CAJ | 학술논문

目的观察重构型caspase-3及其N-端融合蛋白的表达对HeLa细胞的凋亡诱导作用. 方法用反转录PCR获取人caspase-3基因，用重组PCR方法构建人重构型caspase-3及其融合蛋白基因，将基因克隆入表达载体pcDNA3，脂质体法转染HeLa细胞，通过细胞计数、电镜观察、MTT法等检测被转染细胞的生长及其凋亡状况. 结果成功获得了caspase-3基因，构建了重构型caspase-3基因和重构型caspase-3与绿脓杆菌外毒素（PE）转膜结构域部分肽段的融合蛋白基因及其表达载体，与对照组相比，在转染后1～4 d内，两种基因的表达均导致HeLa细胞存活数减少，大量细胞发生凋亡，且二者活性相当. 结论重构型caspase-3及其融合蛋白可以诱导转染的HeLa细胞发生凋亡.
목적관찰중구형caspase-3급기N-단융합단백적표체대HeLa세포적조망유도작용. 방법용반전록PCR획취인caspase-3기인，용중조PCR방법구건인중구형caspase-3급기융합단백기인，장기인극륭입표체재체pcDNA3，지질체법전염HeLa세포，통과세포계수、전경관찰、MTT법등검측피전염세포적생장급기조망상황. 결과성공획득료caspase-3기인，구건료중구형caspase-3기인화중구형caspase-3여록농간균외독소（PE）전막결구역부분태단적융합단백기인급기표체재체，여대조조상비，재전염후1～4 d내，량충기인적표체균도치HeLa세포존활수감소，대량세포발생조망，차이자활성상당. 결론중구형caspase-3급기융합단백가이유도전염적HeLa세포발생조망.
AIM　To investigate the induction of HeLa cells to apoptosis by the expression of reversed caspase-3 and its N-terminal fusion protein. METHODS　RT-PCR and recombi-nant PCR were used to amplify human caspase-3 gene and construct reversed caspase-3 and its fusion protein genes. Following cloning into expression vector pcDNA3 and transfecting HeLa cells, we evaluated the growth condition and apoptosis of these cells by cell counting, electron microscope photographs, as well as MTT assays. RESULTS　We successfully obtained human caspase-3 gene, the genes and expression vectors of reversed caspase-3 and its fusion protein with part of the translocation domain of Pseudomonas exotoxin A (PE). Compared with the control group, the expression of both genes caused in a similar degree the decrease of living cell numbers and massive occurrence of apoptosis. CONCLUSION　Both reversed caspase-3 and its fusion protein can effectively induce HeLa cells to apoptosis.