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)

It has been proposed that capacitative Ca influx into both pancreatic acinar cells and HT-29 colonic cells is regulated by stimulation of nitric oxide synthase (NOS). NO, in turn, controls cGMP levels through effects on guanylate cyclase. We tested this possibility by measuring Ca (and Ba) entry into human embryonic kidney 293 cells and into 293 cells that had been transfected with the neuronal NOS gene (293/NOS). 293 cells had undetectable levels of NOS, while 293/NOS cells exhibited very high levels [Bredt D.S., Ferris C.D., Snyder S.H. Nitric oxide synthase regulatory sites. J Biol Chem 1992; 267: 10976-10981]. Ca (or Ba) entry into single cells was measured as the rate of increase of the Fura-2 fluorescence ratio (digital imaging microscopy) during rapid changes from Ca-free (or Ba-free) to Ca- (or Ba-) containing solutions (using high K to depolarize the membrane potential). cGMP levels (EIA method) were measured to correlate to rates of Ca entry. 100 microM ATP caused release of Ca from internal stores, but no sustained plateau due to Ca entry in either 293 or 293/NOS cells. Cyclopiazonic acid (CPA, which inhibits the Ca pump of the internal store, allowing Ca to leak from the store) caused apparent Ca entry to increase 5-10-fold from similar, low levels in both 293 and 293/NOS cells. CPA-stimulated Ca entry was unaffected by the NOS inhibitor N-nitro-L-arginine (L-NA) in either 293 or 293/NOS cells. In 293 cells [cGMP] was low; ATP and CPA both increased [cGMP] by 2-fold, and the guanylate cyclase inhibitor LY83583 and L-NA decreased [cGMP] by 50-75%. [cGMP] was 20-fold higher in 293/NOS cells than in 293 cells; these [cGMP] were not affected by ATP and CPA, but were effectively decreased by 80-90% by L-NA and by LY83583. Thus, [cGMP] and Ca or Ba entry showed no relationship to each other: Ca entry was small into cells in which [cGMP] was either low (resting 293, CPA + L-NA or CPA + LY83583), intermediate (ATP-treated 293) or high (resting 293/NOS). Similarly, Ca entry was high into cells in which [cGMP] was low (CPA + L-NA- or CPA + LY83583-treated 293), intermediate (CPA-treated 293 and CPA + L-NA-treated 293/NOS) or high (CPA- or ATP-treated 293/NOS). We conclude that, as in most other non-excitable cells, Ca entry into 293 cells is stimulated by loss of Ca from the store but, unlike pancreatic and colonic cells, this capacitative Ca entry does not appear to be regulated by NO and cGMP. Therefore, although capacitative entry across the plasma membrane may be regulated by NO and cGMP in Gl epithelial cells, this regulation does not occur in all cells.
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PMID:Does nitric oxide regulate capacitative Ca influx in HEK 293 cells? 913 96

1. In airway epithelium, nitric oxide (NO) is synthesized in the setting of inflammation by inducible nitric oxide synthase (iNOS). Although the role of epithelial derived NO in the regulation of human airways is unknown, prostaglandin E2 (PGE2) is recognised as an important inhibitory mediator in human airways. Cyclo-oxygenase (COX) is the rate limiting enzyme in the production of prostanoids and since inflammatory pathways enhance the expression of an inducible COX (COX-2), both COX-2 and iNOS may be co-expressed in response to an inflammatory stimulus. Although regulation of the COX-2 pathway by NO has been demonstrated in animal models, its potential importance in human airway epithelium has not been investigated. 2. The effect of endogenous and exogenous NO on the COX-2 pathway was investigated in the A549 human airway epithelial cell culture model. Activity of the COX-2 pathway was assessed by PGE2 EIA, and iNOS pathway activity by nitrite assay. A combination cytokine stimulus of interferon gamma (IFNgamma) 100 u ml(-1), interleukin-1beta (IL-1beta) 1 u ml(-1) and lipopolysaccharide (LPS) 10 microg ml(-1) induced nitrite formation which could be inhibited by the competitive NOS inhibitor N(G)-nitro-L-arginine-methyl-ester (L-NAME). IL-1beta alone (1-50 u ml(-1) induced PGE2 formation without significant nitrite formation, a response which was inhibited by the COX-2 specific inhibitor nimesulide. Submaximal stimuli used for further experiments were IFNgamma 100 u ml(-1), IL-1beta 1 u ml(-1) and LPS 10 microg ml(-1) to induce both the iNOS and COX-2 pathways, and IL-1beta 3 u ml(-1) to induce COX-2 without iNOS activity. 3. Cells treated with IFNgamma 100 u ml(-1), IL-1beta I u ml(-1) and LPS 10 microg ml(-1) for 48 h either alone, or with the addition of L-NAME (0 to 10(-2) M), demonstrated inhibition by L-NAME of PGE2 (3.61 +/- 0.55 to 0.51 +/- 0.04 pg/l0(4) cells; P<0.001) and nitrite (34.33 +/- 8.07 to 0 pmol/10(4) cells; P<0.001) production. Restoration of the PGE2 response (0.187 +/- 0.053 to 15.46 +/- 2.59 pg/10(4) cells; P<0.001) was observed after treating cells with the same cytokine stimulus and L-NAME 10(-6) M, but with the addition of the NOS substrate L-arginine (0 to 10(-5) M). 4. Cells incubated with IL-1beta 3 u ml(-1) for 6 h, either alone or with addition of the NO donor S-nitroso-acetyl-penicillamine (SNAP) (0 to 10(-4) M), demonstrated increased PGE2 formation (1.23 +/- 0.03 to 2.92 +/- 0.19 pg/10(4) cells; P< 0.05). No increase in PGE2 formation was seen when the experiment was repeated in the presence of the guanylate cyclase inhibitor methylene blue (50 microM). Cells treated with SNAP alone did not demonstrate an increased PGE2 formation. Cells incubated with IL-1beta 3 u ml(-1) for 6 h in the presence of dibutyryl cyclic guanylate monophosphate (0 to 10(-3) M) also demonstrated an increased PGE2 response (2.56 +/- 0.21 to 4.53 +/- 0.64 pg/10(4) cells; P<0.05). 5. These data demonstrate that in a human airway epithelial cell culture system, both exogenous and endogenous NO increase the activity of the COX-2 pathway in the setting of inflammatory cytokine stimulation, and that this effect is likely to be mediated by guanylate cyclase. This suggests a role for NO in the regulation of human airway inflammation.
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PMID:Regulation of the inducible cyclo-oxygenase pathway in human cultured airway epithelial (A549) cells by nitric oxide. 925 31

Endogenous carbon monoxide (CO) shares with nitric oxide (NO) a role as a putative neural messenger in the brain. Both gases are believed to modulate CNS function via an increase in cytoplasmic cGMP concentrations secondary to the activation of soluble guanylate cyclase (sGC). Recently CO and NO were proposed as a possible mediator of febrile response in hypothalamus. NO has been reported to activate both the constitutive and inducible isoform of the cyclooxygenase (COX). Thus, we investigated whether CO arising from heme catabolism by heme oxygenase (HO) is involved in the febrile response via the activation of COX in the hypothalamus. PGE2 which is a final mediator of febrile response released from primary cultured hypothalamic cells was taken as a marker of COX activity. PGE2 concentration was measured with EIA kits. Exogenous CO (CO-saturated medium) and hemin (a substrate and potent inducer of HO) evoked an increase in PGE2 release from hypothalamic cells, and these effects were blocked by methylene blue (an inhibitor of sGC). And membrane permeable cGMP analogue, dibutyryl-cGMP elicited significant increases in PGE2 release. These results suggest that there may be a functional link between HO and COX enzymatic activities. The gaseous product of hemin through the HO pathway, CO, might play a role through the modulation of the COX activity in the hypothalamus.
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PMID:Effects of heme oxygenase system on the cyclooxygenase in the primary cultured hypothalamic cells. 1179 44