The activation of caspase-8,caspase-10 in Jurkat and U937 cells treated by anti-human DR5 monoclonal antibody-mDRA-6

Objective: To analyze the activation of caspase-8,caspase-10 in Jurkat and U937 cells treated by anti-human DR5 monoclonal antibody-mDRA-6. Methods: The activation of caspase-8,caspase-10,caspase-3 and PARP in Jurkat and U937 cells trea- ted by mDRA-6 were detected by Western blot,respectively. The caspase-8 and caspase-10 molecules of death-inducing signaling com- plex( DISC) in Jurkat and U937 cells treated by mDRA-6 were detected by immunoprecipitation. The mRNA expression of caspase-8 and caspase-10 in Jurkat and U937 cells were detected by real-time PCR. The inhibition and apoptosis of Jurkat and U937 cells after treating with mDRA-6 or / and caspases inhibitors were analyzed using MTT and FITC-Annexin V / PI staining. Results: When the cells were treated with mDRA-6,analysis by immunoblotting indicated that caspase-8,but not caspase-10 was activated in Jurkat cells; while caspase-10,but not caspase-8,was activated in U937 cells; caspases-3 and PARP were activated in both cells. Immunoprecipi- tation revealed that caspase-8 was a participator of DISC in Jurkat cells treated with mDRA-6,while caspase-10 was a participator of DISC in U937 cells treated with mDRA-6. caspase-8 mRNA expression was not difference in Jurkat and U937 cells,but caspase-10 mRNA expression in U937 cells was higher than that in Jurkat cells. When Jurkat and U937 cells were incubated with the inhibitors of caspase-8,caspase-10 and caspase-3 for one hour,Jurkat cells inhibition induced by mDRA-6 were reduced by 72. 94%( t = 20. 27, P 0. 01),8. 09%( t = 1. 82,P 0. 05) and 31. 20%( t = 8. 06,P 0. 01),U937 cells inhibition induced by mDRA-6 were reduced by 4. 53%( t = 0. 90,P 0. 05),69. 09%( t = 17. 25,P 0. 01) and 50. 20%( t = 13. 06,P 0. 01),respectively. While the Jurkat and U937 cells were incubated with the inhibitors of caspase-8 and caspase-10 for one hour,Jurkat cells apoptosis induced by mDRA- 6 were reduced by 80. 82% and 8. 34%,U937 cells apoptosis induced by mDRA-6 were reduced by 2. 50% and 48. 47%,respective- ly. Conclusion: mDRA-6 can induce apoptosis of both cell lines through activating the caspase signal pathway,but apoptotic signal pathways underlying are cell type dependent. caspase-8 is an initiator caspase for Jurkat cell,caspase-10 is an initiator caspase for U937 cell. The activation of caspase-10 in U937 cells may be correlate with its higher mRNA expression.