Binding of tumour necrosis factor-alpha (TNF-alpha) to TNF-RI induces caspase(s)-dependent apoptosis in human cholangiocarcinoma cell lines

Cholangiocarcinoma (CCA), a tumour of the bile duct epithelium, occurs with a higher incidence in South-east Asian countries than in Europe and North America. The prognosis is poor, due to the unavailability of early diagnosi s and the tumours being relatively resistant to chemotherapy. In the presen t study one of the fatal routes of this tumour was studied. This death was stimulated by TNF-alpha. TNF-alpha at a concentration of 760 pg/ml and 100 pg/ml in the presence of 1 mu g/ml actinomycin D induced 50% cell death of the two established human cholangiocarcinoma cell lines HuCCA-1 and HuCCA-1 Nu, respectively. Preincubation of both cell lines with MoAb to TNF-RI or T NF-RII before TNF-alpha treatment showed that only the MoAb specific to TNF -RI inhibited death. The death of these two cell lines was proved to be apo ptosis. Western blot analysis of extracts from both cell lines demonstrated a cleavage of poly (ADP-ribose) polymerase (PARP) within 6-8 h following T NF-alpha treatment. The degradation of PARP was prevented by a MoAb to TNF- RI indicating that the TNF-RI but not TNF-RII was involved in TNF-induced a poptosis in these two human cholangiocarcinoma cell lines. Moreover, peptid e inhibitor for caspase II subfamily, Ac-DEVD-CHO, reduced the cytolysis of TNF-alpha-treated cholangiocarcinoma cells. The inhibitor also prevented d egradation of PARP. These results indicate that the interaction between TNF -alpha and TNF-RI alone generated a sufficient signal to activate a caspase II subfamily-dependent apoptosis in human cholangiocarcinoma cell lines.