上调miR-21表达水平对胃癌Kato Ⅲ细胞中CD44 mRNA及蛋白表达的影响

Influence of restoration of miR-21 expression level in CD44 mRNA and protein expressions in human gastric cancer Kato Ⅲ cells

目的:探讨上调微小RNA-21(microRNA-21,miR-21)表达对胃癌KatoⅢ细胞中肿瘤干细胞(CSCs)标志物CD44mRNA及蛋白表达的影响,为胃癌细胞的恶性行为相关研究提供依据。方法:常规培养胃癌KatoⅢ细胞株,分为空白对照组和实验组,实验组又分为KatoⅢ/miR-21组和KatoⅢ/control组,分别予以miR-21模拟物(hsa-miR-21mimics)或其阴性对照(hsa-miR-21control)转染KatoⅢ细胞;通过荧光定量RT-PCR和Western blotting法分别检测CD44mRNA和蛋白在KatoⅢ/control组细胞和KatoⅢ/miR-21组细胞中的表达水平。结果:miR-21在KatoⅢ/control组细胞和KatoⅢ/miR-21组细胞中的表达水平分别为0.109 91±0.013 99和1.000 00±0.107 41,两者比较差异有统计学意义(P<0.01);CD44mRNA在KatoⅢ/control组细胞和KatoⅢ/miR-21组细胞中的相对表达水平分别为0.674 58±0.064 53和2.342 23±0.223 47,两者比较差异有统计学意义(P<0.01);CD44蛋白在KatoⅢ/control组细胞和KatoⅢ/miR-21组细胞中的相对水平分别为0.045 929±0.007 396和0.176 048±0.009 900,两者比较差异有统计学意义(P<0.01)。结论:外源性上调miR-21表达水平能够诱导KatoⅢ细胞中CD44mRNA及蛋白的过表达,可能诱导CD44+胃癌CSCs的转化。

Objective To investigate the influence of restoration of oncogenic microRNA-21（miR-21）in CD44 mRNA and protein expressions,one of the most common cancer stem cell（CSC）markers,in human gastric cancer KatoⅢcells,and to provide a new theoretical foundation for research on gastric cancer.Methods Human gastric cancer KatoⅢcells were cultured and divided into blank control group,KatoⅢ/miR-21group and KatoⅢ/control group.Hsa-miR-21mimics and hsa-miR-21control were transfected into Kato Ⅲ cells in Kato Ⅲ/miR-21group and KatoⅢ/control group.Then the expressions of CD44mRNA and protein in the above cells were detected by RT-PCR and Western blotting method.Results The relative expression levels of miR-21in KatoⅢ/miR-21cells was 1.000 00±0.107 41,which was significantly increased compared with Kato Ⅲ/control cells（0.109 91± 0.013 99）,there was significant difference（P0.01）.The mRNA level of CD44in Kato Ⅲ/miR-21cells was 2.342 23±0.223 47,which was significantly increased compared with Kato Ⅲ/control cells（0.674 58± 0.064 53）,there was significant difference（P0.01）.The protein level of CD44in Kato Ⅲ/miR-21cells was 0.176 048±0.009 900,which was also significantly increased as compared with Kato Ⅲ/control cells（0.045 929±0.007 396）,there was significant difference（P 0.01）.Conclusion Restoration of miR-21 expression level can induce the over-expressions of CD44mRNA and protein in gastric cancer Kato Ⅲ cells,and may induce the transformation of gastric cancer CSCs.