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:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cloned BK channel alpha subunit from human myometrium was stably expressed in Chinese hamster ovary cells, either alone (CHOalpha cells) or in combination with the auxiliary beta subunit (CHOalpha+beta cells). We studied basic channel properties and the effects of cGMP- and cAMP-dependent protein kinases on the BK channel activity. Coexpression of alpha and beta subunits enhanced the Ca2+ and voltage sensitivity of the BK channel, and decreased the inhibitory potency of iberiotoxin. Blocking and stimulating effects on BK channel activity by charybdotoxin and nitric oxide, respectively, were independent of the beta subunit. The cGMP kinase Ialpha and cAMP kinase failed to affect BK channel activity in CHOalpha and CHOalpha+beta cells at different [Ca2+]i and voltages. In contrast, BK channels in freshly isolated myometrial cells from postmenopausal women responded to cAMP kinase and cGMP kinase with a fourfold and twofold decrease in their open probability (NPo), respectively. These effects could be reversed by alkaline phosphatase and remained unaffected by the phosphatase inhibitor okadaic acid (100 nM). In 28% of myometrial cells, however, cAMP and cGMP kinases increased NPo 2-fold and 3.5-fold, respectively. This stimulation was enhanced rather than reversed by alkaline phosphatase and was abolished by 100 nM okadaic acid. The results suggest that in stably transfected CHO cells the expressed BK channel is not regulated by cAMP kinase and cGMP kinase. However, in native myometrial cells stimulatory and inhibitory regulation of BK channels by cAMP kinase and cGMP kinase was observed, suggesting that channel regulation by the protein kinases requires factors that are not provided by CHO cells. Alternatively, failure of regulation may have been due to the primary structure of the myometrial BK channel protein used in this study.
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PMID:Regulation of stably expressed and native BK channels from human myometrium by cGMP- and cAMP-dependent protein kinase. 971 6

Amenorrheic athletes have been likened to postmenopausal women, with low estrogen levels and osteopenia. It has been suggested that estrogen exerts its antiresorptive actions on bone via a nitric oxide (NO)-dependent mechanism. This study investigated whether the mechanism of bone loss in amenorrheic athletes is similar to that of postmenopausal women with reduced NO levels and high bone turnover. Eleven amenorrheic athletes, 15 eumenorrheic athletes, and 10 sedentary controls were studied. Spine and hip bone mineral density was measured using dual-energy x-ray absorptiometry. Bone turnover was assessed by biochemical markers of formation (osteocalcin and bone-specific alkaline phosphatase) and resorption (deoxypyridinoline). NO metabolites were measured from 24-h urine samples using a chemiluminescence assay. Spine, but not hip, bone mineral density was reduced in the amenorrheic group, compared with the eumenorrheic (P = 0.0001) and control (P = 0.04) groups. Osteocalcin, bone-specific alkaline phosphatase, and deoxypyridinoline were similar in all groups. NO metabolites were lower in the amenorrheic group, compared with controls (P = 0.035), despite a higher dietary intake of nitrates. Unlike postmenopausal women, amenorrheic athletes do not have raised bone turnover but do have reduced NO metabolites and spinal osteopenia. The results show, however, that reduced NO production is a common denominator in both conditions and further support the importance of NO in estrogen-mediated protection of skeletal mass and strength.
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PMID:Decreased nitric oxide levels and bone turnover in amenorrheic athletes with spinal osteopenia. 974 3

Overproduction of tumor necrosis factor (TNF-), interleukin-1beta (IL-1beta), and nitric oxide (NO) is believed to be detrimental during the progression of acute pancreatitis, yet little is known about the hepatic production of these mediators and their role in mediating pancreatitis-induced hepatic dysfunction. Rats were randomized to receive a single intraperitoneal injection of the macrophage-pacifying compound, CNI-1493 (1.0 mg/kg), or vehicle 1 hour before the induction of retrograde bile salt pancreatitis. Sham-operated animals served as controls. Animals were killed 18 hours later, with serum and livers harvested to determine the degree of hepatocellular injury and the induction of TNF-, IL-1beta, and inducible nitric oxide synthase (iNOS). In addition, serum TNF- and nitrites (end-product of NO breakdown) were determined in each group to assess the mechanism of action of CNI-1493. TNF-, IL-1beta, and iNOS gene expression (by reverse-transcription polymerase chain reaction) as well as aspartate transaminase (AST), alanine transaminase (ALT), and lactic dehydrogenase (LDH) (but not alkaline phosphatase [ALP]) increased following the development of pancreatitis (all P < .05). Macrophage pacification significantly prevented the induction of TNF- and IL-1beta mRNA (but not iNOS), resulting in lessened serum AST, ALT, and LDH (all P < .05). Serum TNF- protein and nitrites correlated with gene induction in that both were increased following the onset of pancreatitis, and TNF- protein production was significantly attenuated in animals receiving CNI-1493. Hepatocellular, but not bile duct, injury occurs during experimental pancreatitis that is associated with hepatic TNF-, IL-1beta, and iNOS mRNA gene induction, as well as TNF- protein and nitrite production. Preventing the production of TNF- and IL-1beta by macrophage pacification attenuates the hepatocellular damage, suggesting that these mediators play a role in pancreatitis-induced hepatic injury.
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PMID:Macrophage pacification reduces rodent pancreatitis-induced hepatocellular injury through down-regulation of hepatic tumor necrosis factor alpha and interleukin-1beta. 979 13

Previous studies have shown evidence of constitutive and cytokine-inducible nitric oxide (NO) synthase activity in cultured osteoblast-like cells from various species. Although cytokine-induced NO production has been found to inhibit osteoblast growth, the role of constitutive NO production in regulating osteoblast function is less clear and the isoforms of nitric oxide synthase (NOS) that are expressed by human osteoblasts have not been determined. Here, we investigated NOS expression in cultured human osteoblast-like cells and studied the effects of constitutive and cytokine-induced NO on osteoblast growth and differentiation. Low levels of NO were produced constitutively by osteoblast-like cells as reflected by analysis of medium nitrite concentrations, and evidence of ecNOS mRNA, protein, and bioactivity was found in primary osteoblasts (hOBs), TE85, and MG63 osteosarcoma cells. None of the osteoblast-like cells expressed nNOS, however, and iNOS was produced only by hOB cells after stimulation with the cytokines IL-1beta, TNF-alpha, and IFN-gamma. The NOS inhibitor, L-NMMA, did not affect growth or alkaline phosphatase activity in unstimulated osteoblasts. Incubation of hOB cells with cytokines inhibited growth and stimulated alkaline phosphatase activity and these effects were abrogated by L-NMMA. Cytokines also inhibited growth of TE85 cells and MG63 cells, but these effects appeared to be NO independent because they were not influenced by L-NMMA. Our experiments show that human osteoblasts constitutively produce NO through the ecNOS pathway, but demonstrate that this does not appear to exert an appreciable effect on osteoblast growth or differentiation under basal conditions. In contrast, IL-1beta, TNF-alpha, and IFN-gamma exerted growth-inhibiting and differentiation-inducing effects on osteoblasts that were partly NO dependent, indicating that NO may act predominantly as a modulator of cytokine-induced effects on osteoblast function.
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PMID:Expression and functional role of nitric oxide synthase isoforms in human osteoblast-like cells. 1007 9

Shear stress causes release of nitric oxide (NO) from microvascular endothelial cells in vivo and stimulates their growth in vitro. After chronic electrical stimulation of lower hind limb skeletal muscles in the rat, measurements of capillary diameters and red blood cell velocity indicated that shear stress is increased in these vessels as a potential source of NO. This study therefore investigated whether NO is involved in capillary growth in stimulated muscles. Control rats or those stimulated for 2 or 7 days were treated with the NO synthase inhibitor, N(G)-nitro-l-arginine (l-NNA, 10 mg.day(-1) in drinking water), or water alone. After bromodeoxyuridine (BrdU) administration, extensor digitorum longus muscles were removed and frozen. Capillary supply was assessed in cryostat sections as capillary:fiber (C:F) ratio after staining for alkaline phosphatase; proliferation of capillary-linked and interstitial nuclei was evaluated by immunostaining for BrdU incorporation. C:F was not increased after 2 days of stimulation but the increase after 7 days (1.88 +/- 0.50 vs control 1.45 +/- 0.04, P < 0.001) was abolished by l-NNA (1.55 +/- 0.04, NS). The labeling index for BrdU-positive nuclei colocalized with capillaries as a percentage of total interstitial nuclei increased in muscles stimulated for 2 days (11.3 +/- 2.2%) and 7 days (10.6 +/- 0.8%) compared with controls (2.9 +/- 0.5%, P < 0.01) and was eliminated by l-NNA at both time points (3.1 +/- 0.6 and 1.0 +/- 0.6%, respectively; both P < 0.05 vs stimulated). A transient increase in BrdU labeling of interstitial nuclei not associated with capillaries (possibly fibroblasts) after 2 but not 7 days stimulation was eliminated by l-NNA treatment. These results suggest that NO is involved in capillary growth in chronically stimulated muscles possibly via its shear-stress-induced release from capillaries or from interstitial fibroblasts.
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PMID:Inhibition of capillary growth in chronically stimulated rat muscles by N(G)-nitro-l-arginine, nitric oxide synthase inhibitor. 1062 70

To determine the roles of nitric oxide (NO) and its metabolite, peroxynitrite (ONOO(-)), on osteoblastic activation, we investigated the effects of a NO donor [ethanamine, 2, 2'-(hydroxynitrosohydrazono)bis- (dNO)], an O(-2) donor (pyrogallol), and an ONOO(-) scavenger (urate) on alkaline phosphatase (ALPase) activity and osteocalcin gene expression, which are indexes of osteoblastic differentiation. dNO elevated ALPase activity in the osteogenic MC3T3-E1 cell line. The combination of dNO and pyrogallol reduced both ALPase activity and osteocalcin gene expression. Because both indexes were recovered by urate, ONOO(-), unlike NO itself, inhibited the osteoblastic differentiation. Furthermore, treatment with a combination of the proinflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) was found to yield ONOO(-) as well as NO and O(-2). The reductions in ALPase activity and osteocalcin gene expression were also restored by urate. We conclude that ONOO(-) produced by TNF-alpha and IL-1beta, but not NO per se, would overcome the stimulatory effect of NO on osteoblastic activity and inhibit osteoblastic differentiation.
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PMID:Peroxynitrite production by TNF-alpha and IL-1beta: implication for suppression of osteoblastic differentiation. 1082 5

Previous studies have shown that, in unstimulated mammary epithelial cells from virgin mice, prolactin receptors are retained intracellularly because of their incomplete N-glycosylation. Activation of the nitric oxide/cGMP pathway stimulates Nacetylglucosamine (NAG) transferase I activity, completion of terminal glycosylation, and redistribution of the receptors to the cell surface. In this study, it was shown that nitric oxide could stimulate the phosphorylation of NAG transferase I in intact cells and that the cGMP-dependent protein kinase (PKG) could directly phosphorylate the purified enzyme. Furthermore, this modification was associated with enhanced enzymatic activity. Conversely, this stimulation of activity was blocked in intact cells by coincubation with a PKG inhibitor and reversed in the immunoprecipitated enzyme by alkaline phosphatase treatment. Kinetic analysis revealed that this effect on enzyme activity was due to an increase in V(max) without any change in K(m). Therefore, it appears that the nitric oxide/cGMP pathway activates NAG transferase I via direct phosphorylation by PKG.
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PMID:Rapid hormonal regulation of N-acetylglucosamine transferase I. 1082 30

Recently, we showed that supplementation with nitric oxide (NO) via donor nitroglycerin (NG) alleviated the ovariectomy and corticosteroid-induced bone loss in rats. In humans, high doses or frequent applications of NG (i.e., for angina) lead to rapid loss of its efficacy in relieving angina. To examine whether there is a similar effect on the loss of efficacy of NG on bone, we examined the frequency-dependent effects of NG on bone mineral density (BMD), bone mass, trabecular bone volumes (BV/TV), and blood pressure in rats. Thirty 7-month-old female Brown Norway rats underwent ovariectomy, and an additional six rats were sham-operated. The ovariectomized rats were treated either with vehicle (ovariectomized control), 17beta-estradiol (E2; positive control), or 0.2 mg NG (via dermal application) once, twice, or three times a day. Before and at the end of the 10-week treatment period, BMD of the lumbar spine was measured by dual-energy X-ray absorptiometric (DXA) scanning and expressed as a percentage change. BMD in ovariectomized rats was significantly lower (-2.5 +/- 2.0%) compared with the sham-operated rats (+6.3 +/- 5.3%; p < 0.01). Estrogen therapy completely abolished the ovariectomy-induced potential bone loss (+5.9 +/- 3.4%). Application of NG once daily also completely prevented (+6.2 +/- 2.8%; p < 0.01) the ovariectomy-induced bone loss (i.e., it was as effective as estrogen). However, the beneficial effects of NG on BMD were significantly reduced with increased frequency of application of NG (+1.9 +/- 2.1%, twice a day and -0.2 +/- 3.3% three times a day). Estrogen or once daily administration of NG preserved femur weights, BV/TV, and decreased urinary deoxypyridinoline levels as expected. However, a higher level of serum osteocalcin and bone-specific alkaline phosphatase levels were maintained only with once daily administration of NG. There were no adverse effects of these doses of NG on blood pressure, but a tendency to lower blood pressure was noticed with increased frequency of NG. These results confirmed our previous findings that NO donors counteract the bone loss associated with estrogen deficiency. However, these beneficial effects of maintaining BMD are lost with increased frequency of NG application.
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PMID:Frequency-dependent effect of nitric oxide donor nitroglycerin on bone. 1084 Nov 80

Nitric oxide is a gas radical regulating cell behaviour in the cardiovascular, immune, and central nervous systems. It has now been established as an important signalling molecule in bone. However, the effects of this gas radical on osteoblastic function are still unclear; in fact, while NO seems to be involved in anabolic processes mediated by mechanical strain, sex hormones and fracture healing, it also mediates catabolic processes in response to inflammation. We show here that a slow and moderate release of nitric oxide stimulates the replication of primary rat osteoblasts and alkaline phosphatase activity, while a rapid release and high concentrations of NO inhibit proliferation and induce apoptosis. We demonstrate that both the stimulatory and apoptosis-inducing effects of NO on primary osteoblasts are mediated by the second messenger cGMP, since both are abolished by the guanylate cyclase inhibitor ODQ.
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PMID:The biphasic effects of nitric oxide in primary rat osteoblasts are cGMP dependent. 1091 63

Nitric oxide (NO) is synthesised by a group of enzymes called nitric oxide synthases (NOS) and oxidizes to its stable end-products nitrite (NO2-) and nitrate (NO3-) We have previously reported in an in vivo rat model that NO is an important regulator for rat bone fracture healing. This study examines the effects of NO on alkaline phosphatase (ALP) activity in a rat fracture callus explant culture system. Explants of rat femoral fracture callus from days 4, 7, 14 and 28 post fracture induced NO2 release and ALP activity in a biphasic temporal manner, with the highest activity on day 7 and the lowest activity on day 14. Inhibition of NOS by co-incubation with an NOS inhibitor, S-(2-aminoethyl) isothiouronium bromide hydrobromide (AETU), inhibited ALP activity by an average of 50% at each time point (P <0.01). Supplementation with NO donor 3-morpholinosydnonomine hydrochloride (SIN-1) at low doses (25 and 0.025 microM) increased ALP activity by 20% (P < 0.01). ALP mRNA and histochemical ALP activity were localised to osteoblast-like and chondrocyte-like cells within fracture callus. The current study provides evidence that NO plays a regulatory role in ALP activity during rat fracture healing.
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PMID:Nitric oxide regulates alkaline phosphatase activity in rat fracture callus explant cultures. 1093 91


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