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:4.6.1.2 (guanylate cyclase)
8,497 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cells isolated from the trabecular meshwork (TM) of a male glaucoma patient were transformed by transfection with an origin defective mutant of SV40 virus. Transformation dramatically increased the growth rate of these cells (designated HTM-3 cells), allowing biochemical and pharmacological characterization. The HTM-3 cells had cytoskeletal components that were reported to be present in TM tissue and non-transformed TM cells. Vimentin, tubulin and smooth muscle specific alpha-actin, but not desmin, were localized in these cells by immunocytochemistry. The extracellular matrix components collagen types I, III and IV, fibronectin and laminin were found in HTM-3 cells as well as their non-transformed parental cells. As predicted, the protein profile of the HTM-3 cells revealed by two-dimensional gel electrophoresis was different from that of the non-transformed cells, probably due to the enhanced growth characteristics of these cells. Furthermore, HTM-3 cells had various intracellular second messenger systems that responded to pharmacological agents. Forskolin, prostaglandin E2, beta-adrenergic and adenosine A2 agonists stimulated the adenylyl cyclase in these cells, whereas muscarinic, serotonergic, dopaminergic and other agonists were ineffective. Sodium nitroprusside increased the intracellular concentration of cGMP, demonstrating the presence of a functional guanylyl cyclase. Phospholipase C activity in these cells was also detected. Muscarinic agonists, histamine and bradykinin, but not adrenergic, serotonergic agonists or prostaglandins, increased phosphoinositide turnover. These drug responses of HTM-3 cells agree with published data on primary TM cells and TM tissues, suggesting that the transformed cells may be a valid substitute for certain pharmacological studies of TM.
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PMID:Preliminary characterization of a transformed cell strain derived from human trabecular meshwork. 815 26

Thromboxane (TX) stimulation of fibronectin (FN) synthesis in mesangial cells (MC) is dependent on protein kinase C (PKC)-mediated increases in transforming growth factor beta (TGF beta), and is suppressed by increases in cellular cGMP. The current studies evaluate the role of cGMP-dependent and -independent actions of nitric oxide (NO) in modulating the responses of MC to the TX analogue U46619. TX-stimulated increases in PKC activity, TGF beta, and FN synthesis in MC were suppressed by either 8-Br-PET-cGMP or the NO donor S-nitroso-N-acetylpenicillamine (SNAP). By contrast, NO, but not cGMP, inhibited basal PKC activity, TGF beta bioactivity and FN synthesis. The cGMP-dependent protein kinase 1-alpha inhibitor 8-(4-chlorophenylthio)guanosine 3',5'-cyclic monophosphorothioate (Rp) restored the PKC, TGF beta, and the FN synthetic responses to TX when added to MC before exposure of the cells to either cGMP or SNAP. However, neither Rp nor the guanylate cyclase inhibitor Ly83583 significantly altered SNAP inhibition of basal PKC. In addition, Rp failed to alter the decreases in basal TGF beta bioactivity and FN synthesis seen in the presence of SNAP. In contrast to the FN response to U46619, cGMP and SNAP did not affect the stimulation of FN synthesis by exogenous TGF beta. The later findings are consistent with inhibitory actions of NO and cGMP at, or proximal to, U46619-induced increases in TGF beta in the suppression of TX-signaled increases in FN synthesis. Thus, NO depresses basal PKC and TGF beta bioactivity in MC by mechanisms that are largely independent of cGMP, whereas NO inhibition of these MC responses to TX is mediated primarily by increases in cGMP and activation of protein kinase 1-alpha.
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PMID:Nitric oxide suppresses increases in mesangial cell protein kinase C, transforming growth factor beta, and fibronectin synthesis induced by thromboxane. 882 14

Animal and clinical investigations have reported that exposure to hyperbaric O(2) improved the outcome of some reperfusion injuries. Animal studies have suggested that this may be due to an inhibition of leukocyte adherence to injured endothelium. This investigation tested the hypothesis that exposure to hyperbaric O(2) would inhibit beta2-integrin-dependent adherence of human neutrophils. Subjects were exposed to O(2) at partial pressures of up to 3 atmospheres absolute (ATA; 1 ATA = 0.1 MPa) for 45 min, and neutrophil binding to nylon columns and to fibrinogen-coated surfaces was measured. Exposure to O(2) at 2.8 or 3.0 ATA inhibited beta2-integrin-dependent neutrophil adherence but had no effect on the cell-surface expression of beta2-integrins, respiratory burst in response to phorbol ester, or non-beta2-integrin-dependent adherence to plastic plates coated with a fibronectin-like protein. beta2-Integrin adherence was restored by incubating blood with 8-bromoguanosine 3',5'-cyclic monophosphate (cGMP) and hyperbaric O(2) inhibited synthesis of cGMP by neutrophils stimulated with N-formyl-Met-Leu-Phe (FMLP). In studies of cell fractions, the activity of membrane guanylate cyclase was found to be increased by incubation with FMLP as well as by atrial natriuretic peptide (ANP) plus ATP. Hyperbaric O(2) had no effect on the basal activity of soluble or membrane-bound guanylate cyclase. However, hyperbaric O(2) inhibited the function of both the extracellular binding domain of membrane guanylate cyclase as well as intracellular catalytic activity. There are approximately 7,300 membrane guanylate cyclase molecules per cell, based on binding studies with ANP, with a dissociation constant of approximately 450 pM. Hyperbaric O(2) inhibits the function of human neutrophil beta2-integrins by a process linked to impaired synthesis of cGMP.
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PMID:Inhibition of human neutrophil beta2-integrin-dependent adherence by hyperbaric O2. 912 10

Interactions of mesangial cells (MCs) with components of the extracellular matrix (ECM) profoundly influence the MC phenotype, such as attachment, contraction, migration, survival and proliferation. Here, we investigated the effects of exogenous nitric oxide (NO) on the process of MC adhesion to ECM molecules. Incubation of rat MCs with the NO donor S-nitroso-N-acetylpenicillamine (SNAP) dose- and time-dependently inhibited MC adhesion and spreading on various ECM substrata, being more pronounced on collagen type I than on collagen type IV, laminin or fibronectin. In contrast, SNAP did not inhibit MC adhesion to L-polylysine-coated plates. The inhibitory effects of SNAP were reduced by hemoglobin and enhanced by superoxide dismutase. The anti-adhesive action of SNAP was mimicked not only by other NO donors but also by 8-bromo-cGMP, and significantly reversed by the soluble guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3,-alpha]quinoxalin-1-one (ODQ). Moreover, SNAP and 8-bromo-cGMP decreased the adhesion-induced phosphorylation of focal adhesion kinase (pp125FAK). In the presence of SNAP or 8-bromo-cGMP, adherent MCs exhibited disturbed organization of alpha-actin filaments and reduced numbers of focal adhesions, as shown by immunocytochemistry. In additional experiments with adherent MCs, it was found that exposure to SNAP or 8-bromo-cGMP for 12 and 24 hours induced detachment of MCs. The results indicate that exogenous NO interferes with the establishment and maintenance of MC adhesion to ECM components. This inhibitory NO effect is mediated predominantly by cGMP-signaling. Disturbance of MC attachment to ECM molecules could represent an important mechanism by which NO affects MC behavior in vitro and in vivo.
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PMID:Exogenous nitric oxide inhibits mesangial cell adhesion to extracellular matrix components. 950 4

We examined the effect of nitric oxide (NO) on cell adhesion using cultured human pulmonary microvascular endothelial cells (PMVEC). Attachment of these cells to fibronectin was significantly inhibited by NO donors, spermine NONOate and S-nitroso-N-acetyl-penicillamine or L-arginine, but not 8-bromoguanosine-3',5'-cyclic-monophosphate. Similar results were obtained with the electrical cell-substrate impedance sensor (ECIS) technique. Addition of NO donors or L-arginine, but not 8-bromoguanosine-3',5'-cyclic-monophosphate or N2,2'-O-dibutyrylguanosine-3',5'-cyclic-monophosphate, to confluent PMVEC monolayers resulted in a transient decrease in cell adhesion, which was quantitated by the ECIS. Exposure to 1 U/ml alpha-thrombin reduced the monolayer electrical resistance by approximately 50%. The observed response was significantly suppressed by pretreatment of cells with intracellular calcium chelator, 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid or NO synthase inhibitor, N(G)-nitro-L-arginine methyl ester, but not guanylate cyclase inhibitor, 6-anilino-5,8-quinoline-quinone. Selective knockout of endothelial NO synthase with antisense oligodeoxynucleotides also significantly reduced thrombin-induced decrease in monolayer resistance. Our findings indicate that thrombin stimulates calcium-dependent release of NO from PMVEC, which mediates the retraction of endothelial cells via a cGMP-independent pathway. Our results suggest that NO modulates cell-matrix and/or cell-cell adhesion in PMVEC and that this molecule might modify microvascular permeability in the human lung.
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PMID:Role of nitric oxide in human pulmonary microvascular endothelial cell adhesion. 1089 23

1. The nitric oxide synthase (NOS) inhibitor, N(omega)-nitro-L-arginine methyl ester (L-NAME), inhibits both rat and human eosinophil chemotaxis in vitro. Here, the role of nitric oxide (NO) in human eosinophil cell surface integrin expression and function was investigated. 2. Human peripheral blood eosinophils were treated with L-NAME (0.01 - 1.0 mM) and their adhesion to human fibronectin and serum observed. Adhesion of cells to fibronectin and serum increased by 24.0+/-4.6 and 43.8+/-4.7%, respectively, when eosinophils were treated with 1.0 mM L-NAME. Increased adhesion by L-NAME could be abolished when cells were co-incubated with VLA-4- and Mac-1-specific monoclonal antibodies (mAbs). 3. The NO donor, sodium nitroprusside (2.5 mM), significantly inhibited eosinophil adhesion to fibronectin and serum by 34.3+/-4.5 and 45.2+/-5.6%, respectively. This inhibition was accompanied by a 4 fold increase in the levels of intracellular cyclic GMP. 4. Flow cytometrical analysis demonstrated that L-NAME induced an increased expression of CD11b (Mac-1) on the eosinophil cell surface of 36.3+/-7.4%. L-NAME had no effect upon CD49d (VLA-4) expression. 5. Treatment of human eosinophils, in vitro, with H-[1,2,4] oxadiazolo quinoxalin-1-one (ODQ) (0.1 mM), an inhibitor of soluble guanylate cyclase, also significantly increased eosinophil adhesion to fibronectin and serum by 73.5+/-17.9 and 91.7+/-12.9%, respectively. This increase in adhesion could also be inhibited by co-incubation with the Mac-1 and VLA-4-specific mAbs. 6. In conclusion, results indicate that NO, via a cyclic GMP-dependent mechanism, inhibits the adhesion of human eosinophils to the extracellular matrix (ECM). This inhibition is accompanied by a decrease in the expression and function of the eosinophil's adhesion molecules, in particular, the expression of the Mac-1 integrin and the function of the VLA-4 integrin.
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PMID:Nitric oxide regulates human eosinophil adhesion mechanisms in vitro by changing integrin expression and activity on the eosinophil cell surface. 1158 18

Nitric oxide is an important messenger that regulates mast cell activity by modifications to gene expression and intracellular pathways associated with exocytosis and adhesion. Integrin interactions with extracellular matrix components modulate an array of cell activities, including mediator production and secretion. To investigate the molecular mechanisms underlying NO regulation of mast cell function, we studied its effects on adhesion of a human mast cell line (HMC-1) to fibronectin (FN). The NO donors S-nitrosoglutathione and S-nitroso-N-acetylpenicillamine strongly down-regulated the adhesion of HMC-1 to FN. Inhibitors of soluble guanylate cyclase and protein kinase G did not alter the response of cells to NO. A peroxynitrite scavenger did not affect modulation of adhesion by NO, nor could the effect of NO be mimicked by the peroxynitrite-producing compound 3-morpholinosydnonimine. NO donors inhibited the cysteine protease, calpain, while calpain inhibitors mimicked the effect of NO and led to a decrease in the ability of HMC-1 cells to adhere to FN. Thus, NO is an effective down-regulator of human mast cell adhesion. The mechanism for this action does not involve peroxynitrite or activation of soluble guanylate cyclase. Instead, a portion of NO-induced down-regulation of adhesion may be attributed to inhibition of the cysteine protease, calpain, an enzyme that has been associated with control of integrin activation in other cell types. The inhibition of calpain is most likely mediated via nitrosylation of its active site thiol group. Calpain may represent a novel therapeutic target for the regulation of mast cell activity in inflammatory disorders.
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PMID:Inhibition of calpain is a component of nitric oxide-induced down-regulation of human mast cell adhesion. 1249 11

Idiopathic pulmonary fibrosis is an incurable fibrosing disorder that progresses relentlessly to respiratory failure. We hypothesized that a product of heme oxygenase activity, carbon monoxide (CO), may have anti-fibrotic effects. To test this hypothesis, mice treated with intratracheal bleomycin were exposed to low-concentration inhaled CO or ambient air. Lungs of mice treated with CO had significantly lower hydroxyproline accumulation than controls. Fibroblast proliferation, thought to play a central role in the progression of fibrosis, was suppressed by in vitro exposure to CO. CO caused increased cellular levels of p21(Cip1) and decreased levels of cyclins A and D. This effect was independent of the observed suppression of MAPK's phosphorylation by CO but was dependent on increased cGMP levels. Further, CO-exposed cells elaborated significantly less fibronectin and collagen-1 than control cells. This same effect was seen in vivo. Suppression of collagen-1 production did not depend on MAPK or guanylate cyclase signaling pathways but did depend on the transcriptional regulator Id1. Taken together, these data suggest that CO exerts an anti-fibrotic effect in the lung, and this effect may be due to suppression of fibroblast proliferation and/or suppression of matrix deposition by fibroblasts.
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PMID:Carbon monoxide suppresses bleomycin-induced lung fibrosis. 1563 97

cGMP serves as the main second messenger of nitric oxide (NO). Antifibrotic effects of enhancing renal cGMP levels have recently been documented in experimental acute anti-Thy-1 glomerulonephritis. The present study compares the effects of the cGMP production-increasing soluble guanylate cyclase (sGC) stimulator BAY 41-2272 with those of the cGMP degradation-limiting phosphodiesterase inhibitor pentoxifylline (PTX) in a progressive model of renal fibrosis. At 1 wk after induction of anti-Thy-1-induced chronic glomerulosclerosis (cGS), rats were randomly assigned to groups as follows: cGS, cGS + BAY 41-2272 (10 mg x kg body wt(-1) x day(-1)), or cGS + PTX (50 mg x kg body wt(-1) x day(-1)). BAY 41-2272 and PTX reduced systolic blood pressure significantly. At 16 wk, tubulointerstitial expressions of sGC mRNA and NO-induced cGMP synthesis were increased in untreated cGS animals, whereas their glomerular activity was depressed compared with normal controls. Tubulointerstitial and glomerular cGMP production in response to NO were significantly enhanced in animals treated with BAY 41-2272, but not in those treated with PTX. BAY 41-2272 administration resulted in marked reductions of glomerular and tubulointerstitial histological matrix accumulation, expression of TGF-beta1 and fibronectin, macrophage infiltration, and cell proliferation as well as improved renal function. In contrast, only moderate and nonsignificant renoprotective changes were observed in the cGS + PTX group. In conclusion, increasing renal cGMP production through BAY 41-2272 significantly improved renal NO-cGMP signaling and limited progression in anti-Thy-1-induced chronic renal fibrosis, whereas inhibition of cGMP degradation by PTX was only moderately effective. The findings indicate that pharmacological enhancement of renal cGMP levels by sGC stimulation represents a novel and effective antifibrotic approach in progressive kidney disorders.
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PMID:Enhancing cGMP in experimental progressive renal fibrosis: soluble guanylate cyclase stimulation vs. phosphodiesterase inhibition. 1604 4

Increased leukocyte adhesion to vascular endothelium contributes to vaso-occlusion in sickle cell disease. Since nitric oxide bioavailability is decreased in sickle cell disease and nitric oxide may inhibit leukocyte adhesion, we investigated whether stimulation of NO-signaling pathways can reduce the adhesive properties of neutrophils from sickle cell disease individuals (sickle cell diseaseneu). sickle cell diseaseneu presented greater adhesion in vitro to both fibronectin and ICAM-1 than control neutrophils. Co-incubation of sickle cell diseaseneu with the nitric oxide-donor agents, sodium nitroprusside and dietheylamine NONOate (DEANO), and the guanylate cyclase stimulator, BAY41-2272, all significantly reduced the increased adhesion to fibronectin/ICAM-1. Oxadiazolo[4,3-a]quinoxalin-1-one, a guanylate cyclase inhibitor, reversed sodium nitroprusside/DEANO-diminished adhesion to fibronectin, implicating cGMP-dependent signaling in this mechanism. Interestingly, intracellular cGMP was significantly higher in neutrophils from sickle cell disease individuals on hydroxyurea (sickle cell diseaseHUneu). Accordingly, sickle cell diseaseHUneu adhesion to fibronectin/ICAM-1 was significantly lower than that of sickle cell diseaseneu. Agents that stimulate the nitric oxide/cGMP-dependent pathway may have beneficial effects on leukocyte function if used in these subjects.
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PMID:Increased adhesive properties of neutrophils in sickle cell disease may be reversed by pharmacological nitric oxide donation. 1832 23


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