Role of AKT kinase in sphingosine-induced apoptosis in human hepatoma cells

Our previous work has shown that a number of sphingolipid metabolites inclu ding sphingosine, sphinganine, and other long-chain bases potently induced apoptosis in human hepatoma cells. in this study, we examined the possibili ty that sphingosine may trigger apoptosis in human hepatoma cells via inhib ition of anti-apoptotic pathways. We investigated the effect of sphingosine on AKT kinase, a serine/threonine kinase which was found to protect cells from apoptosis induced by a variety of extracellular stresses. Our results indicated that sphingosine inhibited basal and serum-stimulated AKT kinase activity in a dose-dependent manner in hepatoma cells. Additionally, sphing osine-induced inhibition of AKT kinase was correlated with induction of apo ptosis in these cells. Pretreatment of insulin, a potent stimulator of AKT kinase, partially reversed the inhibition of AKT kinase by sphingosine and counteracted the apoptotic action of this sphingolipid. Expression of activ ated AKT kinase partially protected cells from sphingosine-induced apoptosi s, whereas expression of kinase-dead AKT kinase had no effect. The molecula r mechanism by which AKT kinase suppressed the apoptotic action of sphingos ine was investigated. Our results showed that increased release of cytochro me C from mitochondria and subsequent activation of caspase-3 were detected in sphingosine-treated hepatoma cells. On the contrary, expression of acti vated AKT kinase in Hep3B cells attenuated cytochrome C release and caspase -3 activation induced by sphingosine. Taken together, these findings sugges t that suppression of AKT kinase is one of the mechanisms by which sphingos ine induces apoptosis in hepatoma cells and activation of AKT kinase may in hibit sphingosine-induced apoptosis by blocking a step upstream of cytochro me C release and caspase-3 activation. (C) 2001 Wiley-Liss, Inc.