miR-296-5p靶向MICB促进口腔鳞癌生长的分子机制  

Molecular mechanism of miR-296-5p promoting growth of oral squamous cell carcinoma by targeting MICB

作　　者：王藜篥 张涛(指导) 金珊 周祥文[1,3] 王青 姚雨彤[1] 杨扉扉 WANG Li-Li;ZHANG Tao;JIN Shan;ZHOU Xiang-Wen;WANG Qing;YAO Yu-Tong;YANG Fei-Fei(Department of Stomatology,Guizhou Orthopedics Hospital,Guiyang 550004,China)

机构地区：[1]贵州省骨科医院,贵阳550004 [2]贵州省人民医院,贵阳550000 [3]遵义医学院附属口腔医院,遵义563000 [4]贵阳市口腔医院,贵阳550000

出　　处：《中国免疫学杂志》2019年第18期2243-2248,共6页Chinese Journal of Immunology

基　　金：2019年度贵阳市科学技术局大健康科技计划(No.2019LH);贵州省科学技术厅基础研究计划(黔科合基础[2019]1182号)

摘　　要：目的:研究微小RNAmiR-296-5p对口腔鳞癌细胞存活及自然杀伤细胞因子分泌的影响,并探讨其机制。方法:运用免疫组织化学方法检测癌旁组织、口腔鳞癌组织中MICB的蛋白表达;qRT-PCR法检测组织和口腔鳞癌细胞Cal27中miR-296-5p的表达;将anti-miR-296-5p组(转染anti-miR-296-5p)、miR-296-5p组(转染miR-296-5pmimics)、pcDNA3.1-MICB组(转染pcDNA3.1-MICB)、anti-miR-296-5p+si-MICB组(anti-miR-296-5p和si-MICB共转染),均用脂质体转染至Cal27细胞;MTT法检测各组细胞的存活率;ELISA法检测各组细胞中TNF-α、IFN-γ的含量;Westernblot检测各组细胞中MICB的蛋白表达;双荧光素酶报告基因检测实验检测各组细胞的荧光活性。结果:与癌旁组织相比,口腔鳞癌组织中MICB的蛋白表达显著降低,miR-296-5p的表达显著升高(P<0.05);抑制miR-296-5p、过表达MICB均可显著下调Cal27细胞的存活率,上调NK细胞因子TNF-α、IFN-γ的含量;miR-296-5p可抑制野生型MICB细胞的荧光活性,且负向调控MICB的表达;敲减MICB可逆转抑制miR-296-5p对Cal27细胞存活率的下调及对NK细胞因子含量的上调作用。结论:下调miR-296-5p可抑制口腔鳞癌细胞的存活,促进NK细胞的杀伤作用,其机制可能与靶向负调控MICB有关,其可为口腔鳞癌的靶向治疗提供依据。Objective:To study the effect of miR-296-5p on the survival of squamous cell carcinoma cells and the secretion of NK cytokines,and explore its mechanism.Methods:Immunohistochemistry was applied to detect the protein expression of MICB in normal tissues and oral squamous cell carcinoma tissues.qRT-PCR was used to detect the expression of miR-296-5p in tissues and cells.anti-miR-296-5p group(transfected anti-miR-296-5p),miR-296-5p group(transfected miR-296-5p mimics),pcDNA 3.1-MICB(transfected pcDNA 3.1-MICB),anti-miR-296-5p+si-MICB group(co-transfected anti-miR-296-5p and si-MICB),were all transfected into Cal27 cells with liposome;the survival rate of each group was detected by MTT assay.The level of TNF-αand IFN-γin each group was detected by ELISA;the protein expression of MICB in each group was detected by Western blot.The fluorescence activity of the cells in the group was detected by dual luciferase reporter gene assay.Results:Compared with normal tissues,the protein expression of MICB in oral squamous cell carcinoma was significantly decreased,and the expression of miR-296-5p was significantly increased(P<0.05).The inhibition of miR-296-5p and overexpression of MICB could significantly down-regulate the survival rate of Cal27 cells,up-regulate the levels of NK cytokines TNF-αand IFN-γ;miR-296-5p inhibits the fluorescence activity of wild-type MICB cells and negatively regulates the expression of MICB;knockdown MICB could rescue the inhibition of miR-296-5p on Cal27 cell survival rate of down-regulation,and up-regulation of NK cytokine levels.Conclusion:Down-regulation of miR-296-5p can inhibit the survival of oral squamous cell carcinoma and promote the killing effect of NK cells.The mechanism may be related to the targeted negative regulation of MICB,which will provide a basis for targeted therapy of oral squamous cell carcinoma.

关 键 词：MICB miR-296-5p 口腔鳞癌 NK细胞因子 

分 类 号：R739.85[医药卫生—肿瘤]