Phosphatidylinositol-3-kinase p110γ contributes to bile salt-induced apoptosis in primary rat hepatocytes and human hepatoma cells

Background & Aims: Glycochenodeoxycholate (GCDC) and taurolithocholate (TLC) are hepatotoxic and cholestatic bile salts, whereas tauroursodeoxycholate (TUDC) is cytoprotective and anticholestatic. Yet they all act, in part, through phosphatidylinositol-3-kinase(PI3K)-dependent mechanisms (PI3K-paradox). Hepatocytes express three catalytic PI3K Class I isoforms (p110 alpha/beta/gamma), specific functions of which, in liver, are unclear. In other cell types, p110 gamma is associated with detrimental effects. To shed light on the PI3K enigma, we determined whether hydrophobic and hydrophilic bile salts differentially activate distinct p110 isoforms in hepatocytes, and whether p110 gamma mediates bile salt-induced hepatocyte cell death. Methods: Isoform-specific PI3K activity assays were established to determine isoform activation by bile salts in rat hepatocytes. Activation of Akt and JNK was determined by immunoblotting. Following stimulation with hydrophobic bile salts, hepatocellular apoptosis was determined morphologically after Hoechst staining and by analysis of caspase-3/-7 activity or caspase-3 cleavage. Activity or expression of PI3K p110 gamma was inhibited pharmacologically (AS604850) or by knock-down using specific siRNA. Results: All bile salts tested activated p110 beta, while p110 alpha was activated by TUDC and GCDC. Intriguingly, only hydrophobic bile salts activated p110 gamma. Inhibition of p110 gamma attenuated GCDC-induced Akt- and JNK-activation, but did not alter TUDC- or cAMP-induced Akt-signaling in rat hepatocytes. Inhibition or knock-down of p110 gamma markedly attenuated hydrophobic bile salt-induced apoptosis in rat hepatocytes and human hepatoma cell lines but did not alter Fas-, tumor necrosis factor alpha- and etoposide-induced apoptosis. Depletion of Ca(++). prevented GCDC-induced toxicity in rat hepatocytes but did not affect GCDC-induced Akt- and JNK-activation. Conclusions: PI3K p110 gamma is activated by hydrophobic, but not hydrophilic bile salts. Bile salt-induced hepatocyte apoptosis is partly mediated via a PI3K p110 gamma dependent signaling pathway, potentially involving JNK. (C) 2010 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.