L-carnitine prevents doxorubicin-induced apoptosis of cardiac myocytes: role of inhibition of ceramide generation

Besides the well-documented effect of the chemotherapeutic drug doxorubicin on free radical generation, the exact signaling mechanisms by which it cau ses cardiac damage remain largely unknown and are of fundamental importance in understanding anthracycline cardiotoxicity In this study, we describe t hat a 1 h treatment of isolated adult rat cardiac myocytes with doxorubicin (0.5 mu M) induced DNA fragmentation associated with the classical morphol ogical features of apoptosis observed after 7 days of culture, The doxorubi cin toxicity was preceded by an increase in intracellular ceramide levels w ith a concurrent decrease in sphingomyelin, Anthracycline-induced ceramide accumulation resulted from the activation of a sphingomyelinase assayed und er acidic conditions, an effect related to an increase in V-max. Pretreatme nt of cardiac myocytes with L-carnitine (200 mu g/ml), a compound known for its protective effect on cardiac metabolic injuries, was found to dose-dep endently inhibit the doxorubicin-induced sphingomyelin hydrolysis and ceram ide generation as well as subsequent cell death. However, L-carnitine did n ot protect cardiac myocytes from apoptosis induced by exogenous cell-permea nt ceramide, L-carnitine pretreatment did not affect the sphingomyelinase b asal activity but abolished the doxorubicin-induced increase in V-max. More over, in vitro studies conducted on cell extracts or with purified acid sph ingomyelinase demonstrated that L-carnitine exerted a dose-dependent, sphin gomyelinase inhibitory effect (through V-max reduction). Taken together, th ese findings show that by inhibiting a (perhaps novel) drug-activated acid sphingomyelinase and ceramide generation, L-carnitine can prevent doxorubic in-induced apoptosis of cardiac myocytes.