摘要
目的探讨长链非编码RNA SCARNA10对肝癌细胞增殖的影响及其机制。方法收集人肝癌细胞HepG2、MH97H及人正常肝细胞L-7702,采用实时荧光定量PCR(qPCR)法检测细胞中的SCARNA10。将HepG2和MH97H细胞各分为两部分,分别转染小干扰RNA(siRNA)和阴性对照RNA(NC)36 h;收集细胞,采用qPCR法检测细胞中的增殖细胞核抗原(PCNA)mRNA,CCK-8法检测波长450 nm处的光密度(OD)值表示细胞增殖活性,克隆形成实验计数克隆形成数。核质分离提取实验观察SCARNA10在肝癌细胞胞核、胞质中的分布情况,RNA结合蛋白免疫沉淀(RIP)实验验证SCARNA10与SUZ12、EZH2的相互作用。结果HepG2、MH97H细胞中SCARNA10相对表达量均高于L-7702细胞(P均<0.05)。与转染NC比较,转染siRNA的HepG2、MH97H细胞中PCNA mRNA表达量、细胞增殖OD值均降低,细胞克隆数均减少(P均<0.05)。核质分离提取实验显示,SCARNA10在HepG2、MH97H细胞胞核内的表达比例高于胞质内表达比例(P均<0.05)。HepG2细胞中SCARNA10与SUZ12、EZH2的相对富集量分别为5.05±1.39、3.95±0.67,MH97H细胞中分别为6.71±2.60、4.47±1.75。结论在细胞核中,SCARNA10可能通过与SUZ12、EZH2结合从而促进肝癌细胞增殖。
Objective To investigates the effect and mechanism of long non-coding RNA SCARNA10 on the proliferation of hepatic carcinoma cells.Methods We collected the human hepatic carcinoma cell lines HepG2,MH97H,and normal hepatocytes L-7702.The expression of SCARNA10 in the above cells was detected by real-time quantitative PCR(qPCR).HepG2 and MH97H were transfected with small interfering RNA(siRNA)and negative control(NC),respectively;then the cells were collected,qPCR was used to test the mRNA of proliferating cell nuclear antigen(PCNA),CCK-8 kit was used to detect the optical density(OD)value at 450 nm wavelength to indicate the cell proliferation activity,and the clone formation assay was used to count the number of clone formation.The distribution of SCARNA10 in hepatic carcinoma cells was observed by subcellular fractionation assay.RNA immunoprecipitation(RIP)was used to verify the interaction between SCARNA10 and suppressor of zeste12(SUZ12)and enhancer of zeste homolog2(EZH2).Results The relative level of SCARNA10 in HepG2 and MH97H cells was higher than that of L-7702 cells(both P<0.05).Compared with the negative control(NC),the relative level of PCNA,the OD value,and the number of clone formation decreased in HepG2 and MH97H cells transfected with siRNA(all P<0.05).The subcellular fractionation assay indicated that the relative expression of SCARNA10 in the nucleus of HepG2 and MH97H cells was higher than that of cytoplasm(both P<0.05).The relative enrichment of SCARNA10 with SUZ12 and EZH2 was 5.05±1.39 and 3.95±0.67,respectively,in the HepG2 cells,versus 6.71±2.60 and 4.47±1.75,respectively,in the MH97H cells.Conclusion SCARNA10 may promote the proliferation of hepatic carcinoma cells by binding to SUZ12 and EZH2 in the nucleus.
作者
韩亚伟
崔冉亮
李悦国
HAN Yawei;CUI Ranliang;LI Yueguo(Tianjin Medical University Cancer Institute and Hospital,National Clinical Research Center for Cancer,Tianjin Key Laboratory of Cancer Prevention and Therapy,Tianjin’s Clinical Research Center for Cancer,Tianjin 300060,China)
出处
《山东医药》
CAS
2020年第29期10-13,共4页
Shandong Medical Journal
基金
国家自然科学基金面上项目(81871719)
天津医科大学肿瘤医院引进人才与博士启动基金(B1916)。
