The hepatotoxic bile acid, glycochenodeoxycholate (GCDC), modulates hepatocyte cell death through activation of JNK, Akt and Erk. The non-hepatotoxic bile acid, taurocholate, activates Akt and Erk through the sphingosine 1-phosphate receptor 2 (S1PR2). The role of the S1PR2 in GCDC mediated apoptosis and kinase activation is unknown. Studies were done in rat hepatocytes and HUH7 cells and HUH7 cells stably transfected with rat Ntcp, HUH7-Ntcp. Cells were treated with GCDC and apoptosis monitored morphologically by Hoechst staining and biochemically by immunoblotting for the active cleaved fragment of caspase 3. Kinase activation was determined by immunoblotting with phospho-specific antibodies. JTE-013, an inhibitor of S1PR2, significantly attenuated morphologic evidence of GCDC induced apoptosis and prevented caspase 3 cleavage in rat hepatocytes and HUH7-Ntcp cells. In hepatocytes, JTE-013 mildly suppressed, augmented and had no effect on GCDC induced JNK, Akt and Erk phosphorylation, respectively. Similar results were seen in HUH7-Ntcp cells except for mild suppression of JNK and Erk phosphorylation. Knock down of S1PR2 in HUH7-Ntcp augmented Akt, inhibited JNK and had no effect on Erk phosphorylation. GCDC failed to induce apoptosis or kinase activation in HUH7 cells. In conclusion, SIPR2 inhibition attenuates GCDC induced apoptosis and inhibits and augments GCDC induced JNK and Akt phosphorylation, respectively. In addition, GCDC must enter hepatocytes to mediate cell death or activate kinases. These results suggest that SIPR2 activation is pro-apoptotic in GCDC induced cell death, but that this effect is not due to direct ligation of the S1PR2 by the bile acid.
- hepatocytes apoptosis
- Copyright © 2015, American Journal of Physiology- Gastrointestinal and Liver Physiology