Effects of ceramide on the induction of apoptosis and the involvement of p53-independent p21 during apoptosis were studied by using two hepatoma cell line, Hep3B (p53-null) and SK-Hep-1 cells (p53 wild type). Ceramide-induced apoptosis were detected after 24 houres and increased with longer treatment. Initial effects appeared at a 5 μM ceramide treated group and maximal effects occurred at a concentration of 20 μM. The level of p21 mRNA was low in untreated cells and increased by 24 and 48 hours after exposure to 20 μM C_6-ceramide in both cell lines. Furthermore, treatment of SK-Hep-1 cells with ceramide resulted in pRB dephosphorylation in a time- and dose- dependent manner. These results suggest that p53- independent p21 induction pathway might be involved in ceramide induced apoptosis by leading to G1 arrest in human hepatoma cells.