Involvement of cytosolic phospholipase A(2) alpha signalling pathway in spontaneous and transforming growth factor-beta-induced activation of rat hepatic stellate cells.

摘要

BACKGROUND: Hepatic stellate cells (HSCs) are extracellular matrix-producing cells that play a pivotal role in liver fibrogenesis. During liver injury and when cells are placed in vitro, HSCs undergo phenotypic transition from quiescent retinoid-storing cells to activated retinoid-deficient myofibroblast-like cells. Although several mediators including reactive oxygen species, platelet derived growth factor, transforming growth factor-beta (TGF-beta) and tumour necrosis factor-alpha (TNF-alpha) were implicated in HSC activation, the cellular signalling pathways that regulate this process remain incompletely defined. AIMS: The objectives of this study were to evaluate the role of cytosolic phospholipase A(2) alpha (cPLA(2) alpha) and peroxisome proliferator-activated receptor-beta/delta (PPAR-beta/delta) in HSC activation. METHODS: Rat HSCs were isolated, purified, cultured and stimulated with TGF-beta1 in the presence or absence of the selective cPLA(2) alpha inhibitor, arachidonyltrifluoromethyl ketone (AACOCF(3) ). The activation status of HSC was evaluated by immunofluorescent staining of alpha-smooth muscle actin (alpha-SMA) and by measuring the expression of cPLA(2) alpha, cyclooxygenase 2 (COX-2) and PPAR-beta/delta using western blot analysis. RESULTS: Rapid and significant increase in cPLA(2) alpha expression was observed during activation of HSCs. These events preceded the elevation of PPAR-beta/delta and the expression of alpha-SMA. Elevated expression of cPLA(2) alpha, but not COX-2, was also observed during TGF-beta-induced HSC activation. The TGF-beta-induced alpha-SMA expression was blocked by AACOCF(3) . Furthermore, transfection of a cPLA(2) alpha expression vector enhanced the transcription activity of PPAR-beta/delta and the expression of alpha-SMA in HSCs. CONCLUSION: cPLA(2) alpha-mediated induction of PPAR-beta/delta is a novel intracellular signalling pathway in spontaneous and TGF-beta induced activation of HSCs and could be a potential therapeutic target for the treatment of liver fibrosis.