miR-3666在香烟烟雾诱导人支气管上皮细胞恶性转化中的表达变化及其功能研究  

Expression and functional analysis of miR-3666 in tobacco exposed bronchial epithelial BEAS-2B transformed cells

作　　者：余晓凡 刘禧越 张炳勇 王力 YU Xiao-fan;LIU Xi-yue;ZHANG Bing-yong;WANG Li(School of Public Health,Bengbu Medical College,Bengbu,Anhui 233030,China)

机构地区：[1]蚌埠医学院公共卫生学院,安徽蚌埠233030

出　　处：《现代预防医学》2023年第18期3402-3407,共6页Modern Preventive Medicine

基　　金：蚌埠医学院自然科学重点项目(2020byzd040);安徽省高等学校自然科学研究项目(2022AH051492);大学生创新创业训练计划项目(20211036048)。

摘　　要：目的探索microRNA-3666(miR-3666)在香烟烟气长期染毒致使的人永生化支气管上皮细胞BEAS-2B恶性转化过程中的表达调控及其功能研究.方法将BEAS-2B细胞以每孔1×105个的浓度种植于六孔板Transwell膜上室,设置细胞气体染毒装置染毒程序为烟雾浓度20%,染毒时间10 min,每4天染毒一次,2次染毒后收集细胞作为染毒1代,依次获得染烟恶性转化第30代(S30)细胞.通过荧光定量PCR(qPCR)技术检测miR-3666在S30恶性转化细胞以及肺腺癌(LUAD)细胞系A549和H1299中的表达量;通过miRNA mimic转染,过表达S30及LUAD细胞系中的miR-3666;通过CCK8法检测过表达miR-3666后S30细胞和LUAD细胞系恶性增殖能力的变化;通过Transwell侵袭实验检测过表达miR-3666对S30细胞及LUAD细胞系侵袭能力的影响;通过流式细胞术检测miR-3666表达对S30细胞及LUAD细胞系凋亡能力的影响.结果与BEAS-2B细胞相比,染烟恶性转化S30细胞和肺腺癌细胞系中miR-3666表达显著下调(F=47.885,P<0.05);CCK8增殖实验显示过表达miR-3666可使S30细胞和肺腺癌细胞系增殖能力显著下降(P<0.05);Transwell侵袭实验结果表明提升miR-3666表达量,S30细胞和肺腺癌细胞系侵袭能力显著下降(P<0.05);流式细胞凋亡实验结果显示,高表达的miR-3666能有效促进S30细胞和肺腺癌细胞系凋亡(P<0.05).结论染烟30代恶性转化细胞中miR-3666通过抑制细胞恶性增殖和迁移,促进细胞凋亡,从而阻碍肿瘤的进展过程,发挥抑癌基因作用.miR-3666具有成为吸烟所致肺癌表观遗传修饰分子标志物的可能.Objective e To investigate the expression and function of microRNA-3666(miR-3666)in tobacco exposed bronchial epithelial BEAS-2B cells that were malignant transformed.Methods BEAS-2B cells were planted in Transwell membrane at 1 x105 concentrations,20%cigarette smoke was injected through the cell poisoning device for 10 minutes per day,and poisoned every 4 days.After two times of poisoning,we got the first generation of poisoning cells,and then the 30th generation(S30)cells of malignant transformation were obtained.The expression level of miR-3666 were detected by qPCR in malignant transformed S30 cells(S30)as well as lung adenocarcinoma(LUAD)cell lines:A549 and H1299,using BEAS-2B cells as contradiction.CCK8 assays were performed to detect the proliferation ability of miR-3666 in S30 and LUAD cell lines.The invasion ability of miR-3666 in S30 and LUAD cell lines were detected by Transwell assay.Flow cytometry was produced to detect cell apoptosis in S30 and LUAD cell lines.Results Compared to normal BEAS-2B cells,the expression levels of miR-3666 were significantly down-regulated in S30 and LUAD cell lines(F=47.885,P<0.05).CCK8 assay revealed that overexpressed miR-3666 could significantly reduce the proliferation of S30 and LUAD cell lines(P<0.05).Transwell assay showed that with high miR-3666 expression,the invasion ability of S30 and LUAD cell lines significantly reduced(P<0.05).Flow cytometry assay revealed that with high miR-3666 expression,S30 and LUAD cell lines apoptosis rate effectively promoted(P<0.05).Conclusion Our data suggests that miR-3666 functions as a tumor suppressor by inhibiting the malignant proliferation and migration as well as promoting apoptosis of S30 cells,and this miRNA can be a molecular marker for epigenetic modification of lung cancer.

关 键 词：香烟 肺癌 恶性转化 miR-3666 分子标志物 

分 类 号：R734.2[医药卫生—肿瘤]