Sphingosine kinase-1 inhibition protects primary rat hepatocytes against bile salt-induced apoptosis

Sphingosine kinases (SphKs) and their product sphingosine-1-phosphate (SIP) have been reported to regulate apoptosis and survival of liver cells. Cholestatic liver diseases are characterized by cytotoxic levels of bile salts inducing liver injury. It is unknown whether SphKs and/or SIP play a role in this pathogenic process. Here, we investigated the putative involvement of SphK1 and SIP in bile salt-induced cell death in hepatocytes. Primary rat hepatocytes were exposed to glycochenodeoxycholic acid (GCDCA) to induce apoptosis. GCDCA-exposed hepatocytes were co-treated with SIP, the SphK1 inhibitor Ski-II and/or specific antagonists of SIP receptors (S1PR(1) and S1PR(2)). Apoptosis and necrosis were quantified. Ski-II significantly reduced GCDCA-induced apoptosis in hepatocytes (-70%, P <0.05) without inducing necrosis. GCDCA increased the SW levels in hepatocytes (P <0.05). GCDCA induced [Ca2+] oscillations in hepatocytes and co-treatment with the [Ca2+] chelator BAPTA repressed GCDCA-induced apoptosis. Ski-II inhibited the GCDCA-induced intracellular [Ca2+] oscillations. Transcripts of all five SW receptors were detected in hepatocytes, of which S1PR(1) and S1PR(2) appear most dominant. Inhibition of S1PR(1), but not S1PR(2), reduced GCDCA-induced apoptosis by 20%. Exogenous SIP also significantly reduced GCDCA-induced apoptosis (-50%, P