Caspase Pathway Activation and Reactive Oxygen Species Generation in Apoptotic Cell Death of Human Leukemic U937 and K562 Cell Line in Response to King Cobra (Ophiophagus hannah) Venom

Abstract

Resistance and decreasing efficacy of current synthetic drug for chemotherapy of leukemic cancer draws attention for development of newer anticancer agent from natural resources. In the present study, king cobra venom (OHV) significantly inhibited leukemic cell growth in dose and time dependent manner. For U937 and K562 cell line, the IC 50 dose (72 h) was found to be 4.1 µg/ml and 3.9 µg/ml respectively, observed by trypan blue exclusion method and tetrazolium bromide reduction assay. OHV treated morphometry of leukemic cell showed the characteristic features of apoptosis. Both U937 and K562 cells were arrested in the G1 phase of cell cycle with most cells exhibiting the biochemical feature of early and late apoptosis. Mitochondrial membrane potential was lost and reactive oxygen species generated highly in OHV treated leukemic cell line (U937 and K562). Western blot analysis showed OHV increased expression of Bax and decreased expression of Bcl2 in OHV treated cell as compared to untreated control U937 and K562 cell. Upregulation of Cytochrome c, Bid, Bad, Caspase 3/8/9, p21 and NF-κB down regulation of Cyclin D1, CDK4 was also showed by western blot analysis which revealed the possible pathway of OHV in cellular level. The results of this study demonstrated that OHV significantly and selectively induced leukemic cell death through both extrinsic and intrinsic apoptotic pathway.