MiR-663b/ALDH6A1轴通过ROS调节口腔鳞状细胞癌细胞增殖  

MiR-663b/ALDH6A1 axis regulating the proliferation of oral squamous cell carcinoma cells by ROS

作　　者：许朗 黄忠卫 鲁大鹏[3] 韩炎 周丽芝[5] XU Lang;HUANG Zhong-wei;LU Da-peng;HAN Yan;ZHOU Li-zhi(Department of Stomatology,Beijing Jishuitan Hospital,Beijing 100035,China;Department of Stomatology,Beijing Royal Hospital of Integrated Traditional Chinese and Western Medicine,Beijing 102200,China;Integrated Emergency Treatment Center,Beijing Stomatological Hospital Affiliated to Capital Medical University,Beijing 100050,China;Department of Stomatology,Wen’an Hospital,Langfang Hebei 065800,China;Department of Stomatology,Eighth Hospital of Tangshan City,Tangshan Hebei 063000,China)

机构地区：[1]北京积水潭医院口腔科,北京100035 [2]北京王府中西医结合医院口腔科,北京102200 [3]首都医科大学附属北京口腔医院急诊综合治疗中心,北京100050 [4]文安县医院口腔科,河北廊坊065800 [5]河北省唐山市第八医院口腔科,河北唐山063000

出　　处：《局解手术学杂志》2022年第9期747-753,共7页Journal of Regional Anatomy and Operative Surgery

基　　金：北京大学口腔医学院青年基金(pkuss20200108)。

摘　　要：目的 探讨microRNA-663b(miR-663b)对口腔鳞状细胞癌(OSCC)细胞增殖的调控作用及机制。方法 利用基因表达汇编(GEO)数据库的OSCC组织芯片数据进行差异表达基因分析;采用基因富集分析(GSEA)探寻OSCC与miR-663b相关的生物学功能。将miR-663b mimic以及醛脱氢酶6家族成员A1(ALDH6A1)过表达质粒转染至人OSCC细胞系HSC-3、CAL-27;采用CCK-8实验、克隆形成实验及裸鼠皮下移植瘤实验检测miR-663b对HSC-3、CAL-27细胞体内、外增殖的影响;采用双荧光素酶报告基因实验验证miR-663b与ALDH6A1的靶向关系。结果 GEO数据库组织芯片数据显示,miR-663b在OSCC细胞中表达上调;GSEA发现miR-663b生物学功能与调节细胞增殖、氧化还原反应及活性氧(ROS)相关。功能试验中,过表达miR-663b显著促进HSC-3、CAL-27细胞的体外增殖、克隆形成能力以及裸鼠皮下移植瘤的生长(P <0.05),下调ALDH6A1蛋白表达和ROS水平(P <0.05),升高NADPH/NADP~+比值(P <0.05);而上调ALDH6A1表达则显著抑制HSC-3、CAL-27细胞体外增殖和和克隆形成能力,上调ALDH6A1蛋白表达和ROS水平(P <0.05),降低NADPH/NADP~+比值(P <0.05)。双荧光素酶报告基因实验结果证实miR-663b能靶向结合ALDH6A1 mRNA的3’UTR区。结论 miR-663b在OSCC细胞中表达上调,并可通过靶向抑制ALDH6A1表达调节ROS水平,进而促进OSCC细胞增殖发挥致癌基因作用。Objective To explore the regulation effect and mechanism of microRNA-663b ( miR-663b) on the proliferation of oral squamous cell carcinoma ( OSCC) cells. Methods The differential expression genes were analyzed by OSCC tissue microarray data from gene expression omnibus ( GEO) database. The biological functions related to miR-663b in OSCC were explored by genomic enrichment analysis ( GSEA) . The plasmids with overexpression of miR-663b mimic and aldehyde dehydrogenase 6 family member A1 ( ALDH6A1) were transfected into human OSCC cell lines ( HSC-3,CAL-27) . The effects of miR-663b on the proliferation of HSC-3 and CAL-27 cells in vivo and in vitro were detected by CCK-8 experiment,clone formation assay and subcutaneous xenograft assay of nude mice. The targeting relationship between miR-663b and ALDH6A1 was verified by dual luciferase reporter gene assay. Results The tissue microarray data from GEO database showed that the expression of miR-663b was up-regulated in OSCC cells. GSEA showed that the biological functions of miR-663b were related to regulating the cells proliferation,redox reactions and reactive oxygen species ( ROS) . In functional test,overexpression of miR-663b significantly promoted the in vitro proliferation and clone formation ability of HSC-3 and CAL-27 cells,as well as the growth of subcutaneous xenograft tumors in nude mice,down-regulated the expression of ALDH6A1 protein and ROS levels,and increased NADPH / NADPratio ( P < 0. 05) . However,up-regulation of ALDH6A1 expression significantly inhibited the in vitro proliferation and clone formation ability of HSC-3 and CAL-27 cells,up-regulated the expression of ALDH6A1 protein and ROS levels,and reduced NADPH / NADPratio ( P < 0. 05) . The results of dual luciferase reporter gene assay confirmed that miR-663b could target 3 ’UTR region of ALDH6A1 mRNA. Conclusion The expression of miR-663b is up-regulated in OSCC cells,which can regulate ROS levels by targeting the inhibition of ALDH6A1 expression,thereby promoting the proliferation of OSCC

关 键 词：口腔鳞状细胞癌 miR-663b 醛脱氢酶6家族成员A1 活性氧 增殖 

分 类 号：R246.83[医药卫生—针灸推拿学]