circ_PITX1/miR-129-5p对食管癌细胞增殖凋亡及放射敏感性的影响

Effect of circ_PITX1/miR-129-5p on proliferation,apoptosis and radiosensitivity of esophageal cancer

目的探讨circ_PITX1对食管癌细胞增殖凋亡及放射敏感性的影响及分子机制。方法选取2018年1月至2021年1月51例食管癌患者癌组织及癌旁组织,运用实时荧光定量PCR(RT-qPCR)测定circ_PITX1和miR-129-5p的表达;将食管癌细胞EC109分为对照组、干扰circ_PITX1组、miR-129-5p Inhibitor组、干扰circ_PITX1+miR-129-5p Inhibitor组、2 Gy组、2 Gy+干扰circ_PITX1组、2Gy+干扰circ_PITX1+miR-129-5p Inhibitor组;四甲基偶氮唑盐比色法(MTT)检测细胞增殖抑制率;流式细胞术检测细胞凋亡率;克隆形成实验检测细胞克隆数;蛋白质印迹(Western blot)法检测蛋白表达;circ_PITX1和miR-129-5p的靶向关系通过双荧光素酶报告实验分析。结果食管癌组织中circ_PITX1表达比癌旁组织增加,miR-129-5p表达比癌旁组织减少(P<0.05)。单独沉默circ_PITX1或沉默circ_PITX1后用射线照射,EC109细胞抑制率和凋亡率升高,克隆数减少,Cleaved-caspase3和Cleaved-caspase9表达水平升高(P<0.05)。circ_PITX1靶向调控miR-129-5p的表达。抑制miR-129-5p可逆转沉默circ_PITX1对EC109细胞增殖、凋亡及放射敏感性的影响。结论沉默circ_PITX1通过靶向食管癌细胞的miR-129-5p,抑制细胞增殖,促进凋亡及增强其放射敏感性。

Objective To explore the effect of circ_PITX1 on proliferation,apoptosis and radiosensitivity of esophageal cancer(EC)and its molecular mechanism.Methods Totally 51 cases cancerous tissues and adjacent tissues were collected which offered by EC patients in our hospital from January 2018 to January 2021.The expression levels of circ_PITX1 and miR-129-5p were assessed by real-time fluorescent quantitative PCR(RT-qPCR);The esophageal EC109 cells were allocated to control group,interference circ_PITX1 group,miR-129-5p Inhibitor group,interference circ_PITX1+miR-129-5p Inhibitor group,2Gy group,2Gy+interference circ_PITX1 group,2Gy+interference circ_PITX1+miR-129-5p Inhibitor group.Methyl thiazolyl tetrazolium(MTT)was performed to detect proliferation inhibition rate;flow cytometry was used to detect cell apoptosis rate;clone formation experiment was applicated to detect the number of cell clones;Western blot method was conducted to detect protein expression;the targeting relationship between circ_PITX1 and miR-129-5p was determined by dual luciferase reporter assays.Results The significantly higher expression of circ_PITX1 and significantly lower expression of miR-129-5p in esophageal cancer tissue than those in adjacent tissues(P<0.05).Silencing of circ_PITX1 alone or silencing circ_PITX1 and then irradiated with radiation.The inhibition rate and apoptosis rate of EC109 cells were increased,the number of clones was decreased,and the expression levels of Cleaved-caspase3 and Cleaved-caspase9 were increased(P<0.05);circ_PITX1 targeted regulation the expression of miR-129-5p,it can reverse the effects of silencing circ_PITX1 on the proliferation,apoptosis and radiosensitivity of EC109 cells by Inhibition of miR-129-5p.Conclusion Silencing of circ_PITX1 can inhibit the proliferation of esophageal cancer cells by upregulating miR-129-5p,promote cell apoptosis and enhance the radiosensitivity of cells.