摘要
背景与目的:膀胱癌是常见的泌尿系统恶性肿瘤之一,蛋白质精氨酸甲基转移酶7(protein arginine methyltransferase 7,PRMT7)在消化道肿瘤中的作用已有研究报道,但在膀胱癌中的生物学作用及机制尚未明确,本文旨在探究PRMT7对人膀胱癌细胞增殖和迁移能力的影响及其可能的作用机制。方法:体外培养人膀胱癌细胞系5637、T24、RT112、UM-UC-3和输尿管上皮永生化细胞系SV-HUC-1,采用蛋白质印迹法(Western blot)检测PRMT7的表达情况。采用RNA干扰技术沉默PRMT7基因的表达,设立阴性对照组(si-NC)及实验组(siPRMT7#1、siPRMT7#2),采用质粒转染法过表达PRMT7基因,设立阴性对照组(p-PCMV3)及实验组(p-PRMT7)。采用实时荧光定量聚合酶链反应(real-time fluorescence quantitative polymerase chain reaction,RTFQ-PCR)及Western blot验证PRMT7的转染效率,采用细胞计数试剂盒-8(cell counting kit-8,CCK-8)和集落形成实验检测细胞增殖能力,采用transwell实验检测细胞迁移能力,采用Western blot检测增殖和迁移相关靶蛋白Cyclin D1、CDK4和MMP9以及Notch信号转导通路关键蛋白Notch1、HEY1和Hes1的表达情况。使用转染siPRMT7#2的细胞加入Notch信号特异性抑制剂γ-分泌酶抑制剂DAPT,进一步验证PRMT7在膀胱癌中的表达情况。结果:Western blot结果显示,PRMT7在膀胱癌细胞中低表达(P<0.05)。在5637细胞中沉默PTMT7后,细胞增殖和迁移能力显著增强(P<0.05),增殖和迁移相关靶蛋白Cyclin D1、CDK4和MMP9以及Notch信号通路关键蛋白Notch1、HEY1和Hes1的表达明显增加(P<0.05);而在T24细胞中过表达PRMT7后,呈相反的趋势。使用DAPT后,明显逆转了PRMT7在膀胱癌细胞中的表达。结论:PRMT7抑制膀胱癌细胞增殖和迁移,其作用机制与Notch信号转导通路有关。
Background and purpose:Bladder cancer is one of the common tumor of the urinary system.The protein arginine methyltransferase 7(PRMT7)has been reported in gastrointestinal tumors,but its biological role and mechanism in bladder cancer are still unknown.In this study,the effect of PRMT7 on the proliferation and migration of human bladder cancer cells and its possible mechanism were investigated.Methods:Human bladder cancer cell lines 5637,T24,RT112,UM-UC-3 and ureteral epithelial immortalized cells SV-HUC-1 were cultured in vitro,and the expression of PRMT7 was detected by Western blot.The expression of PRMT7 gene was silenced by RNA interference,and the negative control group(si-NC)and experimental group(siPRMT7#1 and siPRMT7#2)were established.PRMT7 gene were overexpressed by plasmid transfection,and the negative control group(p-PCMV3)and experimental group(p-PRMT7)were established.The transfection efficiency of PRMT7 was verified by real-time fluorescence quantitative polymerase chain reaction(RTFQ-PCR)and Western blot,cell proliferation was detected by cell counting kit-8(CCK8)assay and colony formation assay,and cell migration was detected by Transwell assay.Western blot was used to detect the expressions of proliferation and migration related target proteins including Cyclin D1,CDK4 and MMP9,as well as the key proteins Notch1,HEY1 and Hes1 in Notch signaling pathway.Notch signaling specific inhibitorsγ-secretase inhibitor DAPT was added to siPRMT7#2 transfected cells to further verify PRMT7 expression in bladder cancer.Results:Western blot results showed that PRMT7 expression was low in bladder cancer cells(P<0.05).After silencing PTMT7 in 5637 cells,cell proliferation and migration were significantly enhanced(P<0.05),and the expressions of proliferation and migration related target proteins including Cyclin D1,CDK4 and MMP9,and the key proteins of Notch signaling pathway Notch1,HEY1 and Hes1 were significantly increased(P<0.05).The overexpression of PRMT7 in T24 cells showed an opposite trend.DAPT signific
作者
王雪梅
程玉
齐洁敏
WANG Xuemei;CHENG Yu;QI Jiemin(Department of Pathology,Chengde Medical University,Chengde 067000,Hebei Province,China)
出处
《中国癌症杂志》
CAS
CSCD
北大核心
2023年第5期437-444,共8页
China Oncology
基金
承德医学院-国家自然科学基金项目培育基金(202114)
承德医学院-校级重点课题基金(201711)
河北省重点学科建设项目[冀教高(2013)4号病理学与病理生理学]
河北省硕士在读研究生创新能力培养资助项目(CXZZSS2022141)。
