Distinct effects of ceramide-generating pathways in prostate adenocarcinoma cells

Ceramide, generated by the hydrolysis of sphingomyelin, mediates the actions of several cytokines such as tumour necrosis factor-alpha (TNF-alpha) interferon-gamma and interleukin-1beta (IL-1beta), including their inhibitory effect on tumour proliferation. We have evaluated the role of ceramide in the proliferation of prostate cancer by using the human prostate adenocarcinoma LNCaP cell line. Treatment of LNCaP cells with neutral or acidic sphingomyelinase or addition of C8- or C2-ceramide, two cell permeable analogues of endogenous ceramide, induced a profound inhibition of cell proliferation. This effect appeared after 24 h, was still present after 72 h of exposure to the drugs and exhibited concentration-dependency (10-200 and 5-200 mU ml(-1) for neutral and acidic sphingomyelinase, respectively, and 1-25 microM for C8-ceramide). The inhibitory effect on cell growth caused by neutral sphingomyelinase and ceramides was rapidly reversible as LNCaP cells rapidly regained their previous proliferation rate following withdrawal of the treatment. IL-1beta produced profound inhibition of LNCaP cell proliferation and caused enhanced ceramide formation. No clear features of apoptotic cell death were detectable by either oligonucleosome formation, cytofluorimetric analysis or nuclear staining following exposure of LNCaP cells to neutral sphingomyelinase, ceramide or IL-1beta. However, clear changes in LNCaP cell cycle distribution were detectable following these treatments. In contrast, treatment with acidic sphingomyelinase or TNF-alpha induced apoptotic death detectable by flow cytometric analysis and bisbenzimide staining. In conclusion, our data demonstrate that preferential activation of distinct enzymatic pathways by cytokines may lead to different outcomes in the viability of LNCaP cells.