Relationships of apoptotic signaling mediated by ceramide and TNF-α in U937 cells

Abstract

It is commonly assumed that ceramide is a second messenger that transduces signaling leading to apoptosis. We tested this hypothesis by investigating the role of ceramide in TNF-α-initiated apoptotic signaling using the histiocytic lymphoma cell line U937. We found considerable differences between cell killing by TNF-α and by ceramide. U937 cells treated with INF-α are committed early and irreversibly to the apoptotic pathway and start to die 90 min after treatment. U937 cells treated with ceramide start to die 12 h after the initial treatment. The cell death signaling initiated by TNF-α is transduced within minutes of exposure to INF-α and it is irreversible. Exogenous ceramide increases the intracellular level of ceramide rapidly, significantly, and well above the physiological levels, within minutes, but cellular commitment to death does not occur until after the first 6 h of incubation. Furthermore, the endogenous ceramide in U937 cells treated with TNF-α increases well after the commitment to the apoptotic pathway. The differences between ceramide and TNF-α in the kinetics and the commitment to the apoptotic pathway suggest that, (a) ceramide is not a second messenger in the apoptotic signaling of TNF-α, (b) ceramide elevations, in TNF-α treated cells, are a consequence rather than a cause of apoptosis and (c) exogenously added ceramide and TNF-α kill cells via different pathways.