人脐带间充质干细胞向肝细胞分化过程中长链非编码RNA核富集转录本1的作用  

Role of long non-coding RNA nuclear-enriched abundant transcript 1 in the differentiation of human umbilical cord mesenchymal stem cells into hepatocytes

作　　者：王丹丹[1] 禹亚彬[2] 刘世奇 严雨楼 张建淮[2] Wang Dandan;Yu Yabin;Liu Shiqi;Yan Yulou;Zhang Jianhuai(Nanjing Medical University,Nanjing 210000,Jiangsu Province,China;Department of Hepatobiliary Surgery,Affiliated Huaian No.1 People’s Hospital of Nanjing Medical University,Huaian 223300,Jiangsu Province,China)

机构地区：[1]南京医科大学,江苏省南京市210000 [2]南京医科大学附属淮安第一医院肝胆外科,江苏省淮安市223300

出　　处：《中国组织工程研究》2022年第30期4847-4851,共5页Chinese Journal of Tissue Engineering Research

基　　金：淮安市自然科学项目(HAB202016),项目负责人:禹亚彬;南京医科大学附属淮安第一医院高层次人才科研项目(YGRX201902),项目负责人:禹亚彬。

摘　　要：背景:人脐带间充质干细胞在体外借助细胞因子可以诱导分化为肝样细胞,但是形成的肝样细胞功能上尚不够成熟。研究发现长链非编码RNA与细胞分化关系密切,并通过干预某些长链非编码RNA的表达达到了促进细胞分化的目的。目的:探讨长链非编码RNA核富集转录本1对人脐带间充质干细胞向肝细胞分化的影响。方法:建立人脐带间充质干细胞向肝细胞分化体系,用糖原染色和吲哚菁绿摄取实验评价肝细胞功能特性,qRT-PCR检测肝细胞特异性基因及核富集转录本1的mRNA表达。然后通过构建慢病毒干扰核富集转录本1表达,转染7 d后得到稳定转染的细胞株,然后进行肝向诱导分化4周,设立转染空质粒的阴性对照组,未进行病毒转染的对照组,qRT-PCR、Western blot检测肝细胞特异性基因和蛋白表达。结果与结论:①经糖原染色后可见细胞胞质内有红、紫色糖原颗粒,吲哚菁绿摄取后可见大量深绿色吲哚菁绿阳性细胞;②随着诱导时间延长,白蛋白、细胞色素酶P4503A4 mRNA表达量逐渐增加,核富集转录本1的mRNA表达量逐渐降低;③与阴性对照组和未转染对照组比较,在干扰核富集转录本1表达后,白蛋白、细胞色素酶P4503A4 mRNA和蛋白表达均明显增加;④结果表明,长链非编码RNA核富集转录本1可能参与了人脐带间充质干细胞向肝细胞的分化。BACKGROUND:Human umbilical cord mesenchymal stem cells can be induced to differentiate into hepatocytes with the help of cytokines in vitro,but the resulting hepatoid cells are not yet functionally mature.It has been gradually found that long non-coding RNA(lncRNA)is closely related to cell differentiation and the purpose of promoting cell differentiation is achieved by interfering with the expression of some lncRNAs OBJECTIVE:To explore the role of long non-coding RNA nuclear-enriched abundant transcript 1(lncRNA NEAT1)in the differentiation of human umbilical cord mesenchymal stem cells into hepatocytes.METHODS:A differentiational cell line system of human umbilical cord mesenchymal stem cells into hepatocytes was established.Periodic Acid-Schiff staining and indocyanine green intake test were used to assess hepatocellular functional characteristics of the cells.Quantitative real-time reverse transcription polymerase chain reaction was used to detect the expression of hepatocyte specific genes and lncRNA NEAT1.The lentivirus was constructed to interfere with NEAT1 expression,and stably transfected cell lines were obtained 7 days after transfection,and hepatotropic differentiation was induced for 4 weeks.A negative control group for transfection with empty plasmids and a control group without virus transfection were set up.The expression levels of hepatocyte specific genes and proteins were detected by quantitative real-time reverse transcription polymerase chain reaction and western blot assay.RESULTS AND CONCLUSION:(1)After Periodic Acid-Schiff staining,there were red and purple glycogen particles in the cytoplasm of cells,and a large number of indocyanine green positive cells were observed after indocyanine green ingestion.(2)With the extension of induction time,the mRNA expression levels of albumin and cytochrome enzyme P4503A4 mRNA increased gradually,while the mRNA expression levels of NEAT1 decreased gradually.(3)Compared with the negative control group and the control group,the mRNA and protein expressio

关 键 词：长链非编码RNA NEAT1 间充质干细胞 肝细胞 分化 肝细胞特异性基因 转染 

分 类 号：R459.9[医药卫生—治疗学] R329[医药卫生—临床医学]