重组人肿瘤坏死因子-α联合肿瘤坏死因子受体1基因转染杀伤舌癌细胞的实验研究

Gene transfection of recombinant human tumor necrosis factor-α with tumor nec rosis factor-receptor-1 inhibit the proliferation of tongue carcinoma Tca 8113 cells

目的 :观察重组人肿瘤坏死因子 α(rhTNF α)对建系细胞 (T T)作用效应与机制。方法 :反转录PCR方法克隆人TNFR1基因 ,并将其稳定转染入舌癌细胞中 ,建系并鉴定 ;通过MTT法检测、DNA降解片段琼脂糖凝胶电泳及电镜观察检测rhTNF α对T T细胞的作用效果与机制。结果 :成功构建含TNFR1基因的真核表达质粒 pcDNA3 TNFR1,并建立表达TNFR1蛋白活性的舌癌细胞系T T ;较之亲本Tca 8113细胞及转染空载体的T pc细胞 ,rhTNF α对T T细胞具有更强的细胞毒作用 ,且通过介导细胞凋亡的方式发挥其作用。结论 :重组人肿瘤坏死因子 α联合肿瘤坏死因子受体 1基因转染可有效地杀伤舌癌细胞 ,为舌癌基因治疗提供理论依据。

Objective: To study the effects of tumor necro si s factor receptor 1 (TNFR1) and recombinant human tumor necrosis factor-α(rhT NF-α) on the proliferation of human tongue carcinoma Tca8113 cells. Methods: TNFR1 gene cDNA was amplified by RT-PCR and clone into pcDNA3 vector, then transfected into Tca8113 cells. TNFR 1 protein in t he gene-transfected cells was detected by Western blot, the effect of rhTNF-α on the proliferation of the cells was studied by MTT ussay, DNA fragmentation w as observed by agarose gel eletrophoresis assay and cell morphology was studied by electron microscope. Results: pcDNA3-TNFR1 was transfeted in to Tca8113 cells and the expression of TNFR 1 protein was confirmed by W estern blot. The gene transfected cells were more sensititve to the growth inhi bition of rhTNF-α than those in the parental cells and the vector transfected cells. In the TNFR 1 transfected cells, DNA fragments and apoptotic changes were observed after the cells had been treated with rhTNF-α.Conclusion: TNFR1 gene transfection may induce Tca8113 cells to be more sensit ive to rhTNF-α. The growth inhibition may be related to apoptosic induction.