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)

Estrogen replacement therapy is cardioprotective in postmenopausal women; however, the precise molecular mechanisms for this modulation are not fully elucidated. We previously showed that chronic estrogen replacement therapy reduced angiotensin-converting enzyme (ACE) activity in tissue extracts and serum with an associated reduction in plasma angiotensin II. A reverse transcriptase-polymerase chain reaction assay was developed to determine whether estrogen treatment regulates tissue ACE mRNA concentration. Total RNA was isolated from kidney cortex, kidney medulla, lung, and aorta of ovariectomized Sprague-Dawley rats after 21 days of chronic 17beta-estradiol replacement therapy (5 mg pellet per rat SC) or placebo. A marked decrease in densitometric intensity ratios of amplified ACE cDNA to elongation factor-1alpha control cDNA was observed in all tissues from placebo-treated rats compared with the estradiol-treated rats (renal cortex: 0.29+/-0.04 versus 0.14+/-0.02; renal medulla: 0. 37+/-0.04 versus 0.24+/-0.03; lung: 4.49+/-0.37 versus 2.49+/-0.59; and aorta: 0.41+/-0.04 versus 0.29+/-0.02; all P<0.05). A comparable reduction in ACE activity was detected in tissue extracts from kidney cortex, kidney medulla, and lung of hormone-treated animals. Incubation of purified rat lung ACE with 1 or 10 micromol/L 17beta-estradiol had no effect on enzyme activity. These results suggest that estrogen treatment regulates tissue ACE activity by reducing ACE mRNA concentrations. Thus, the beneficial cardiovascular effects of estrogen may be mediated in part by downregulation of ACE with a consequent reduction in the circulating levels of the vasoconstrictor angiotensin II, a decrease in the metabolism of the vasodilator bradykinin, and an increase in the production of the vasorelaxant angiotensin-(1-7).
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PMID:Estrogen regulation of angiotensin-converting enzyme mRNA. 993 Nov 24

In the kidney and colon 11beta-hydroxysteroid dehydrogenase type 2 (11beta-HSD2) inactivates cortisol to cortisone, thereby protecting the non-selective mineralocorticoid receptor from cortisol. Deficiency of 11beta-HSD2 results in cortisol-mediated sodium retention and hypertension, suggesting that the physiological regulation of 11beta-HSD2 in mineralocorticoid target tissues may be important in modulating sodium homoeostasis and blood pressure control. Using the human epithelial colon cell line SW-620, reverse transcriptase-polymerase chain reaction and enzyme kinetic analysis indicated expression of only 11beta-HSD2 (Km for cortisol 66 nmol/l). Bradykinin (10(-8) to 10(-12) mol/l), frusemide (10(-4) to 10(-9) mol/l), benzamiloride hydrochloride (10(-5) to 10(-10) mol/l) and atrial natriuretic peptide (10(-6) to 10(-10) mol/l) had no effect on 11beta-HSD2 expression. Using a range of concentrations of angiotensin II (2x10(-8) to 2x10(-5) mol/l) a significant reduction in activity was seen but only at supra-physiological concentrations, [e.g. 2x10(-6) mol/l at 4 h pretreatment: 36.7+/-2.0 pmol cortisone. h-1.mg-1 (mean+/-S.E.M.) compared with 45.1+/-1.7 pmol.h-1.mg-1 in control; P<0.05]. The angiotensin-converting enzyme inhibitors captopril, enalapril, lisinopril, perindopril, quinapril and trandolapril at 10(-7) mol/l, but not fosinopril, significantly increased 11beta-HSD2 activity after pretreatment for 16 or 24 h (P<0.05-P<0.01 compared with control). No effects were seen at 4 h pretreatment. Hydrochlorothiazide (10(-7) mol/l) significantly decreased 11beta-HSD2 activity (P<0.05 compared with control) at 4 h pretreatment. Commonly used diuretics, atrial natriuretic peptide and physiological concentrations of angiotensin II and bradykinin do not alter 11beta-HSD2 activity. In contrast, a series of angiotensin-converting enzyme inhibitors significantly increase 11beta-HSD2 activity in vitro. This may explain how intrarenal infusions of angiotensin-converting enzyme inhibitors increase renal sodium excretion independent of circulating concentrations of angiotensin II. The interaction between angiotensin-converting enzyme inhibitors and 11beta-HSD2 may be an additional mechanism by which the former can lower blood pressure.
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PMID:Regulation of 11beta-hydroxysteroid dehydrogenase type 2 by diuretics and the renin-angiotensin-aldosterone axis. 1033 75

Bradykinin (BK)-stimulated colonic Cl- secretion was studied in T84 colonic adenocarcinoma cells by measuring BK (50 nM)-evoked changes in cytosolic free [Ca2+] ([Ca2+]i), membrane conductance and transepithelial ion transport. In T84 cells grown on impermeable supports, BK stimulated a transient increase in [Ca2+]i as assessed by fura-2 ratio imaging. In cell-attached, patch-clamp recordings, BK transiently activated low-conductance K channels. These channels were activated/inactivated reversibly in inside-out patches by switching [Ca2+]i in the bath between 30 nM and 100 nM. Excised channels recorded with 160 mM [K+] in bath and pipette exhibited an inwardly rectifying current/voltage-relation, conductances of 10+/-1 pS and 34+/-4 pS (n=10) at positive and negative voltages, respectively, and a 15-fold lower permeability for Na+ than for K+. The mean open probability of these channels did not depend on voltage but increased with increasing [Ca2+]i with an apparent concentration for a half-maximal response (EC50) of 110 nM, resembling that of hSK4 K+ channels. Application of the reverse transcriptase-polymerase chain reaction technique showed hSK4 messenger ribonucleic acid (mRNA) to be expressed in T84 cells. Macroscopic currents in T84 cells showed a similar dependence on [Ca2+]i. Whole cell conductance (in nS/10pF) increased from 0.5+/-0. 1 (n=6) at 10 nM [Ca2+]i in the pipette solution to 1.5+/-0.2 (n=7) at 100 nM, and to 2.0+/-0.5 (n=7) at 1 microM due to activation of a K+ conductance. In Ussing-chambered T84 monolayers grown on filters, BK did not evoke a short-circuit current (Isc). When, however, the monolayers were pre-stimulated by forskolin (1 microM), BK further enhanced Cl-secretion (DeltaIsc=21+/-5 microA/cm2, n=10) transiently and biphasically. In conclusion, BK enhances cyclic adenosine monophosphate-stimulated Cl- secretion in T84 cells, probably via basolateral, Ca2+-liganded activation of low-conductance hSK4-type K+ channels.
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PMID:Bradykinin-stimulated Cl- secretion in T84 cells. Role of Ca2+-activated hSK4-like K+ channels. 1037 87

Bacterial infection of the amniotic cavity is one of the most frequent causes of preterm delivery. Bacterial products activate a network of autocrine and paracrine mediators in fetal membranes and decidua, with prostaglandins finally inducing contractions of the myometrium. Bradykinin and its B2-receptor (B2R) seem to be part of this network. In cultured decidua-derived cells, bradykinin stimulates the release of arachidonic acid, interleukin-6 (IL-6), and interleukin-8 (IL-8). These effects are prevented by the specific B2R antagonist Hoe 140. Using a pooled antiserum against peptide sequences of the B2R protein, the receptor can be visualized immunocytochemically. The cells contain mRNA for the B2R, as shown by reverse transcriptase polymerase chain reaction (RT-PCR). Binding studies reveal specific and saturable binding sites for bradykinin with characteristics of the B2R. Binding of bradykinin to the cells is enhanced by the inflammatory mediator interleukin-1beta. In summary, human decidua-derived cells express the B2R, its expression is upregulated in response to interleukin-1beta, and bradykinin stimulates the secretion of further mediators by these cells. Thus, bradykinin and the B2R could play a central role in decidual activation. If so, B2R antagonists would add to established tocolytic therapies that are applied together with antibiotics in cases of chorioamnionitis at low gestational age.
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PMID:The bradykinin B2-receptor in human decidua. 1063 76

Ca(2+)-activated K(+) (K(Ca)) channels have been suggested to play a role in the control of endothelial functions such as regulation of vascular tone and cell proliferation. We established a method for single-cell reverse transcriptase-polymerase chain reaction analysis in combination with the patch-clamp technique to characterize K(Ca) channel expression and function in single endothelial cells (ECs) within the endothelial monolayer of intact human mesenteric arteries (MAs) and in disease states. We tested whether endothelial K(Ca) channel expression and function are altered in MAs obtained from patients with colonic adenocarcinoma (CA) compared with those in MAs from non-cancer patients with inactive diverticulitis. Expression of the intermediate-conductance K(Ca) channel (hIK1) was detected in non-cancer and CA patients. In whole-cell patch-clamp measurements, only ECs expressing hIK1 exhibited corresponding K(Ca) currents, whereas respective K(Ca) currents were missing in hIK1-negative ECs. This heterogeneity of hIK1 expression patterns is indicative of a specialized subset of ECs within the endothelial monolayer. In CA patients, compared with non-cancer patients, a 2.5-fold increase in hIK1-expressing ECs per MA was observed (P:<0.05). However, K(Ca) current densities in hIK1-expressing ECs of both groups were similar. In addition to hIK1, expression of the large-conductance K(Ca) channel (hSlo) was detected in single ECs from CA patients. The increased K(Ca) channel expression in CA patients resulted in a 2. 7-fold increase of bradykinin-induced endothelial hyperpolarization compared with controls (P:<0.05). This increased expression and function of K(Ca) channels might indicate an altered functional state of the endothelium in cancer patients and could play a role in tumor angiogenesis.
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PMID:Expression and function of endothelial Ca(2+)-activated K(+) channels in human mesenteric artery: A single-cell reverse transcriptase-polymerase chain reaction and electrophysiological study in situ. 1098 42

Bradykinin has vasodilatory and tissue-protective effects exerted via its B(2) type receptor, whereas the B(1) receptor is constitutively absent but inducible by inflammation and toxins. In previous studies, we found that B(2) receptor gene knockout mice exhibit overexpression of the B(1) receptor, which assumes a vasodilatory function and is further upgraded in renovascular hypertension. The present study was designed to explore the effects of excess angiotensin II (ANG II) on B(1) receptor and B(2) receptor gene expression in mouse cardiomyocytes and rat vascular smooth muscle cells (VSMC) in vivo (after a 3-day infusion of 30 ng/min ANG II in 11 wild-type and in 13 genetically engineered mice with deleted B(2) receptor gene) and in vitro (ANG II added in rat VSMC culture in the presence or absence of AT(1) or AT(2) receptor antagonist). Expression of B(1) and B(2) receptor mRNA was assessed by reverse transcriptase-polymerase chain reaction. ANG II infusion caused upregulation by 30% of the already significantly overexpressed B(1) receptors in cardiomyocytes of the B(2) receptor gene knockout mice, but in the wild-type mice it upregulated only the B(2) receptor mRNA by 47%. The addition of ANG II in VSMC culture produced a time-dependent induction of B(1) and upregulation of B(2) receptor gene expression, maximal at 3 h (by fivefold), declining almost to baseline by 24 h. The addition of losartan completely blocked this effect, whereas the AT(2) blocker PD-123319 made no difference, indicating that this is an AT(1)-mediated effect of ANG II. The data indicate that excess ANG II in subpressor doses in vivo upregulates expression of the B(2) receptor, but in its absence, the already overexpressed B(1) receptor is further upregulated, evidently assuming a counterregulatory response; in vitro, it transiently upregulates both bradykinin receptors.
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PMID:Effects of ANG II on bradykinin receptor gene expression in cardiomyocytes and vascular smooth muscle cells. 1155 71

The activation of soluble guanylate cyclase by bradykinin and sodium nitroprusside (SNP), a direct activator of soluble guanylate cyclase, was evaluated in androgen-sensitive LNCaP and androgen-independent PC3 and DU145 prostate cancer cells. Bradykinin and SNP activated soluble guanylate cyclase in LNCaP cells, but not in PC3 and DU145 cells. Western blot analysis revealed that the bradykinin B2 receptor, Gqalpha, phospholipase Cgamma and endothelial nitric oxide synthase were expressed in LNCaP, PC3 and DU145 cells. However, both Western blotting and reverse transcriptase--polymerase chain reaction indicated that soluble guanylate cyclase was only expressed in LNCaP cells. These results demonstrate that the impaired bradykinin-soluble guanylate cyclase pathway in PC3 and DU145 cells is likely due to lack of expression of soluble guanylate cyclase.
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PMID:The bradykinin/soluble guanylate cyclase signaling pathway is impaired in androgen-independent prostate cancer cells. 1182 65

We detected the expression of inducible bradykinin (BK) B1 receptor mRNA in the rat ileum by the reverse transcriptase-polymerase chain reaction (RT-PCR) method, when the isolated ileum was suspended for at least 1 hr in an aerated Tyrode's solution at 37 degrees. The induction of this mRNA was both time- and temperature-dependent, and was followed by a contractile response to des-Arg9-BK at around 3 hr of incubation; this response increased in magnitude with time and was maximal at 6 hr. In contrast, the contraction in response to BK and the expression of B2 receptor mRNA were constant throughout this 6-hr incubation period. The contraction due to des-Arg9-BK was selectively suppressed by B1 receptor antagonists, i.e. des-Arg9[Leu8]-BK and des-Arg10-HOE140, but not by the B2 antagonists D-Arg-[Hyp3,Thi5,8,D-Phe7]-BK and HOE140. The inducible des-Arg9-BK contractile response was suppressed by continuous in vitro exposure of the ileum to cycloheximide or actinomycin D, but neither inhibitor affected the contraction induced by BK, suggesting that the B1 receptor could be induced de novo. In vitro and ex vivo treatment of the ileum with dexamethasone suppressed the induction of the contractile response to des-Arg9-BK, but had no significant effect on the expression of B1 receptor mRNA. Some protein kinase C inhibitors, i.e. H7 and calphostin C, suppressed the expression of B1 receptor mRNA and diminished the contractile response to des-Arg9-BK. These results suggest that the de novo synthesis of the B1 receptor in the ileum preparation can be up-regulated at the transcriptional level (a process in which a specific isoform of protein kinase C may be involved). Additionally, these data suggest that the contractile response to des-Arg9-BK involves a process sensitive to some post-transcriptional action of dexamethasone.
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PMID:Effects of dexamethasone and protein kinase C inhibitors on the induction of bradykinin B1 mRNA and the bradykinin B1 receptor-mediated contractile response in isolated rat ileum. 1209 82

Vascular endothelial growth factor (VEGF) plays multifunctional roles in vascular permeability, repair and remodelling processes, in addition to the maintenance of vascular structure and function. In the present study, the potential of airway epithelial cell lines, BEAS-2B cells and A549 cells, to release and express VEGF in unstimulated and stimulated conditions was evaluated. The secretion and expression of VEGF were evaluated by enzyme-linked immunosorbant assay and by reverse transcriptase-polymerase chain reaction. The isoforms of released VEGF were determined by high-performance liquid chromatography. BEAS-2B cells and A549 cells released VEGF constitutively. Interleukin (IL)-1beta and tumour necrosis factor (TNF)-alpha augmented the release of VEGF in a time- and dose-dependent manner. The released VEGF was 165 amino acid residues in either condition. Pseudomonas aeruginosa lipopolysaccharide (LPS), interferon (IFN)-gamma, smoke extract (SE), neutrophil elastase (NE), and bradykinin stimulated the release of VEGF. Keracinocyte growth factor (KGF), which reduces vascular permeability, also stimulated both cells to release VEGF. VEGF messenger ribonucleic acid (mRNA) was expressed both time- and dose-dependently at 2 h, and declined after 2 h in response to IL-1beta and TNF-alpha. The expression of VEGF mRNA in airway epithelial cells was also augmented by LPS, IFN-gamma, SE, NE, and KGF stimulation. These data suggest that airway epithelial cells may regulate the maintenance of vascular structure and function, as well as vascular permeability, repair and remodelling processes, in a variety of lung conditions by expressing vascular endothelial growth factor.
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PMID:Vascular endothelial growth factor mRNA and protein expression in airway epithelial cell lines in vitro. 1250 3

We previously demonstrated that cyclic ADP-ribose (cADPR) elicits Ca2+ release in airway smooth muscle (ASM) cells through ryanodine receptor channels. CD38 is a cell surface protein that catalyzes the synthesis and degradation of cADPR. In inflammatory diseases such as asthma, augmented Ca2+ responses and Ca2+ sensitivity contribute to increased ASM contractility in response to agonists. In this study, we investigated the regulation of CD38 expression and the role of cADPR-mediated Ca2+ release in airway inflammation. Human ASM cells in culture between the second and fifth passages were exposed to tumor necrosis factor alpha (TNF-alpha), interleukin 1beta, or interferon gamma, or bovine serum albumin (controls). CD38 expression was measured by reverse transcriptase-polymerase chain reaction (RT-PCR), real-time PCR, and Western blot analysis, and ADP-ribosyl cyclase activity was assayed with nicotinamide guanine dinucleotide as the substrate. Ca2+ responses to acetylcholine, bradykinin, and thrombin were measured in fura-2AM-loaded cells by fluorescence microscopy. Cytokines caused significant augmentation of CD38 expression, ADP-ribosyl cyclase activity, and Ca2+ responses to the agonists, compared with the control. TNF-alpha effects were greater than those of the other two cytokines. The cADPR antagonist 8-bromo-cADPR attenuated the Ca2+ responses to the agonists in control and cytokine-treated cells, with the magnitude of inhibition correlating with the level of CD38. This study provides the first demonstration of a role for CD38-cADPR signaling in a model of inflammatory airway disease.
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PMID:CD38/cyclic ADP-ribose-mediated Ca2+ signaling contributes to airway smooth muscle hyper-responsiveness. 1251 17


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