生物钟基因Per1对鼻咽癌细胞CNE2侵袭迁移及与JAK2-STAT3通路相关蛋白表达的影响  

Effects of biological clock gene Per1 on the invasion and migration of nasopharyngeal carcinoma cells CNE2 and the expression of JAK2-STAT3 pathway-related proteins

作　　者：罗盼 吴伟莉 金风 龙金华 李媛媛 曾艳 王子琪 叶永英 陈宇 张芒 唐红 LUO Pan;WU Weili;JIN Feng;LONG Jinhua;LI Yuanyuan;ZENG Yan;WANG Ziqi;YE Yongying;CHEN Yu;ZHANG Mang;TANG Hong(Department of Oncology,Affiliated Cancer Hospital of Guizhou Medical University/Affiliated Hospital of Guizhou Medical University,Guizhou Guiyang 550000,China)

机构地区：[1]贵州医科大学附属肿瘤医院/贵州医科大学附属医院肿瘤科,贵州贵阳550000

出　　处：《现代肿瘤医学》2023年第15期2823-2828,共6页Journal of Modern Oncology

基　　金：国家自然科学基金项目(编号:82060555);贵州省卫生健康委科学技术基金项目(编号:gzwj-2022-022);贵州医科大学国家自然资金培育项目(编号:19NSP047)。

摘　　要：目的:探讨生物钟基因Per1对鼻咽癌细胞CNE2侵袭转移及上皮-间质转化(EMT)的影响,并阐述其与JAK2-STAT3信号通路相关蛋白的关系。方法:实时荧光定量PCR法(qPCR)以及蛋白质印迹法(WB)检测CNE2细胞中Per1基因及蛋白水平;构建Per1基因过表达及干扰慢病毒载体并转染CNE2细胞;实验分为过表达组(Per1-OE)、过表达对照组(Per1-OENC)、干扰组(Per1-sh)、干扰对照组(Per1-shNC);利用划痕愈合、Transwell实验检测Per1基因对各组细胞侵袭、迁移能力;WB检测Per1基因对EMT相关蛋白:E-cadherin、N-cadherin、Vimentin表达的影响;WB检测Per1基因对JAK2-STAT3信号通路相关蛋白表达的影响。结果:WB和PCR结果显示:与人永生化鼻咽上皮细胞NP69相比,Per1在鼻咽癌细胞CNE2中表达降低。通过构建Per1基因过表达及干扰慢病毒载体,有效地上调和下调了CNE2中Per1在转录及蛋白水平的表达,并分别成功筛选出稳转细胞株。划痕实验结果显示:Per1-OE组较Per1-OENC组24 h、48 h细胞迁移率均明显下降(P=0.04,P=0.01),Per1-sh组较Per1-shNC组24 h、48 h细胞迁移率均明显升高(P=0.01,P=0.005)。Transwell实验结果显示:Per1-sh组较Per1-shNC组穿过Transwell小室细胞数明显增加(P=0.02),Per1-OE组较Per1-OENC组穿过Transwell小室细胞数明显减少(P=0.001)。WB结果显示:JAK2、p-JAK2、STAT3蛋白在各组细胞中表达无统计学差异(P>0.05)。Per1-sh组较Per1-shNC组p-STAT3、N-cadherin、Vimentin的蛋白表达下降,E-cadherin表达升高(P<0.05),Per1-OE组较Per1-OENC组p-STAT3、N-cadherin、Vimentin的蛋白表达均升高,E-cadherin表达下降(P<0.05)。结论:过表达Per1基因后可抑制鼻咽癌细胞CNE2侵袭、迁移,使N-cadherin、Vimentin、p-STAT3蛋白表达下降,E-cadherin表达增加;干扰Per1基因后则相反,进而表明Per1基因在鼻咽癌细胞CNE2中可能扮演着抑癌基因的角色;同时我们推测Per1基因与p-STAT3的表达有一定的相关性,但其具体作用机制有待Objective:To explore the the effects of biological clock gene Per1 on invasion and epithelial-mesenchymal transition(EMT)of nasopharyngeal carcinoma cells CNE2 and its relationship with the expression of JAK2-STAT3 signaling pathway-related proteins.Methods:Per1 gene and protein levels in CNE2 cells were detected by real-time fluorescence quantitative PCR(qPCR)and Western blot(WB).Per1 gene overexpression and interference lentivirus vector was constructed and transfed into CNE2 cells.The experiment was divided into overexpression group(Per1-OE),overexpression control group(Per1-OENC),interference group(Per1-sh),interference control group(Per1-shNC).The ability of Per1 gene to invade and migrate cells in each group was detected by scratch healing and Transwell test.WB was used to detect the effect of Per1 gene on the expression of EMT-related proteins:E-cadherin,N-cadherin and Vimentin.WB detected the effects of Per1 gene on the espression of JAK2-STAT3Y signaling pathway-related proteins.Results:WB and PCR results showed that compared with human immortalized nasopharyngeal epithelial cell NP69,the expression of Per1 in nasopharyngeal carcinoma cell CNE2 was decreased.By constructing Per1 gene overexpression and interfering with lentivirus vector,effectively up-regulates and down-regulates the expression of Per1 in the transcription and protein levels of CNE2,and successfully screened the stable transformation cell lines.The scratch test results showed that the cell migration rate of Per1-OE group was significantly lower than that of Per1-OENC group at 24 h and 48 h(P=0.04,P=0.01),and that of Per1-sh group was significantly higher than that of Per1-shNC group at 24 h and 48 h(P=0.01,P=0.005).The results of Transwell experiment showed that the number of cells passing through Transwell cells in Per1-sh group was significantly higher than that in Per1-shNC group(P=0.02),and the number of cells passing through Transwell cells in Per1-OE group was significantly lower than that in Per1-OENC group(P=0.001).WB results sho

关 键 词：鼻咽癌细胞CNE2 生物钟基因Per1 JAK2-STAT3信号通路 侵袭迁移 

分 类 号：R739.62[医药卫生—肿瘤]