoxide exposure leading to carboxyhaemoglobin levels similar to those observed in smokers increases endothelin-1 gene expression and induces myocardial hypertrophy in the rat.
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PMID:Chronic carbon monoxide exposure in vivo induces myocardial endothelin-1 expression and hypertrophy in rat. 1056 19


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