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

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‐κB (helenalin). Moreover, thrombin‐stimulated phosphorylation of c‐Jun/AP‐1 and p65/NF‐κB was attenuated by tanshinone IIA and helenalin, respectively, suggesting that thrombin induces COX‐2 expression via PAR1/MAPKs/AP‐1 or the NF‐κB 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‐κB pathways is, at least in part, required for COX‐2/PGE₂/EP₂‐triggered cell proliferation in human cardiomyocytes.

DOI: 10.1111/bph.12794

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