Restoration of TNF-α-induced ceramide generation and apoptosis in resistant human leukemia KG1a cells by the p-glycoprotein blocker PSC833

Tumor necrosis factor (TNF-alpha) is a cytokine with antitumor activity against several cellular models, TNF-alpha-induced apoptosis seems to be mediated by a signaling pathway termed 'sphingomyelin-ceramide' pathway, which consists of the hydrolysis of sphingomyelin and the production of its breakdown product ceramide, Our study shows that KG1a cells, which are inherently resistant to TNF-alpha and do not produce ceramide upon cytokine stimulation, can be sensitized by the use of the P-glycoprotein inhibitor PSC833, Coincubation with 1 mu M of this cyclosporin derivative restored the apoptotic potential of 10 ng/ml TNF-alpha, This effect was associated with the restoration of ceramide generation (315%) and activation of neutral, but not acid sphingomyelinase activity (143%), Furthermore, we demonstrate that treatment of KG1a cells with 1 mu M PSC833 led to a threefold increase in inner plasma membrane sphingomyelin content and basal neutral sphingomyelinase activity, These results support the hypothesis whereby resistance to TNF-alpha-mediated apoptosis of certain leukemic cells is linked to the disposability of the sphingomyelin pool, These data also suggest a role for P-glycoprotein in sphingomyelin transverse plasma membrane asymmetry.