CD80-链亲和素融合蛋白锚定的鼻咽癌CNE2细胞增强T细胞的杀伤活性

CD80-streptavidin-decorated CNE2 cells enhance cytotoxicity of T cells

目的:制备CD80-链亲和素(streptavidin,SA)融合蛋白,研究CD80-SA锚定的鼻咽癌CNE2细胞对T细胞杀伤活性的影响。方法:pET21a-CD80-SA-6His质粒转化大肠杆菌BL21(DE3),IPTG诱导CD80-SA融合蛋白的表达,经Ni-NTA亲和层析纯化后透析复性。流式细胞仪检测CD80-SA融合蛋白在CNE2细胞表面的锚定效率;LDH释放法检测CD80-SA锚定的CNE2细胞对T细胞杀伤活性的影响。结果:成功制备和纯化了CD80-SA融合蛋白。CNE2细胞表面低表达CD80分子[(2.2±0.18)%]。CD80-SA融合蛋白可有效地锚定于生物素化的CEN2细胞表面,锚定效率可达73%。CD80-SA锚定的CNE2细胞可有效激活T细胞的杀伤作用,效靶比在1∶1、1∶20、1∶40时T细胞的杀伤率分别约为(37±3.12)%、(51±2.63)%和(58±2.47)%,均显著高于对照CNE2细胞激活的T细胞(均P<0.01)。结论:CD80-SA融合蛋白可有效锚定于生物素化的CNE2细胞表面,从而增强T细胞对CNE2细胞的杀伤活性。

Objective：To prepare CD80-streptavidin（CD80-SA）fusion protein and immobilize it on the surface of nasopharyngeal carcinoma CNE2 cells（CD80-SA-CNE2 cells）,so as to investigate the effect of CD80-SA-CNE2 cells on cytotoxicity of T cells.Methods：pET21a-CD80-SA-6His expression plasmid was transformed into E.coli BL21（DE3）.The CD80-SA fusion protein was induced by IPTG,purified by Ni-NTA affinity chromatography and refolded by dialysis.The immobilization rate of CD80-SA on CNE2 cell surface was analyzed by flow cytometry,and the effect of CD80-SA-CNE2 cells on cytotoxicity of T cells was detected by LDH assay.Results：CD80-SA fusion protein was successfully prepared and purified.CD80 was lowly expressed on CNE2 cells（[2.233±0.176]%）.CD80-SA fusion protein was effectively immobilized on the surface of biotinylated-CNE2 cells,with the immobilization rate being 73%.Moreover,CD80-SA-CNE2 cells effectively induced cytotoxicity of T cells;the cytotoxic rates of T cells were（37±3.12）%,（51±2.63）% and（58±2.47）% at the E∶T ratios of 1∶1,1∶20 and 1∶40,respectively,which were significantly higher than those of control CNE2 cells（all P〈0.01）.Conclusion：CD80-SA fusion protein can be effectively immobilized on the surface of biotinylated-CNE2 cells,enhancing the cyctotoxicty of T cells against CNE2 cells.