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Received:June 21, 2022 Published Online:September 20, 2022
Received:June 21, 2022 Published Online:September 20, 2022
中文摘要: 目的 探讨短发夹RNA(shRNA) 靶向沉默接触蛋白-1(CNTN1)基因表达对人乳腺癌MDA-MB-468细胞凋亡及迁移侵袭能力的影响及其机制。方法 将人乳腺癌MDA-MB-468细胞株分为3组:常规更换培养基的为空白组,转染无义shRNA片段的为空载体组,转染靶向沉默CNTN1表达的shRNA载体片段的为沉默组。采用western blot 法检测CNTN1蛋白的表达水平。通过流式细胞技术、细胞划痕以及Transwell实验分别检测凋亡率和迁移、侵袭能力的改变,采用western blot 法检测细胞中E-钙黏蛋白(E-cadherin)和上皮细胞间质化相关基因蛋白[锌指E盒结合蛋白1 (ZEB-1)、波形蛋白(Vimentin)、扭曲蛋白(Twist)、锌指转录因子(Snail) ]的表达情况。结果 与空白组和空载体组比较,沉默组MDA-MB-468细胞中CNTN1蛋白表达水平明显降低(P<0.05),凋亡率增加(P<0.05),迁移和侵袭能力均明显减弱(P<0.05),ZEB-1、Vimentin、Twist和Snail的表达水平明显被抑制,而E-cadherin表达水平明显升高。结论 沉默CNTN1表达可通过调节上皮细胞间质转化促进乳腺癌细胞株MDA-MB-468凋亡增加,迁移和侵袭能力减弱。
Abstract:Objective To investigate the effect of short hairpin RNA (shRNA) targeted silencing of CNTN1 gene expression on apoptosis, migration and invasion of human breast cancer MDA-MB-468 cells and its mechanism. Methods Human breast cancer MDA-MB-468 cell lines were divided into three groups: blank group replaced with routine culture medium, empty vector group transfected with nonsense shRNA fragment, and silencing group transfected with shRNA vector fragment targeting silencing CNTN1 expression. The expression level of CNTN1 protein was detected by western blot. The changes of apoptosis rate, migration and invasion ability were detected by flow cytometry, cell scratch and transwell experiment. Western blot was used to detect the expressions of E-cadherin, and epithelial interstitial related genes [zinc finger E-box binding protein 1 (ZEB-1), Vimentin, Twist protein, zinc finger transcription factor (Snail)]. Results The expression level of CNTN1 protein in MDA-MB-468 cells in the silencing group decreased significantly (P<0.05), the apoptosis rate increased (P<0.05), and the ability of migration and invasion decreased significantly (P<0.05). The expression levels of ZEB-1, Vimentin, Twist and Snail in the silencing group were significantly inhibited, while the expression level of E-cadherin increased significantly (P<0.05). Conclusion Silencing CNTN1 expression promotes the increase of apoptosis and the decrease of migration and invasion of breast cancer cell line MDA-MB-468 by regulating epithelial mesenchymal transformation.
文章编号: 中图分类号:R737.9 文献标志码:A
基金项目:陕西省重点研发计划项目(2021SF-218);陕西省自然科学基础研究计划项目(2020JM-680);陕西省肿瘤医院院内国家自然科学基金孵育项目(SC211007)
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