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肿瘤坏死因子相关凋亡诱导配体(TNF-related apoptosis inducing ligand,TRAIL)是TNF超家族中的成员,能够广泛诱导肿瘤细胞凋亡,对正常细胞无明显毒副作用.TRAIL已成为肿瘤治疗领域的研究热点.人脑胶质瘤是神经系统肿瘤中最常见类型,占颅内肿瘤50%～60%,5年存活率为20%～30%.本研究探讨可溶性TRAIL蛋白对人脑胶质瘤细胞(U251)的抑制作用.由大肠杆菌表达系统表达的TRAIL多为包涵体,为获得可溶性的蛋白,将hTRAIL95～281功能区基因片段插入到pHisSUMO表达载体,经IPTG低温诱导表达,Ni-NTA Agarose纯化后获得可溶性SUMO-hTRAIL,经SUMO ProteaseⅠ切去SUMO融合标签后获得成熟可溶hTRAIL蛋白.以U251细胞为靶细胞,通过MTT法检测TRAIL对肿瘤细胞的抑制作用.结果证明,TRAIL对U251细胞的抑制呈剂量依赖关系,最大抑制率为53.9%.流式细胞仪检测TRAIL诱导U251细胞凋亡实验中,对照组细胞存活率为92.2±0.8%,实验组细胞存活率为35.5±1.2%,证明重组蛋白具有生物学活性,并在体外能明显诱导U251肿瘤细胞发生死亡.本研究结果为TRAIL蛋白在临床上应用于肿瘤治疗奠定了基础.
Tumor necrosis factor-related apoptosis inducing ligand (TNF-related apoptosis inducing ligand, TRAIL) is a member of the TNF superfamily, can widely induce tumor cell apoptosis, no obvious side effects on normal cells.TRAIL has become a field of cancer treatment Research focus.Globosarcoma is the most common type of nervous system tumors, accounting for 50% to 60% of intracranial tumors, 5-year survival rate of 20% to 30% .This study was to explore soluble TRAIL protein on human glioma (U251) .According to the expression system of E.coli, TRAIL was mostly inclusion body. To obtain soluble protein, the hTRAIL95 ~ 281 functional region gene fragment was inserted into pHisSUMO expression vector and induced by low temperature induction of IPTG. Ni-NTA Soluble SUMO-hTRAIL was obtained after purification by Agarose, and mature soluble hTRAIL protein was obtained after SUMO fusionase was removed by SUMO Protease I. U251 cells were used as target cells, and the inhibitory effect of TRAIL on tumor cells was detected by MTT assay. The inhibition of cells in a dose-dependent manner, the maximum inhibition rate was 53.9%. Flow cytometry TRAIL induced apoptosis in U251 cells, the control group cell survival rate was 92.2 ± 0.8%, the experimental group Cellular survival was 35.5 ± 1.2%, have demonstrated biologically active recombinant protein, and in vitro U251 tumor could induce cell death. The results of this study for the treatment of tumors TRAIL protein applied in clinical basis.