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)

Previous studies have shown that transforming growth factor-beta1 (TGF-beta1) stimulates protein kinase C (PKC) via a mechanism that is independent of phospholipase C or tyrosine kinase, but involves a pertussis toxin-sensitive G-protein. Maximal activation occurs at 12 h and requires new gene expression. To understand the signaling pathways involved, resting zone chondrocytes were incubated with TGF-beta1 and PKC activity was inhibited with chelerythrine, staurosporine or H-7. [(35)S]Sulfate incorporation was inhibited, indicating that PKC mediates the effects of TGF-beta1 on matrix production. However, there was little, if any, effect on TGF-beta1-dependent increases in [(3)H]thymidine incorporation, and TGF-beta1-stimulated alkaline phosphatase was unaffected, indicating that these responses to the growth factor are not regulated via PKC. TGF-beta1 caused a dose-dependent increase in prostaglandin E(2) (PGE(2)) production which was further increased by PKC inhibition. The increase was regulated by TGF-beta1-dependent effects on phospholipase A(2) (PLA(2)). Activation of PLA(2) inhibited TGF-beta1 effects on PKC, and inhibition of PLA(2) activated TGF-beta1-dependent PKC. Exogenous arachidonic acid also inhibited TGF-beta1-dependent increases in PKC. The effects of TGF-beta1 on PKC involve genomic mechanisms, but not regulation of existing membrane-associated enzyme, since no direct effect of the growth factor on plasma membrane or matrix vesicle PKC was observed. These results support the hypothesis that TGF-beta1 modulates its effects on matrix production through PKC, but its effects on alkaline phosphatase are mediated by production of PGE(2) and protein kinase A (PKA). Inhibition of PKA also decreases TGF-beta1-dependent proliferation. We have previously shown that PGE(2) stimulates alkaline phosphatase through its EP2 receptor, whereas EP1 signaling causes a decrease in PKC. Thus, there is cross-talk between the two pathways.
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PMID:Transforming growth factor-beta1 regulation of resting zone chondrocytes is mediated by two separate but interacting pathways. 1077 Oct 99

The effects of dietary polyunsaturated fatty acids (PUFA) on ex vivo bone prostaglandin E(2) (PGE(2)) production and bone formation rate were evaluated in rats. Weanling male Sprague-Dawley rats were fed AIN-93G diet containing 70 g/kg of added fat for 42 d. The dietary lipid treatments were formulated with safflower oil and menhaden oil to provide the following ratios of (n-6)/(n-3) fatty acids: 23.8 (SMI), 9.8 (SMII), 2.6 (SMIII), and 1.2 (SMIV). Ex vivo PGE(2) production in liver homogenates and bone organ cultures (right femur and tibia) were significantly lower in rats fed diets with a lower dietary ratio of (n-6)/(n-3) fatty acids than in those fed diets with a higher dietary ratio. Regression analysis revealed a significant positive correlation between bone PGE(2) and the ratio of arachidonic acid (AA)/eicosapentaenoic acid (EPA), but significant negative correlations between bone formation rate and either the ratio of AA/EPA or PGE(2) in bone. Activities of serum alkaline phosphatase isoenzymes, including the bone-specific isoenzyme (BALP), were greater in rats fed a diet high in (n-3) or a low ratio of (n-6)/(n-3), further supporting the positive action of (n-3) fatty acids on bone formation. These results demonstrated that the dietary ratio of (n-6)/(n-3) modulates bone PGE(2) production and the activity of serum BALP in growing rats.
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PMID:Dietary ratio of (n-6)/(n-3) polyunsaturated fatty acids alters the fatty acid composition of bone compartments and biomarkers of bone formation in rats. 1095 24

Prostaglandin E(2) (PGE(2)) has been shown to exert a bone anabolic effect in young and adult rats. In this study we tested whether it possesses a similar effect on bone formation and bone mass in aging rats. Fifteen-month-old rats were injected daily with either PGE(2) at 5 mg/kg or vehicle for 14 days. PGE(2) treatment stimulated the rate of cancellous bone formation (a approximately 5.5-fold increase in bone formation rate), measured by the incorporation of calcein into bone-forming surfaces at the tibial proximal metaphysis. This effect resulted in increased cancellous bone area (+54%) at the same site. Since PGE(2) treatment resulted in a much higher proportion of bone surface undergoing bone formation and thus lined with osteoblasts, we tested the hypothesis that PGE(2) stimulates osteoblast differentiation from bone marrow precursor cells both in vivo and in vitro. We found that ex vivo cultures of bone marrow stromal cells from rats injected for 2 weeks with PGE(2) at 5 mg/kg per day yielded more ( approximately 4-fold) mineralized nodules and exhibited a greater (by 30-40%) alkaline phosphatase activity compared with cultures from vehicle-injected rats, attesting to a stimulation of osteoblastic differentiation by PGE(2). We also compared the osteogenic capacity of bone marrow from aging (15-month-old) versus young (5-week-old) rats and its regulation by PGE(2) in vitro. Bone marrow stromal cell cultures from aging rats exhibited a greatly diminished osteogenic capacity, reflected in reduced nodule formation ( approximately 6% of young animals) and lower alkaline phosphatase activity ( approximately 60% of young animals). However, these parameters could be stimulated in both groups of animals by incubation with 10-100 nM PGE(2). The magnitude of this stimulation was greater in cultures from aging rats (+550% vs +70% in nodule formation of aging compared with young rats). In conclusion, we demonstrate here that PGE(2) exerts a bone anabolic effect in aging rats, similar to the effect we and others have reported in young, growing rats. The PGE(2)-stimulated bone formation, which augments bone mass, most likely results from recruitment of osteoblasts from their bone marrow stromal precursors.
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PMID:Systemic prostaglandin E2 increases cancellous bone formation and mass in aging rats and stimulates their bone marrow osteogenic capacity in vivo and in vitro. 1113 77

Previous studies suggest that the enhanced expression of the osteoblastic phenotype exhibited by MG63 osteoblast-like cells on rough Ti surfaces (R(a) 4-5 microm) involves increased production of prostaglandin. Inhibition of prostaglandin synthesis by indomethacin blocks surface-roughness-dependent decreases in cell proliferation and increases in alkaline phosphatase activity and the production of osteocalcin and TGF-beta1. This study examined the hypothesis that the increase in expression of the osteoblastic phenotype noted in MG63 cells cultured on rough Ti surfaces is mediated by inducible cyclooxygenase-2 (Cox-2) whereas Cox-1 modulates prostaglandin production and phenotypic expression of the cells under standard conditions and on smooth Ti surfaces. MG63 cells were cultured on tissue culture plastic, smooth Ti (PT, R(a) = 0.60 microm), and two rough Ti surfaces with differing morphologies (SLA, R(a) = 3.97 microm and TPS, R(a) = 5.21 microm). At 24 h after plating, media were replaced with media containing the general Cox inhibitor indomethacin (10(-7)M), the Cox-1 inhibitor resveratrol (1 or 10 microM), or the Cox-2 inhibitor NS-398 (1 or 10 microM). Media were changed again after 48 h. Five days after plating, osteocalcin, PGE(2), and TGF-beta1 content of the conditioned media were determined. Cell numbers were assessed in the same cultures used for determination of osteocalcin production. Cell layer protein and alkaline phosphatase specific activity were assessed in cultures used to measure PGE(2) and TGF-beta1. Indomethacin, resveratrol, and NS-398 had no effect on cell number. Indomethacin blocked the surface-roughness-dependent increase in PGE(2) production by up to 80%. Similarly, resveratrol inhibited up to 50% of the PGE(2) production on smooth surfaces and up to 80% on rough surfaces. In contrast, NS-398 had no effect on PGE(2) production by cells on smooth surfaces but caused a 60% reduction in cultures on rough surfaces. Indomethacin reduced alkaline phosphatase on all surfaces below basal levels. However, neither resveratrol nor NS-398 had an effect. Indomethacin blocked the stimulatory effect of surface roughness on osteocalcin production while resveratrol only partially reduced osteocalcin production, and NS398 completely blocked the surface-dependent increase. TGF-beta1 production on rough surfaces was blocked by indomethacin. The effects of resveratrol and NS-398 were dose dependent, but neither agent caused total inhibition of the increase noted on SLA, and only resveratrol blocked the increase on TPS. These results indicate that both Cox-1 and Cox-2 are involved in the response of osteoblasts to surface roughness with respect to production of PGE(2), TGF-beta1, and osteocalcin. While prostaglandin mediates the effects of surface roughness on alkaline phosphatase, neither Cox-1 nor Cox-2 appears to be involved, at least with respect to the two inhibitors used.
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PMID:Both cyclooxygenase-1 and cyclooxygenase-2 mediate osteoblast response to titanium surface roughness. 1125 88

Periprosthetic osteolysis and implant loosening is associated with the presence of ultrahigh molecular weight polyethylene (UHMWPE) wear debris particles. Osteoblast phenotypic expression in vitro is affected by UHMWPE particles, suggesting that bone formation may also be affected by wear debris. Here we tested the hypothesis that the response of osteoblasts to UHMWPE can be modified by changes in UHMWPE particle chemistry. We used four different commercially available preparations of GUR UHMWPE particles to determine if chemical composition (+/- Ca-stearate) or polymer molecular weight (3.1-4.2 million or 5.4-6.5 million g/mol) modulates osteoblast response. Particles were characterized by size distribution, morphology, and number of particles added to the culture medium. They had an average equivalent circle diameter ranging from 0.46-1.26 microm. MG63 cell response was assessed by measuring cell number, cellular and cell layer alkaline phosphatase, and prostaglandin E2 (PGE2) production. There were dose-dependent effects of the particles on cell response. Cell number and PGE, production were increased, while alkaline phosphatase specific activity was decreased. In addition, there was a marked difference between cultures treated with particles containing Ca-stearate and as a function of polymer molecular weight. Particles of higher molecular weight caused a greater stimulation of proliferation and inhibition of alkaline phosphatase than particles of lower molecular weight. The presence of Castearate exerted a more pronounced depression of osteoblast phenotype as well as a significantly greater increase in PGE2 release by the cells. The present study shows that chemical composition and polymer molecular weight of UHMWPE are capable of modulating osteoblast response to particles. The results suggest that osteoblast differentiation is inhibited by UHMWPE particles, whereas cell proliferation and PGE2 production are stimulated. This may have direct effects on osteoblasts and bone formation, but also paracrine effects on cells of the monocytic lineage inducing bone resorption and promoting inflammation which may lead to aseptic loosening. The present results suggest that the cellular events in aseptic loosening may be modulated or even accelerated by changes in the composition of the UHMWPE used to fabricate implants.
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PMID:Effect of polymer molecular weight and addition of calcium stearate on response of MG63 osteoblast-like cells to UHMWPE particles. 1134 88

Growth plate chondrocyte function is modulated by the vitamin D metabolite 1alpha,25-(OH)(2)D(3) via activation of protein kinase C (PKC). In previous studies with cells derived from prehypertrophic and upper hypertrophic zones of rat costochondral cartilage (growth zone cells), inhibition of prostaglandin production with indomethacin caused a decrease in the stimulation of PKC activity, suggesting that changes in prostaglandin levels mediate the 1alpha,25-(OH)(2)D(3)-dependent response in these cells. Growth zone cells also respond to PGE(2) directly, indicating that prostaglandins act as autocrine or paracrine regulators of chondrocyte metabolism in the growth plate. The aim of the present study was to identify which PGE(2) receptor subtypes (EP) mediate the effects of PGE(2) on growth zone cells. Using primers specific for EP1-EP4, reverse transcription-polymerase chain reaction (RT-PCR) amplified EP1 and EP2 cDNA in a RT-dependent manner. In parallel experiments, we used EP subtype-specific agonists to examine the role of EP receptors in 1alpha,25-(OH)(2)D(3)-mediated cell proliferation and differentiation. 17-Phenyl-trinor-PGE(2) (PTPGE(2)), an EP1 agonist, decreased [3H]-thymidine incorporation in a dose-dependent manner and augmented the 1alpha,25-(OH)(2)D(2)-induced inhibition of [3H]-thymidine incorporation. PTPGE(2) also caused significant increases in proteoglycan production, as measured by [35S]-sulfate incorporation, and alkaline phosphatase specific activity. 1alpha,25-(OH)(2)D(3)-induced alkaline phosphatase activity was only slightly stimulated by PTPGE(2). In contrast, 1alpha,25-(OH)(2)D(3)-induced PKC activity was synergistically increased by PTPGE(2), whereas EP1 antagonists SC-19220 and AH6809 inhibited PKC activity in a dose-dependent manner. The EP2, EP3 and EP4 agonists had no effect on the various cell-induced responses measured. EP1 receptor-induced responses were blocked by the phospholipase C inhibitor U73122, and reduced by PKA inhibitors. EP1 receptor-induced PKC activity was insensitive to pertussis toxin or choleratoxin but blocked by the G-protein inhibitor GDPbetaS, suggesting the involvement of G(q). These results suggest that the EP1 receptor subtype mediates various PGE(2)-induced cellular responses in growth zone chondrocytes leading to decreased proliferation and enhanced differentiation, as well as the effect of 1alpha,25-(OH)(2)D(3) on cellular maturation.
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PMID:Characterization of PGE(2) receptors (EP) and their role as mediators of 1alpha,25-(OH)(2)D(3) effects on growth zone chondrocytes. 1159 7

The aim of the present study was to determine whether dietary intake of monounsaturated (MUFA) and/or polyunsaturated fatty acids (PUFA) of the (n- 3) and (n-6) series could improve intestinal damage and reduce inflammation in experimental ulcerative colitis (UC). Rats were treated with 80 mg/kg body of 2,4,6-trinitrobenzenesulfonic acid and fed for 1 or 2 wk diets enriched in olive oil (OO), fish oil (FO), or purified pig brain phospholipids (BPL), as sources of monounsaturated and PUFA of the (n-3) and (n-3) + (n-6) series. Evaluation of macroscopic and microscopic colonic damage was assessed. Ultrastructural and histologic changes were analyzed as well as plasma and colonic mucosa fatty acid profiles and some biochemical markers of injury and inflammation [alkaline phosphatase (AP), mieloperoxidase (MPO), prostaglandin E(2) (PGE(2)) and leukotriene B(4)]. Fatty acid profiles of both plasma and mucosa mostly reflected the dietary fatty acid composition. Plasma MUFA proportions were higher in UC animals fed the OO diet compared with FO or BPL groups 1 and 2 wk and (n-3) long chain PUFA (LC-PUFA) were higher in the FO than in the OO and BPL groups. At 1 wk, UC led to lower MUFA mucosa levels and (n-3)LC-PUFA were higher in the FO group compared with the OO and BPL groups. Rats with UC fed FO at 1 wk showed significantly less macroscopic and microscopic colonic damage. They also have lower AP and MPO activities and PGE(2) levels compared with the OO and BPL groups and showed enhanced histological repair, less necrotic areas within the mucosa, and more goblet cells with mature mucin granules. These results suggest that the use of balanced diets containing (n-3) LC-PUFA could ameliorate the inflammation and mucosal damage in UC.
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PMID:Dietary polyunsaturated fatty acids improve histological and biochemical alterations in rats with experimental ulcerative colitis. 1177 1

Mesenchymal cells isolated from chick limb-buds, when plated in micromass culture, differentiate into chondrocytes, forming a mineralizable matrix. Because these cells produce prostanoids during differentiation, this system was used to test the hypothesis that E-series prostanoids are involved in chondrocyte-mediated calcification. Prostaglandins E(2) (PGE(2)) and E(1) (PGE(1)), and the E-series analog, misoprostol (MP), increased chondrocyte cAMP content in the presence of the phosphodiesterase inhibitor IBMX. The increases for PGE(1) and PGE(2) at their saturation concentrations were twofold to threefold greater than for MP at its saturation concentration. At culture day 7, 9, or 11 (the day that mineralization commenced), the maximal cyclic adenosine monophosphate (cAMP) production was at a concentration of 250--500 ng/ml PGE(2), 500--1000 ng/ml PGE(1), and less-than-or-equal5000 ng/ml MP. At these concentrations, PGE(1) and PGE(2), but not MP, stimulated chondrocyte differentiation. (45)Ca accumulation in mineralizing, as compared to nonmineralizing, similarly treated control cultures was not altered by the addition of indomethacin and/or prostanoid when the phosphate source was inorganic phosphate. Because the prostanoids decrease alkaline phosphatase activity, initial beta-glycerophosphate-mediated mineralization was inhibited by each of the prostanoids.
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PMID:The Effect of Misoprostol and Prostanoids On cAMP Production and Calcification in a Differentiating Chick Limb-Bud Culture System. 1186 48

Chronic crystal-associated arthropathies such as gout and pseudogout can lead to local bone destruction. Because osteoblasts, which orchestrate bone remodeling via soluble factors and cell-to-cell interactions, have been described in contact with microcrystals, particularly in uratic foci of gout, we hypothesized that microcrystals of monosodium urate monohydrate (MSUM) and of calcium pyrophosphate dihydrate (CPPD) could alter osteoblastic functions. MSUM and CPPD adhered to human osteoblastic cells (hOB) in vitro and were partly phagocytized as shown by scanning electron microscopy. MSUM and CPPD dose-dependently stimulated the production of PGE(2) in hOB as assessed by enzyme immunoassay, a response that was synergistically enhanced in the presence of IL-1. The mechanism of this synergism was, at least in part, at the level of the expression of cyclooxygenase-2 as evaluated by immunoblot analysis. MSUM and CPPD also stimulated the expression of IL-6 and IL-8 and reduced the 1,25-dihydroxyvitamin D(3)-induced activity of alkaline phosphatase and osteocalcin in hOB (with no synergism with IL-1). MSUM- or CPPD-stimulated expression of IL-6 in hOB pretreated with the selective cyclooxygenase-2 inhibitor NS-398 was increased, unlike that induced by IL-1 alone which was partially reduced. MSUM-, CPPD- or IL-1-induced expression of IL-8 was unchanged by pretreating hOB with NS-398. These results suggest that inflammatory microcrystals alter the normal phenotype of hOB, redirecting them toward reduced bone formation and amplified osteoblast-mediated bone resorption, abnormalities that could play a role in the bone destruction associated with chronic crystal-induced arthritis.
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PMID:Inflammatory microcrystals alter the functional phenotype of human osteoblast-like cells in vitro: synergism with IL-1 to overexpress cyclooxygenase-2. 1199 89

Transforming growth factor beta 1 (TGF-beta1) affects growth plate chondrocytes through Smad-mediated mechanisms and has been shown to increase protein kinase C (PKC). This study determined if PKC mediates the physiological response of rat costochondral growth zone (GC) chondrocytes to TGF-beta1; if the physiological response occurs via type II or type III TGF-beta receptors, and, if so, which receptor mediates the increase in PKC; and the signal transduction pathways involved. Treatment of confluent GC cells with TGF-beta1 stimulated [(3)H]thymidine and [(35)S]sulfate incorporation as well as alkaline phosphatase (ALPase) and PKC specific activities. Inhibition of PKC with chelerythrine, staurosporine, or H-7 caused a dose-dependent decrease in these parameters, indicating that PKC signaling was involved. TGF-beta1-dependent PKC and the physiological response of GC cells to TGF-beta1 was reversed by anti-type II TGF-beta receptor antibody and soluble type II TGF-beta receptor, showing that TGF-beta1 mediates these effects through the type II receptor. The increase in [3H]thymidine incorporation and ALPase specific activity were also regulated by protein kinase A (PKA) signaling, since the effects of TGF-beta1 were partially blocked by the PKA inhibitor H-8. The mechanism of TGF-beta1 activation of PKC is through phospholipase A(2) (PLA(2)) and not through phospholipase C (PLC). Arachidonic acid increased PKC in control cultures and was additive with TGF-beta1. Prostanoids are required, as indomethacin blocked the effect of TGF-beta1, and Cox-1, but not Cox-2, is involved. TGF-beta1 stimulates prostaglandin E(2) (PGE(2)) production and exogenous PGE(2) stimulates PKC, but not as much as TGF-beta1, suggesting that PGE(2) is not sufficient for all of the prostaglandin effect. In contrast, TGF-beta1 was not regulated by diacylglycerol; neither dioctanoylglycerol (DOG) nor inhibition of diacylglycerol kinase with R59022 had an effect. G-proteins mediate TGF-beta1 signaling at different levels in the cascade. TGF-beta1-dependent increases in PGE(2) levels and PKC were augmented by the G protein activator GTP gamma S, whereas inhibition of G-protein activity via GDP beta S, pertussis toxin, or cholera toxin blocked stimulation of PKC by TGF-beta1, indicating that both G(i) and G(s) are involved. Inhibition of PKA with H-8 partially blocked TGF-beta1-dependent PKC, suggesting that PKA inhibition on the physiological response was via PKA regulation of PKC signaling. This indicates that multiple interacting signaling pathways are involved: TGF-beta1 stimulates PLA(2) and prostaglandin release via the action of Cox-1 on arachidonic acid. PGE(2) activates the EP2 receptor, leading to G-protein-dependent activation of PKA. PKA signaling results in increased PKC activity and PKC signaling regulates proliferation, differentiation, and matrix synthesis.
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PMID:Transforming growth factor-beta1 regulation of growth zone chondrocytes is mediated by multiple interacting pathways. 1206 64


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