miR-515-5p通过靶向HDAC2影响食管癌发生发展的分子机制

miR-515-5p effects the carcinogenesis of esophageal cancer via targeting HDAC2 and its molecular mechanism

目的:探讨miR-515-5p对食管癌细胞增殖、迁移和侵袭的影响及其分子机制。方法:选取2020年6月至2020年12月在河北医科大学第四医院手术切除的60例食管癌患者的癌组织标本和20例健康成人的食管上皮组织标本,以及食管癌细胞TE1、Eca109、KYSE30和KYSE170,用q PCR法检测食管癌组织和细胞中miR-515-5p的表达水平。在Eca109细胞中转染miR-515-5p模拟物以及其阴性对照物、在TE1细胞中转染miR-515-5p抑制剂以及其阴性对照物,q PCR法检测转染效率,用CCK-8法、Transwell实验分别检测转染细胞的增殖、迁移及侵袭能力。采用生物信息学方法分析预测miR-515-5p的下游靶基因,双荧光素酶报告基因实验验证组蛋白去乙酰化酶2(histone deacetylase 2,HDAC2)为miR-515-5p的靶基因。应用GEPIA和TCGA数据集分析HDAC2在食管癌组织中的表达及其与患者临床特征的关系。结果:miR-515-5p在食管癌组织及细胞中表达降低(均P<0.01)。过表达miR-515-5p抑制食管癌Eca109细胞的增殖、迁移和侵袭能力(P<0.05或P<0.01),而敲低miR-515-5p表达则可增强食管癌TE1细胞的增殖、侵袭和迁移能力(均P<0.05)。双荧光素酶报告基因分析证明,HDAC2是miR-515-5p的靶基因。q PCR和WB实验结果显示,miR-515-5p对HDAC2 m RNA和蛋白表达具有负调控作用。挽救实验证实miR-515-5P通过靶向HDAC2抑制食管癌Eca109细胞的增殖、迁移和侵袭能力(P<0.05或P<0.01)。结论:miR-515-5p通过靶向HDAC2影响食管癌细胞的增殖、迁移和侵袭能力。

Objective: To investigate the effects of miR-515-5p on proliferation, migration and invasion of esophageal cancer(EC)cells and the molecular mechanism. Methods: Cancer tissue specimens from 60 patients with esophageal cancer that surgically resected at the Fourth Hospital of Hebei Medical University from June 2020 to December 2020, esophageal epithelial tissue specimens from 20healthy adults, as well as esophageal cancer cell lines(TE1, Eca109, KYSE30 and KYSE170) were selected for this study. Quantitative reverse transcription polymerase chain reaction(qPCR) was performed to detect miR-515-5p expression in above mentioned EC tissues and cells. Eca109 cells were transfected with miR-515-5p mimics or its negative controls, and TE1 cells were transfected with miR-515-5p inhibitors or its negative controls. The transfection efficiency was detected by qPCR, and the proliferation, migration and invasion of transfected cells were detected by CCK-8 assay and transwell assay respectively. Bioinformatics tools were used to predict the downstream target genes of miR-515-5p, and HDAC2(histone deacetylase 2) was verified to be a target gene of miR-515-5p by dual luciferase reporter gene assay. GEPIA and TCGA databases were used to analyze the expression of HDAC2 in EC tissues and its correlation with clinical characteristics of EC patients. Results: miR-515-5p was downregulated in EC tissues and cells(all P<0.01).Over-expression of miR-515-5p inhibited the proliferation, migration and invasion ability of Eca109 cells(P<0.05 or P<0.01), while down-regulation of miR-515-5p enhanced the proliferation, migration and invasion ability of TE1 cells(both P<0.05). Furthermore,HDAC2 was demonstrated to be a target gene of miR-515-5p by dual luciferase reporter gene assay. qPCR and Western blotting results showed that miR-515-5p negatively regulated the m RNA and protein expression of HDAC2. Rescue experiments confirmed that miR-515-5p inhibited the proliferation, migration and invasion ability of esophageal cancer Eca109 cells by targeting HDAC2(P<0.05or P<0.01). Conclusion: miR-515-5p affects proliferation, migration and invasion of EC cells by targeting HDAC2.