摘要
目的研究miR-195通过靶向调控SerpinA3促进胃癌细胞增殖、转移及侵袭的分子机制。方法选取胃癌MGC803及NCI-N87细胞株为研究对象,分别转染miR-195及RNA阴性对照序列。采用实时荧光定量PCR法检测细胞中miR-195的表达水平;MTT检测转染miR-195对肿瘤细胞分裂增殖能力的影响;Transwell侵袭实验分析miR-195对细胞侵袭力的作用;细胞划痕实验检测miR-195对细胞转移力的作用;流式细胞仪检测并评价转染miR-195对肿瘤细胞分裂周期的影响;Western blot探究miR-195对肿瘤细胞SerpinA3表达的调节;采用Spearman相关分析法分析miR-195及SerpinA3表达的相关性;荧光素酶实验验证miR-195与SerpinA3的靶向关系。结果 MTT实验表明高表达miR-195可明显抑制细胞的体外增殖能力;Transwell侵袭实验结果提示转染miR-195可导致细胞侵袭能力明显降低;流式细胞术结果表明,转染miR-195后G1和G2期细胞增加,S期细胞减少;划痕试验结果表明,转染miR-195可以抑制细胞的迁移能力;Western blot结果显示,miR-195-mimics组细胞SerpinA3蛋白含量低于miR-NC组(P <0.05);miR-195 mRNA水平与SerpinA3蛋白含量负相关(r=-0.464,P <0.01);miR-195可直接靶向SerpinA3。结论 miR-195可抑制胃癌恶性行为,其机制与调控SerpinA3有关。
Objective To explore the molecular mechanism of miR-195 targeting SerpinA3 inhibiting proliferation,apoptosis,invasion and metastasis of gastric cancer cells.Methods Gastric cancer cell lines MGC803,NCI-N87 were transfected with miR-195 and RNA-negative control sequences respectively.The expression of miR-195 in cells was detected by real-time fluorescence quantitative PCR.The effect of transfected miR-195 on the proliferation of tumor cells was detected by MTT assay.Cell invasion and metastasis was detected by transwell invasion test and cell scratch test.Cell metastasis was detected by flow cytometry.The protein levels of CHEK1 were determined by western blot.The correlation between the expressions of miR-195 and SerpinA3 were analysis by Spearman.The targeted relationship of miR-195 and SerpinA3 was verified by luciferase assay.Results MTT assay showed that high expression of miR-195 could significantly inhibit the proliferation of cells in vitro.Transwell invasion test showed that miR-195 could significantly decrease the invasive ability of cells.Flow cytometry showed that G1 and G2 phase cells increased and S phase cells decreased after miR-195 transfer.The results of scratch test showed that miR-195 could inhibit the migration of cells.Western blot results showed that the content of SerpinA3 protein in miR-195-mimics group was lower than that in miR-NC group.The level of miR-195 mRNA was negatively correlated with the content of SerpinA3 protein(r=-0.464,P<0.01),and miR-195 could directly target SerpinA3.Conclusion MiR-195 can inhibit the malignant behavior of gastric cancer,and its mechanism is related to the regulation of SerpinA3.
作者
罗文
李群芳
严超
萧赪
LUO Wen;LI Qunfang;YAN Chao;XIAO heng(Department of Internal Medicine,Hunan University of Environment and Biology,Hengyang 421001,China)
出处
《实用医学杂志》
CAS
北大核心
2019年第23期3588-3592,共5页
The Journal of Practical Medicine
基金
湖南省教育厅资助课题（编号：16C0557）
