摘要
目的:探究miR-135b-5p通过靶向CMTM3基因调节胃癌细胞的增殖、侵袭和迁移能力。方法:采用实时荧光定量PCR(RT-qPCR)检测胃癌组织和细胞系中CMTM3和miR-135b-5p的表达水平。利用慢性病毒转染技术将miR-135b-5p inhibitor转染至胃癌细胞,RT-qPCR检测转染效率;CCK-8比色法和Transwell实验检测转染后细胞的增殖、侵袭和迁移能力;Western blotting检测转染后细胞中CMTM3蛋白表达水平。利用生物信息学网站预测miR-135b-5p潜在靶基因CMTM3,利用荧光素酶实验进行验证。结果:RT-qPCR检测结果表明,miR-135b-5p在胃癌组织和胃癌细胞株中呈现高表达,CMTM3在胃癌组织中呈现低表达(P<0.01)。RT-qPCR检测转染效率,结果显示,转染miR-135b-5p inhibitor敲低了miR-135b-5p的表达水平(P<0.001);CCK-8和Transwell实验结果表明,敲低miR-135b-5p后抑制了胃癌细胞的增殖、侵袭和迁移能力(P<0.01),Western blotting实验结果表明,敲低miR-135b-5p促进了CMTM3蛋白的表达。生物信息学网站预测CMTM3为miR-135b-5p的潜在靶基因,荧光素酶实验证实了miR-135b-5p直接靶向胃癌细胞中CMTM3基因(P<0.01)。结论:miR-135b-5p作为重要的调节因子,反向调节抑癌靶基因CMTM3,从而促进胃癌细胞的增殖、侵袭和迁移能力。
Objective:To explore the ability of miR-135 b-5 p to regulate the proliferation,invasion and migration of gastric cancer cells by targeting the CMTM3 gene.Methods:The expression of CMTM3 and miR-135 b-5 p in gastric cancer tissues and cells was detected by RT-qPCR.miR-135 b-5 p inhibitor was transferred into gastric cancer cells by chronic virus transfection technique,and the transfection efficiency was detected by RT-qPCR.CCK-8 colorimetric assay and Transwell test were used to detect the proliferation,invasion and migration ability of gastric cancer cells transfected with miR-135 b-5 p inhibitor,respectively.The expression level of CMTM3 protein after transfection was detected by Western blotting.The potential target gene CMTM3 of miR-135 b-5 p was predicted by the biological information website,and the luciferase experiment was used to verify the results.Results:The results of RT-qPCR showed that miR-135 b-5 p was highly expressed in gastric cancer tissue and gastric cancer cells,and CMTM3 showed low expression in gastric cancer tissue(P<0.01).The results of RT-qPCR detection of transfection efficiency showed that the expression level of miR-135 b-5 p was decreased by transfection of miR-135 b-5 p inhibitor(P<0.001).The results of vitro transfection experiments showed that the proliferation,invasion and migration ability of gastric cancer cells were inhibited by knockdown of miR-135 b-5 p(P<0.01).The results of Western blotting assay showed that knockdown of miR-135 b-5 p promoted the expression of CMTM3 protein.The bioinformatics website predicted that CMTM3 was a potential target gene of miR-135 b-5 p,and the fluciferase test confirmed that miR-135 b-5 p was directly targeted at CMTM3 gene(P<0.01).Conclusion:As an important regulatory factor,miR-135 b-5 p inversely regulates the target gene CMTM3,which promotes the proliferation,invasion and migration of gastric cancer cells.
作者
李永红
刘文明
李伟
董链
杨峥嵘
衷敬华
Li Yonghong;Liu Wenming;Li Wei;Dong Lian;Yang Zhengrong;Zhong Jinghua(Oncology Department,Tianmen First People's Hospital,Hubei Tianmen 431700,China;Hubei Key Laboratory of Occupational Hazard Identification and Control,Wuhan University of Science and Technology,Hubei Wuhan 430081,China;Oncology Department,the First Affiliated Hospital of Gannan Medical University,Jiangxi Ganzhou 341000,China)
出处
《现代肿瘤医学》
CAS
2020年第19期3305-3310,共6页
Journal of Modern Oncology
基金
湖北省卫生健康科研基金资助(编号:WJ2019H211)。