Mechanism of apoptosis induced by Mcl-1 inhibitor UMI-77 on gastric cancer MGC-803 cells / 安徽医科大学学报

ABSTRACT

Objective To investigate the mechanism of apoptosis induced by UMI-77 , a novel selective inhibitor of Mcl-1, in gastric cancer MGC-803 cells. Methods MGC-803 cells were treated with UMI-77 in different concen-trations for 24 h, apoptotic rates were determined by Annexin V/PI method using flow cytometry. Mitochondrial membrane potential was determined by JC-1 staining on a flow cytometer. The activation of Caspase-9, Caspase-3 and cleavage of PARP were measured by Western blot analysis. The protein level of Bcl-2, Bcl-XL and Mcl-1 was monitored by Western blot as well. Chemically synthesized Mcl-1 siRNA was transfected into MGC-803 cells using Lipofectamine 2000 Reagent. The efficacy of gene silencing was confirmed by Western blot analysis, and opoptotic rates before and after transfection was measured by flow cytometry using Annexin V/PI staining. Results UMI-77 was effective in induction of apoptosis in gastric cancer MGC-803 cells, apoptotic rates were increased in a dose-de-pendent manner. Mitochondrial membrane potential was collapsed after UMI-77 treatment. Activation of Caspase-9, Caspase-3 and cleavage of PARP occurred at 24 h (P<0. 05). The expression level of Bcl-2 and Bcl-XL were not altered after exposure to UMI-77 , while Mcl-1 was down-regulated after 12 h ( P<0. 05 ) . Transfection with Mcl-1 siRNA successfully decreased the expression level of Mcl-1 in MGC-803 cells ( P<0. 05 ) and blocked apoptosis induced by UMI-77 ( P<0. 05 ) . Conclusion UMI-77 induces apoptosis through activation of the intrinsic path-way in gastric cancer MGC-803 cells, and knocking down Mcl-1 expression abrogates apoptosis by UMI-77.