DC prevents apoptosis by reduction in ceramide; blocking Flk-1 increases ceramide. (a) Apoptosis induced by serum deprivation was quantified by propidium iodide staining and flow cytometry. DC was added to test its dose-dependent effect on the prevention of apoptosis. (b) Immunoblot for activated caspase 3. Cells were deprived of serum, lysed, and the protein analyzed by SDS-PAGE/immunoblotting. Note that the addition of 100 μmol/l DC suppressed activation of caspase 3, which was consistent with prevention of apoptosis (see panel a). (c) HPTLC analysis of ceramide. Cells were cultivated in the absence of serum and treated with reagents as indicated in the figure. Cellular lipids were then isolated and analyzed by HPTLC. Note that serum deprivation leads to the elevation of ceramide by fourfold. DC suppresses this elevation by 70%. (d) HPTLC analysis of ceramide as in panel c. Note that the Flk-1 antagonist SU5416 (10 μmol/l) obliterated DC-induced reduction of ceramide. DC, sodium deoxycholate; HPTLC, high performance thin layer chromatography.