CAJ | 학술논문

目的以细胞周期特异性T淋巴细胞型白血病Molt-4细胞S期凋亡为模式,检测细胞周期蛋白依赖性激酶1( CDK1)在S期细胞周期检测点蛋白磷酸化及蛋白去磷酸化,探讨细胞周期时相特异性细胞凋亡的CDK1的活性改变.方法对数生长期的人类急性淋巴细胞白血病细胞系Molt-4经喜树碱(CPT)诱导细胞凋亡,经不同的作用时间、剂量处理后,以API法检测细胞凋亡的细胞周期特异性,以分选后激光共聚焦荧光显微镜技术( PSC)对API结果进一步验证,从而获取最佳条件下细胞周期特异性细胞凋亡,蛋白电泳分析法检测CDK1的磷酸化、去磷酸化.结果对数生长期的Molt-4细胞经CTP诱导后发生细胞周期特异性细胞凋亡,CPT 0.2 μg/ml处理后6h出现S期特异性细胞凋亡.CTP诱导出现S期特异性细胞凋亡时CDK1-第15位酪氨酸(Tyr15)与对照Western blot结果条带相近,而CDK1-第161位苏氨酸( Thr161)条带较窄.结论细胞凋亡往往伴有细胞周期特异性；经PSC形态学证实的API流式细胞术是有效、快捷的细胞周期特异性细胞凋亡分析方法；CDK1在S期特异性细胞凋亡时磷酸化活性降低.
목적 이세포주기특이성T림파세포형백혈병Molt-4세포S기조망위모식,검측세포주기단백의뢰성격매1( CDK1)재S기세포주기검측점단백린산화급단백거린산화,탐토세포주기시상특이성세포조망적CDK1적활성개변.방법 대수생장기적인류급성림파세포백혈병세포계Molt-4경희수감(CPT)유도세포조망,경불동적작용시간、제량처리후,이API법검측세포조망적세포주기특이성,이분선후격광공취초형광현미경기술( PSC)대API결과진일보험증,종이획취최가조건하세포주기특이성세포조망,단백전영분석법검측CDK1적린산화、거린산화.결과 대수생장기적Molt-4세포경CTP유도후발생세포주기특이성세포조망,CPT 0.2 μg/ml처리후6h출현S기특이성세포조망.CTP유도출현S기특이성세포조망시CDK1-제15위락안산(Tyr15)여대조Western blot결과조대상근,이CDK1-제161위소안산( Thr161)조대교착.결론 세포조망왕왕반유세포주기특이성；경PSC형태학증실적API류식세포술시유효、쾌첩적세포주기특이성세포조망분석방법；CDK1재S기특이성세포조망시린산화활성강저.
Objective To investigate the phosphorylation and dephosphorylation of CDK1 based on the specific cell cycle apoptosis in Molt-4 cells and active variety of CDK1 in cell cycle specific apoptosis.Methods The exponential phase of growth Molt-4 cells (the human acute lymphoblastic leukemia cell line) were induced with dose response and time course of Camptothecin (CPT).The specific cell cycle apoptosis was detected with API method,then cell apoptosis was verified with post sorting confocal method.Meanwhile,the phosphorylation and dephosphorylation of CDK1 were detected by the protein electrophoretic analysis (Western blot).Results The specific cell cycle apoptosis occurred on exponential phase of growth Molt-4 cells after CPT treatment.When Molt-4 cells occured S-phase apoptosis, the apoptosis cell phosphorylation of CDK1-Thr161 band was more narrow than that of control cells, the apoptosis cell phosphorylation of CDK1-Thr15 band almost had the same band with control cells.Conclusion Cell apoptosis frequently developed in different cell cycle phase. API assay is quick and efficient method to analyze specific cell cycle apoptosis. When cell apoptosis take place in S-phase,the phosphorylation activity of CDK1 is inhibited.