COX-2 and cytosolic PLA2 mediate IL-1beta-induced cAMP production in human vascular smooth muscle cells (original) (raw)
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Cytokine, 1995
Vasodilatation is a major feature of infection and inflammation, occurring systemically during sepsis, and locally at sites of infection or inflammation. Recent evidence has implicated a role for the pro-inflammatory cytokine interleukin-1 (IL-1) as a mediator of vasodilatation in sepsis and inflammation. However the mechanism of IL-1-induced vasodilatation in humans is unknown. Much attention has focused on the potential role for vascular smooth muscle cell (VSMC)-derived nitric oxide (NO) in IL-1-induced vasodilatation, as supported by evidence from animal studies. 2,4,10 While IL induces NO production in cultured VSMC derived from rat or rabbit aorta, 4,5 it does not induce NO production in cultured VSMC derived from human blood vessels. 3 A role for prostanoids in the vasodilatory effects of IL-1 is also supported by animal studies. In rabbits in vivo, the hypotension induced by i.v. administration of IL-1 is dependent on prostanoids. 32 Likewise, IL-1 causes prostanoid-dependent relaxation of pial arterioles of piglets in situ, 37 and of isolated rabbit mesenteric arteries in vitro. 25 The latter effect is independent of the endothelium, suggesting that it is a direct action on VSMC. IL-1 also enhances prostanoid production in VSMC derived from human blood vessels including aorta, renal artery and saphenous vein, 1,21 suggesting a potential role of prostanoids in humans. The mechanisms by which prostanoids induce vasodilatation have not been well documented, but are thought to involve activation of adenylate cyclase. 17 While several studies indicate that IL-1 induces prostanoid production in VSMC, none have addressed the potential implications for IL-1 effects on adenylate cyclase activity. The present study shows that IL-1 is a potent activator of prostacyclin production in human saphenous VSMC, and furthermore, that enhanced prostacyclin production results in a prolonged and marked increase in intracellular cAMP. These results estabish IL-1 as a major regulator of cAMP levels in human VSMC, and implicate cAMP as a mediator of IL-1-induced vasodilatation in man.
Journal of Biological Chemistry, 1999
Type II-secreted phospholipase A 2 (type II-sPLA 2) is expressed in smooth muscle cells during atherosclerosis or in response to interleukin-1. The present study shows that the induction of type II-sPLA 2 gene by interleukin-1 requires activation of the NFB pathway and cytosolic PLA 2 /PPAR␥ pathway, which are both necessary to achieve the transcriptional process. Interleukin-1 induced type II-sPLA 2 gene dose-and time-dependently and increased the binding of NFB to a specific site of type II-sPLA 2 promoter. This effect was abolished by proteinase inhibitors that block the proteasome machinery and NFB nuclear translocation. Type II-sPLA 2 induction was also obtained by free arachidonic acid and was blocked by either AACOCF 3 , a specific cytosolic-PLA 2 inhibitor, PD98059, a mitogen-activated protein kinase kinase inhibitor which prevents cytosolic PLA 2 activation, or nordihydroguaiaretic acid, a lipoxygenase inhibitor, but not by the cyclooxygenase inhibitor indomethacin, suggesting a role for a lipoxygenase product. Type II-sPLA 2 induction was obtained after treatment of the cells by 15-deoxy-⌬ 12,14-dehydroprostaglandin J 2 , carbaprostacyclin, and 9-hydroxyoctadecadienoic acid, which are ligands of peroxisome proliferator-activated receptor (PPAR) ␥, whereas PPAR␣ ligands were ineffective. Interleukin-1 as well as PPAR␥-ligands stimulated the activity of a reporter gene containing PPAR␥-binding sites in its promoter. Binding of both NFB and PPAR␥ to their promoter is required to stimulate the transcriptional process since inhibitors of each class block interleukin-1-induced type II-sPLA 2 gene activation. We therefore suggest that NFB and PPAR␥ cooperate at the enhanceosome-coactivator level to turn on transcription of the proinflammatory type II-sPLA 2 gene.
Circulation Research, 2004
Increased levels of inflammatory cytokines contribute to the pathophysiology of pulmonary hypertension. Prostacyclin (PGI 2 ) analogues, which relax pulmonary vessels mainly through cAMP elevation, have a major therapeutic role. In this study, we show that prolonged incubation with bradykinin (BK), interleukin-1β (IL-1β), and transforming growth factor-β 1 (TGF-β 1 ) markedly impairs cAMP accumulation in human pulmonary artery smooth muscle cells in response to short-term incubation with prostaglandin E 2 (PGE 2 ) and the PGI 2 analogues iloprost and carbaprostacyclin. A similar reduction in cAMP accumulation in response to a direct adenylyl cyclase activator, forskolin, suggested that the effect was attributable to downregulation of adenylyl cyclase. Reverse transcriptase–polymerase chain reaction studies showed downregulation of adenylyl cyclase isoforms 1, 2, and 4. The effect of IL-1β, BK, and TGF-β 1 on cAMP levels was abrogated by the selective COX-2 inhibitor NS398. Furthermo...
American Journal of Respiratory and Critical Care Medicine, 2000
We studied the effect of endogenous and exogenous prostaglandin E 2 (PGE 2), a metabolite of arachidonic acid through the cyclooxygenase (COX) pathway, on interleukin (IL)-1 -induced COX-2 expression, using primary cultures of human bronchial smooth-muscle cells (HBSMC). Treatment with exogenous PGE 2 resulted in enhanced expression of IL-1 -induced COX-2 protein and messenger RNA (mRNA) as compared with the effect of the cytokine per se. Inhibition of PGE 2 production with a nonselective COX inhibitor (flurbiprofen, 10 M) resulted in a significant reduction in IL-1 induced COX-2 expression, supporting a role of endogenous COX metabolites in the modulation of COX-2 expression. None of the experimental conditions used in the study affected the expression of constitutive cyclooxygenase (COX-1). Treatment with cycloheximide to inhibit translation, and with dexamethasone or actinomycin D to inhibit transcription, linked the effect of PGE 2 to the transcriptional level of COX-2 mRNA rather than to a potential effect on protein and/or mRNA stabilization. PGE 2 increased adenylate cyclase activity in a concentration dependent manner, and forskolin, a direct activator of adenylate cyclase, caused a marked increase in IL-1 -dependent COX-2, suggesting the existence of a causal relationship between the two events. The same results were observed with salbutamol, a bronchodilator that acts by increasing cyclic adenosine monophosphate. The effect of PGE 2 on COX-2 expression may contribute to the hypothesized antiinflammatory role of PGE 2 in human airways, providing a self-amplifying loop leading to increased biosynthesis of PGE 2 during an inflammatory event.
2003
Objective: The present study was designed to investigate the role of protein kinase C (PKC) and phospholipase D (PLD) in angiotensin II (AngII)-and phorbol ester (PMA)-induced cyclooxygenase-2 (COX-2) expression and prostacyclin (PGI 2 ) production in rat aortic smooth muscle cells (VSMC). Methods: Prostacyclin production in cultured VSMC was determined by radioimmunoassay. PKC activity was examined by measuring the transfer of 32 P from (g-32 P)ATP to histone III-S. COX-2 expression was determined by Western blotting. To measure PLD activity, thin layer chromatography was used. Results: AngII (50 nM) and PMA (100 nM) promoted the translocation of PKC activity from the cytosol to the membranes within 30 min, followed by a strong increase in PLD activity as well as COX-2 expression and PGI 2 production. After 48 h exposure to PMA, PKC was downregulated resulting in a complete suppression of its activity. PKC-downregulation and the PKC inhibitor CGP41251 abolished PMA-and AngII-induced PLD activation, suppressed the stimulatory effect of PMA on COX-2 expression and PGI 2 production and strongly inhibited that of AngII. Furthermore, AngII-and PMA-induced PGI 2 production depended on protein synthesis and COX-2 but not COX-1 activity. Inhibition of PLD-mediated phosphatidic acid (PA) formation by 1% 1-butanol abolished AngII-induced COX-2 expression and PGI 2 secretion, while dioctanoyl PA increased COX-2 expression and PGI 2 production in a time-and concentration-dependent manner. Conclusion: Our results indicate that in VSMC, AngII promotes PGI 2 production to a large extent through a rise in COX-2 expression which is mediated by PA generated from increased PKC-dependent PLD activity.
Prostaglandins, Leukotrienes and Essential Fatty Acids, 1996
Elucidation of the cellular and molecular mechanisms which regulate vascular smooth muscle cell proliferation is critical to the understanding of atherogenesis. The present studies were conducted to evaluate the relationship between prostaglandin E1 (PGG) and cAMP in the regulation of DNA synthesis in mouse vascular smooth muscle cells (SMCs). Quiescent cultures of SMCs were challenged with 10% fetal bovine serum to initiate cell cycle transit and PGE1 (10 pM) or dibutyryl cAMP (1, 10, 100 pM) added at 0, 8, 16, 24, and 32 h. DNA synthesis as measured by [3H] thymidine incorporation and intracellular cAMP levels were measured 24 h following individual treatments. PGE~ modulated DNA synthesis in a cell cycle related fashion, with inhibition only observed in cells challenged 16 h or longer following initiation of cell cycle transit. The decrease in DNA synthesis induced by PGG was associated with increased intracellular cAMP levels at 16 and 24 h, but not 32 h. Exposure of SMCs to dibutyryl-cAMP also inhibited DNA synthesis in a cell cycle related fashion, with the most pronounced effect seen at 16 h. These results demonstrate that the effects of PGE1 are restricted to a defined period within the cell cycle following S phase entry and implicate modulation of intracellular cAMP levels in the inhibitory response.
Regulation of cyclooxygenase-2 expression by cyclic AMP
Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 2007
Prostaglandins (PG) regulate many biological processes, among others inflammatory reactions. Cyclooxygenases-1 and-2 (COX-1 and COX-2) catalyse PG synthesis. Since this step is rate limiting, the regulation of COX expression is of critical importance to PG biology. Contrary to COX-1, which is constitutively expressed, COX-2 expression is subject to regulation. For example, COX-2 levels are increased in inflammatory reactions. Many signalling pathways can regulate COX-2 expression, not least those involving receptors for COX products themselves. Analysis of the intracellular signal transducers involved reveals a crucial role for cAMP, albeit as a modulator rather than direct inducer. Indeed, the influence of cAMP on COX-2 expression is complex and dependent on the cell type and cellular environment. This review aims to summarise various topics related to cAMP-dependent COX-2 expression. Firstly, the main aspects of COX-2 regulation are briefly considered. Secondly, the molecular basis for COX-2 gene (post)-transcriptional regulation is reviewed. Lastly, a detailed overview of the effects of cAMP-dependent signalling on COX-2 mRNA and protein expression in various human and rodent cells is provided. There is a large number of marketed, clinical and preclinical concepts promoting the elevation of intracellular cAMP levels for therapeutic purposes (e.g., β 2-agoinsts, PG receptor agonists, phosphodiesterase inhibitors). In this respect, the role of cAMP in the regulation of COX-2 expression, especially the human enzyme, is of significant clinical importance.