【摘要】 目的:探究白杨素(Chrysin,CR)对食管鳞癌细胞增殖和侵袭的影响及其潜在的分子机制。方法:将食管鳞癌细胞分为对照组(未转染、未CR处理细胞常规培养)、miR-NC组(稳定转染的miR-199a-3p-NC用等量DSMO培养)、miR mimic组(稳定转染的miR-199a-3p mimic用等量DSMO培养)、CR-miR-NC组(稳定转染的miR-199a-3p-NC用终浓度100μmol·L-1CR培养)、CR-miR mimic组(稳定转染的miR-199a-3p mimic用终浓度100μmol·L-1CR培养),48 h后,采用流式细胞仪检测转染效率,采用实时荧光定量PCR(qRT-PCR)检测EC9706细胞中miR-199a-3p和锌指和同源框蛋白1(Zinc-fingers and homeoboxes protein 1,ZHX1)表达情况,采用荧光素活性和蛋白免疫印迹(Western blot)检测miR-199a-3p对ZHX1的调控作用,分别采用MTT和transwell小室实验检测EC9706细胞增殖和侵袭能力。结果:流式细胞仪检测结果显示转染效率达到80%以上;CR-miR-NC组miR-199a-3p的表达水平显著低于miR-NC组(P<0.05);CR-miR mimic组miR-199a-3p的表达水平显著低于miR mimic组(P<0.05),CR-miR-NC组ZHX1的表达水平显著高于miR-NC组(P<0.05);CR-miR mimic组ZHX1的表达水平显著高于miR mimic组(P<0.05)。荧光素酶活性测定显示,在转染ZHX1 3’UTR的细胞中,与miR-199a-3p-NC组相比,miR-199a-3p mimic组荧光素酶活性显著降低(P<0.05),而在转染ZHX13’UTR-Mut的细胞中,miR-199a-3p mimic组与miR-199a-3p-NC组的荧光素酶活性差异无统计学意义(P>0.05),CR-miR-NC组EC9706细胞增殖和侵袭能力显著低于miRNC组(P<0.05),CR-miR mimic组EC9706细胞增殖和侵袭能力显著低于miR mimic组(P<0.05)。结论:CR通过调控miR-199a-3p/ZHX1抑制食管鳞癌细胞增殖和侵袭,为食管鳞癌的靶向治疗提供一定的理论依据。更多还原

【Abstract】 Objective: To investigate the effect of chrysin( CR) on the proliferation and invasion of esophageal squamous carcinoma cells and its potential molecular mechanism. Methods: Esophageal squamous carcinoma cells were divided into five groups: the control group( routine culture of untransfected and untreated CR cells),miR-NC group( stable transfected miR-199 a-3 p-NC was cultured in equal DSMO),miR mimic group( stable transfected miR-199 a-3 p mimic was cultured in equal DSMO),CR-miR-NC group( stable transfected miR-199 a-3 p-NC was cultured with 100 μmol·L-1 CR),and CR-miR mimic group( stable transfected miR-199 a-3 p mimic was cultured with 100 μmol·L-1 CR). After 48 h,the transfection efficiency was detected by flow cytometry,the expressions of miR-199 a-3 p and Zinc-fingers and homeoboxes protein 1( ZHX1) in EC9706 cells were detected by real-time fluorescence quantitative PCR( qRT-PCR),the regulatory effects of miR-199 a-3 p on ZHX1 were detected by fluorescein assay and Western blot assay,and the proliferation and invasion of EC9706 cells were detected by MTT and Transwell chamber experiments,respectively. Results: The results of flow cytometry showed that the transfection efficiency was over 80%; the expression level of miR-199 a-3 p in CRmiR-NC group was significantly lower than that in miR-NC group( P < 0. 05); the expression level of miR-199 a-3 p in CR-miR mimic group was significantly lower than that in miR mimic group( P < 0. 05),and the expression level of ZHX1 in CR-miR-NC group was significantly higher than that in miR-NC group( P < 0. 05); the expression level of ZHX1 in CR-miR mimic group was significantly higher than that in miR mimic group( P < 0. 05). Luciferase activity assay showed that the luciferase activity in the transfected ZHX13 ’UTR cells of miR-199 a-3 p mimic group was significantly lower than that of miR-199 a-3 p-NC group( P < 0. 05). In transfected ZHX13 ’UTR-Mut cells,there was no significant difference in the luciferase activity between miR-199 a-3 p mimic group and miR-199 a-3 p-NC group( P > 0. 05); the proliferation and invasion ability of EC9706 cells in CR-miR-NC group was significantly lower than that in miR-NC group( P < 0. 05),and the proliferation and invasion ability of EC9706 cells in CR-miR mimic group was significantly lower than that in miR mimic group( P < 0. 05). Conclusion: CR can inhibit the proliferation and invasion of esophageal squamous cell carcinoma by regulating miR-199 a-3 p/ZHX1,which provides theoretical basis for the targeted therapy of esophageal squamous cell carcinoma.更多还原