【摘要】 目的:探讨MHC-Ⅰ类链相关分子A(MHC class I chain-related molecule A,MICA)多态性与乳腺癌细胞对NK细胞杀伤敏感性的关系。方法:测序分析乳腺癌细胞系MCF-7、MDA-MB-231、MDA-MB-435S和SK-BR-3的MICA等位基因,用Western blotting和流式细胞术检测MICA重组表达载体转染293T细胞(分别命名为p MCFA5.1、p MCFA4、p231A5R、p231A9和p435A5P)MICA蛋白的表达水平,用LDH法检测NK细胞对转染MICA的293T细胞的杀伤活性,酶联免疫斑点法检测NK细胞穿孔素、颗粒酶B分泌水平。结果:MCF-7细胞表达MICA*008/A5.1和MICA*001/A4,MDA-MB-231和SK-BR-3细胞均表达MICA*019/A5和MICA*002/A9,MDA-MB-435S细胞表达MICA*010/A5。转染MICA后,p MCFA5.1组293T细胞MICA蛋白的表达水平最低(P<0.05),p435A5P组次之(P<0.05),p MCFA4组、p231A5R组和p231A9组表达水平较高(均P<0.05)。NK细胞对转染MICA的293T细胞杀伤活性及穿孔素、颗粒酶B分泌:p435A5P组对NK细胞杀伤的敏感性最低(P<0.05),穿孔素、颗粒酶B分泌水平最低(均P<0.05);p MCFA5.1、p MCFA4、p231A5R和p231A9各组之间比较差异无统计学意义(P>0.05)。结论:MICA基因多态性与乳腺癌细胞对NK细胞杀伤的敏感性密切相关。更多还原

【Abstract】 Objective:To investigate the associations of MHC class I chain-related molecule A(MICA)gene polymorphism with susceptibility of breast cancer cells to NK cell-mediated cytotoxicity.Methods:MICA alleles of breast cancer cell lines(MCF-7,MDAMB-231,MDA-MB-435 S and SK-BR-3)were analyzed by DNA sequencing.MICA protein expression in 293 T cells transfected with MICA recombinant expression vectors(namely p MCFA5.1,p MCFA4,p231 A5 R,p231 A9 and p435 A5 P) was detected by Western blotting and Flow cytometry;the cytotoxicity of NK cells against the above 293 T cells were measured by lactate dehydrogenase(LDH)assay and the release of perforin(PFN)and granzymes B(Gzm B)were measured by ELISPOT assay.Results:DNA sequencing result showed that MICA*008/A5.1 and MICA*001/A4 were expressed in MCF-7 cells,MICA*019/A5 and MICA*002/A9 were expressed in both MDA-MB-231 and SK-BR-3 cells while MICA*010/A5 was expressed in MDA-MB-435 S cells.After the MICA recombinant expression vectors were transfected into 293 T cells,the level of MICA were the lowest in p MCFA5.1 group(P<0.05),following after p435 A5 P group(P<0.05),and highly expressed in the p MCFA4,p231 AR and p231 A9 groups(P<0.05).NK cell-mediated cytotoxicity and the release of PFN and Gzm B:NK cell-mediated cytotoxicity were the lowest in p435 A5 P group(P<0.05),and the ability of inducing NK cells to release PFN and Gzm B was also the lowest in p435 A5 P group(P<0.05),but there were no statistical difference among the other transfected groups(P>0.05).Conclusion:MICA gene polymorphism is closely associated with susceptibility of breast cancer cells to NK cell-mediated cytotoxicity.更多还原