CAJ | 학술논문

目的以人类白血病细胞株K562为研究对象,探讨BH3模拟物S1诱导人类白血病细胞凋亡的机制.方法通过XTT法检测S1作用下K562细胞的生存率；S1作用不同时间,流式细胞术检测细胞凋亡率；分光光度法检测Caspase-3、-8、-9激活；免疫共沉淀检测S1作用后bax、bak释放情况.结果与对照组相比S1能够以时间和浓度依赖的方式诱导K562细胞凋亡,24h的IC50值为13.5μmol/L；流式细胞术结果显示,S1处理12h后K562细胞出现大量早期凋亡,时间延长至24 h后,晚期凋亡比例显著增加；S1通过激活Caspase-3、-9而不是Caspase-8诱导K562细胞通过内源凋亡通路进行凋亡；免疫共沉淀实验检测5 μmol/L的S1处理8h后K562细胞中的抗凋亡蛋白bcl-2和mcl-1受到S1的拮抗,促凋亡蛋白bax和bak分别从抗凋亡蛋白bcl-2和mcl-1上得到释放.结论 S1可能通过拮抗bcl-2、mcl-1蛋白释放bax、bak诱导人类白血病细胞凋亡.
목적 이인류백혈병세포주K562위연구대상,탐토BH3모의물S1유도인류백혈병세포조망적궤제.방법 통과XTT법검측S1작용하K562세포적생존솔；S1작용불동시간,류식세포술검측세포조망솔；분광광도법검측Caspase-3、-8、-9격활；면역공침정검측S1작용후bax、bak석방정황.결과 여대조조상비S1능구이시간화농도의뢰적방식유도K562세포조망,24h적IC50치위13.5μmol/L；류식세포술결과현시,S1처리12h후K562세포출현대량조기조망,시간연장지24 h후,만기조망비례현저증가；S1통과격활Caspase-3、-9이불시Caspase-8유도K562세포통과내원조망통로진행조망；면역공침정실험검측5 μmol/L적S1처리8h후K562세포중적항조망단백bcl-2화mcl-1수도S1적길항,촉조망단백bax화bak분별종항조망단백bcl-2화mcl-1상득도석방.결론 S1가능통과길항bcl-2、mcl-1단백석방bax、bak유도인류백혈병세포조망.
Objective To investigate the apoptosis mechanism induced by BH3 mimetic S1 in human leukemia cell line K562.Methods Cell viability was detected by XTT to S1 in leukemia cell line K562.K562 cells was incubated with S1 for different time,the apoptosis rate of K562 cells was determined by flow cytometry analysis.Caspase-3,-8,and-9 activities were measured by absorption spectra.Co-immunoprecipitation was used to analyze the releasing of bax,bak from bcl-2 and mcl-1.Results Compared with control group,a dose-dependent increase in apoptosis coincided with a dose-dependent decrease in cell viability following S1 treatment suggested that S1 inhibits cell proliferation through the induction of apoptosis.The IC50 value at 24 h for S1 was 13.5 μ mol/L.Exposure of K562 cells to S1 for 12 h resulted in a time-dependent increase in FITC-Annexin-positive/PI-negative early apoptotic cells.The strong increase of FITC Annexin/PI doublepositive cells after a 24 h treatment indicated a shift to late apoptosis.S1 activated Caspase-3 and-9,but not Caspase-8 indicated that S1 induced K562 cells apoptosis via the intrinsic pathway.K562 cells treated with 5 μmol/L of S1 showed a disruption in bcl-2/bax,mcl-1/bak complexes after 8 h S1 treatment.Conclusion The main mechanism that S1 induces K562 cells apoptosis might be through the inhibition of bcl-2/bax,mcl-1/bak complexes dissociation.