微RNA-335对胃癌细胞系SGC-7901细胞增殖、迁移和侵袭的影响及机制

Effect and mechanism of microRNA-335 on proliferation,migration and invasion of gastric cancer cell line SGC-7901

目的探讨微RNA(miR)-335对胃癌细胞系SGC-7901细胞增殖、迁移、侵袭的影响及机制。方法选择2020年1月至2021年7月新乡医学院第三附属医院收治的61例胃癌患者为研究对象,收集患者手术切除的胃癌组织、癌旁组织(距癌灶1~2 cm)及切缘正常组织,采用实时荧光定量聚合酶链式反应法检测人胃癌组织、癌旁组织和正常组织中miR-335的表达。另外,将对数生长期的胃癌细胞系SGC-7901细胞分为miR-335转染组、空白载体组和对照组,miR-335转染组SGC-7901细胞转染miR-335 precursor,空白载体组SGC-7901细胞转染miR-335-NC,对照组细胞不做任何传染。采用四甲基偶氮唑盐法检测3组SGC-7901细胞增殖能力,划痕实验检测3组SGC-7901细胞迁移能力,Transwell小室法检测3组SGC-7901细胞侵袭能力。采用荧光素酶报告基因技术检测miR-335的作用靶点,Western blot法检测3组SCG-7901细胞中p53蛋白的表达。结果miR-335转染组、空白载体组和对照组胃癌组织中miR-335相对表达量显著低于正常组织和胃癌癌旁组织(t=15.238、8.796,P<0.05),癌旁组织与正常组织中miR-335相对表达量比较差异无统计学意义(t=0.293,P>0.05)。miR-335转染组SGC-7901细胞的增殖能力显著低于空白载体组和对照组(t=8.192、9.209,P<0.05),空白载体组与对照组SGC-7901细胞的增殖能力比较差异无统计学意义(t=0.910,P>0.05)。miR-335转染组SGC-7901细胞迁移距离显著短于空白载体组和对照组(t=12.833、12.987,P<0.05),空白载体组与对照组SGC-7901细胞迁移距离比较差异无统计学意义(t=0.623,P>0.05)。miR-335转染组SGC-7901细胞侵袭数目显著少于空白载体组和对照组(t=12.750、14.553,P<0.05),空白载体组与对照组SGC-7901细胞侵袭数目比较差异无统计学意义(t=0.556,P>0.05)。含有p53-3′非翻译区预测基因质粒的miR-335转染组细胞的荧光素酶活性显著低于空白载体组和对照组(t=6.609、7.671,P<0.05),空白载体组与对照组细胞的荧光素酶活性比较差异无统计学意义(t=0.432,P>0.05)。miR-335转染组SGC-7901细胞中p53蛋白相对表达量显著低于空白载体组和对照组(t=7.652、9.227,P<0.05),空白载体组与对照组SGC-7901细胞中p53蛋白相对表达量比较差异无统计学意义(t=1.004,P>0.05)。结论胃癌组织中miR-335呈低表达,miR-335可通过靶向p53抑制胃癌细胞的增殖和侵袭。

Objective To investigate the effect and mechanism of microRNA(miR)-335 on the proliferation,migration and invasion of gastric cancer cell line SGC-7901.Methods A total of 61 patients with gastric cancer admitted to the Third Affiliated Hospital of Xinxiang Medical University from January 2020 to July 2021 were selected as the research objects.The gastric cancer tissues,adjacent tissues(1-2 cm from the cancer focus)and normal tissues at the cutting edge of the patients were collected.The expression of miR-335 in gastric cancer tissues,adjacent tissues and normal tissues was detected by fluorescence quantitative polymerase chain reaction.The gastric cancer cell line SGC-7901 cells in logarithmic growth stage were divided into miR-335 transfection group,blank vector group control group.The SGC-7901 cells in the miR-335 transfection group were transfected with miR-335 precursor,and the SGC-7901 cells in the blank vector group were transfected with miR-335-NC,and the cells in the control group did not have any infection.The proliferation of SGC-7901 cells in the three groups was detected by methyl thiazolyl tetrazolium method,the migration of SGC-7901 cells in the three groups was detected by scratch test,and the invasion of SGC-7901 cells in the three groups was detected by Transwell chamber method.The target of miR-335 was detected by luciferase reporter gene technology,and the expression level of p53 protein in the three groups was detected by Western blot.Results The relative expression of miR-335 in gastric cancer tissue was significantly lower than that in the normal tissue and gastric cancer adjacent tissue(t=15.238,8.796;P<0.05);there was no significant difference in the relative expression of miR-335 between gastric cancer adjacent tissues and normal tissues(t=0.293,P>0.05).The proliferation of SGC-7901 cells in the miR-335 transfection group was significantly lower than that in the blank vector group and control group(t=8.192,9.209;P<0.05),there was no significant difference in the proliferation of SGC-7901 cells between the blank vector group and control group(t=0.910,P>0.05).The migration distance of SGC-7901 cells in the miR-335 transfection group was significantly shorter than that in the blank vector group and control group(t=12.833,12.987;P<0.05),there was no significant difference in the migration distance of SGC-7901 cells between the blank vector group and control group(t=0.623,P>0.05).The number of SGC-7901 cells invasion in the miR-335 transfection group was significantly less than that in the blank vector group and control group(t=12.750,14.553;P<0.05),there was no significant difference in the number of SGC-7901 cell invasion between the blank vector group and the control group(t=0.556,P>0.05).The luciferase activity of the miR-335 transfected cells containing p53-3′-untranslated region prediction gene plasmid was significantly lower than that in the blank vector group and control group(t=6.609,7.671;P<0.05),and there was no significant difference in the luciferase activity of SGC-7901 cell between blank vector group and control group(t=0.432,P>0.05).The relative expression of p53 protein in SGC-7901 cells in the miR-335 transfection group was significantly lower than that in the blank vector group and control group(t=7.652,9.227;P<0.05),there was no significant difference in the relative expression of p53 protein in SGC-7901 cells between the blank vector group and control group(t=1.004,P>0.05).Conclusion The expression of miR-335 is low in gastric cancer,and miR-335 can inhibit the proliferation and invasion of gastric cancer cells by targeting p53.