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PAR1-dependent COX-2/PGE2 production contributes to cell proliferation via EP2 receptors in primary human cardiomyocytes.

ABSTRACT: BACKGROUND AND PURPOSE: Different protease-activated receptors (PARs) activated by thrombin are involved in cardiovascular disease, via up-regulation of inflammatory proteins including COX-2. However, the mechanisms underlying thrombin-regulated COX-2 expression in human cardiomyocytes remain unclear. EXPERIMENTAL APPROACH: Human cardiomyocytes were used in the study. Thrombin-induced COX-2 protein and mRNA expression, and signalling pathways were determined by Western blot, real-time PCR and COX-2 promoter luciferase reporter assays, and pharmacological inhibitors or siRNAs. PGE2 generation and cell proliferation were also determined. KEY RESULTS: Thrombin-induced COX-2 protein and mRNA expression, promoter activity and PGE2 release was attenuated by the PAR1 antagonist (SCH79797) or the inhibitors of proteinase activity (PPACK), MEK1/2 (U0126), p38 MAPK (SB202190) or JNK1/2 (SP600125), and transfection with small interfering RNA (siRNA) of PAR1, p38, p42 or JNK2. These results suggested that PAR1-dependent MAPKs participate in thrombin-induced COX-2 expression in human cardiomyocytes. Moreover, thrombin stimulated phosphorylation of MAPKs, which was attenuated by PPACK and SCH79797. Furthermore, thrombin-induced COX-2 expression was blocked by the inhibitors of AP-1 (tanshinone IIA) and NF-?B (helenalin). Moreover, thrombin-stimulated phosphorylation of c-Jun/AP-1 and p65/NF-?B was attenuated by tanshinone IIA and helenalin, respectively, suggesting that thrombin induces COX-2 expression via PAR1/MAPKs/AP-1 or the NF-?B pathway. Functionally, thrombin increased human cardiomyocyte proliferation through the COX-2/PGE2 system linking to EP2 receptors, as determined by proliferating cell nuclear antigen and cyclin D1 expression. CONCLUSIONS AND IMPLICATIONS: These findings demonstrate that MAPKs-mediated activation of AP-1/NF-?B pathways is, at least in part, required for COX-2/PGE2 /EP2 -triggered cell proliferation in human cardiomyocytes.

SUBMITTER: Chien PT

PROVIDER: S-EPMC4209155 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature

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PAR1-dependent COX-2/PGE2 production contributes to cell proliferation via EP2 receptors in primary human cardiomyocytes.

Chien Peter Tzu-Yu PT Hsieh Hsi-Lung HL Chi Pei-Ling PL Yang Chuen-Mao CM

British journal of pharmacology 20140905 19

<h4>Background and purpose</h4>Different protease-activated receptors (PARs) activated by thrombin are involved in cardiovascular disease, via up-regulation of inflammatory proteins including COX-2. However, the mechanisms underlying thrombin-regulated COX-2 expression in human cardiomyocytes remain unclear.<h4>Experimental approach</h4>Human cardiomyocytes were used in the study. Thrombin-induced COX-2 protein and mRNA expression, and signalling pathways were determined by Western blot, real-ti ...[more]

PMID: 24902855

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Project description:Enteric caliciviruses in the genera Norovirus and Sapovirus are important pathogens that cause severe acute gastroenteritis in both humans and animals. Cyclooxygenases (COXs) and their final product, prostaglandin E2 (PGE2), are known to play important roles in the modulation of both the host response to infection and the replicative cycles of several viruses. However, the precise mechanism(s) by which the COX/PGE2 pathway regulates sapovirus replication remains largely unknown. In this study, infection with porcine sapovirus (PSaV) strain Cowden, the only cultivable virus within the genus Sapovirus, markedly increased COX-2 mRNA and protein levels at 24 and 36 h postinfection (hpi), with only a transient increase in COX-1 levels seen at 24 hpi. The treatment of cells with pharmacological inhibitors, such as nonsteroidal anti-inflammatory drugs or small interfering RNAs (siRNAs) against COX-1 and COX-2, significantly reduced PGE2 production, as well as PSaV replication. Expression of the viral proteins VPg and ProPol was associated with activation of the COX/PGE2 pathway. We observed that pharmacological inhibition of COX-2 dramatically increased NO production, causing a reduction in PSaV replication that could be restored by inhibition of nitric oxide synthase via the inhibitor N-nitro-l-methyl-arginine ester. This study identified a pivotal role for the COX/PGE2 pathway in the regulation of NO production during the sapovirus life cycle, providing new insights into the life cycle of this poorly characterized family of viruses. Our findings also reveal potential new targets for treatment of sapovirus infection. IMPORTANCE:Sapoviruses are among the major etiological agents of acute gastroenteritis in both humans and animals, but little is known about sapovirus host factor requirements. Here, using only cultivable porcine sapovirus (PSaV) strain Cowden, we demonstrate that PSaV induced the vitalization of the cyclooxygenase (COX) and prostaglandin E2 (PGE2) pathway. Targeting of COX-1/2 using nonsteroidal anti-inflammatory drugs (NSAIDs) such as the COX-1/2 inhibitor indomethacin and the COX-2-specific inhibitors NS-398 and celecoxib or siRNAs targeting COXs, inhibited PSaV replication. Expression of the viral proteins VPg and ProPol was associated with activation of the COX/PGE2 pathway. We further demonstrate that the production of PGE2 provides a protective effect against the antiviral effector mechanism of nitric oxide. Our findings uncover a new mechanism by which PSaV manipulates the host cell to provide an environment suitable for efficient viral growth, which in turn can be a new target for treatment of sapovirus infection.

Dataset Information

PAR1-dependent COX-2/PGE2 production contributes to cell proliferation via EP2 receptors in primary human cardiomyocytes.

Publications

PAR1-dependent COX-2/PGE2 production contributes to cell proliferation via EP2 receptors in primary human cardiomyocytes.

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