林卡帅?邱月?董兰?周姗姗?黄俊?秦曙光?何凤

【摘要】目的 探討长链非编码 RNA（lncRNA）在糖尿病肾病（DN）发生发展中的作用。方法 收集临床肾活组织检查标本，采用RNA-seq 测序技术检测DN组与正常对照组（NC组）肾组织中差异表达的lncRNA及mRNA。通过 GO、KEGG数据库分析差异表达mRNA的生物学功能，并通过共表达网络分析预测差异表达lncRNA的相互作用基因。采用实时荧光定量 PCR（qRT-PCR）检测目标lncRNA、mRNA 在DN肾组织中的相对表达水平。结果 RNA-seq 测序结果表明，与NC组相比，DN组共有353个差异表达的lncRNA，其中224个表达上调，129个表达下调。qRT-PCR 结果显示，DN组中CRNDE、PVT1和 BLZF2P 相对表达水平较NC组升高（P均< 0.001），而WT1-AS、TARID 和 ST13P6相对表达水平较NC组降低（P均< 0.001）。共表达网络分析及双变量相关分析显示lncRNA CRNDE 与 NPHS1（编码的 nephrin 是足细胞结构完整性和发挥功能的决定性关键蛋白）呈负相关。结论 lncRNA CRNDE 在DN肾组织中表达明显升高，且与 NPHS1存在负相关关系，lncRNA CRNDE 可能通过调控 NPHS1 的表达促进DN足细胞损伤。

【关键词】糖尿病肾病;长链非编码核糖核酸;CRNDE;NPHS1;足细胞

Long non-coding RNA CRNDE promotes podocyte indury in diabetic nephropathy by regulating NPHS1 expression Lin Kashuai， Qiu Yue， Dong Lan， Zhou Shanshan， Huang Jun， Qin Shuguang， He

Feng. School of Medicine， South China University of Technology， Guangzhou 510006， China

Corresponding author， He Feng， E-mail： eyhefeng@ scut. edu. cn

【Abstract】Objective To investigate the role of long non-coding RNAs （lncRNAs） in the incidence and progression of diabetic nephropathy （DN）. Methods The differentially-expressed lncRNAs and mRNAs in the kidney biopsy specimens were detected by RNA-seq sequencing between the DN and normal control groups （NC）. The biological functions of differentially-expressed mRNAs were analyzed through the GO and KEGG databases， and the interaction among differentially-expressed lncRNAs and mRNAs was analyzed through co-expression network analysis. The relative expression levels of target lncRNAs and mRNAs in DN kidney tissues were detected by real-time fluorescent quantitative PCR （qRT-PCR）. Results RNA-seq sequencing results showed that a total of 353 differentially-expressed lncRNAs in the DN group compared with that in the NC group， in which 224 were up-regulated and 129 were down-regulated. qRT-PCR revealed that the relative expression levels of CRNDE， PVT1 and BLZF2P in the DN group were significantly higher （all P < 0.001）， whereas the relative expression levels of WT1-AS， TARID and ST13P6 were remarkably lower than those in the NC group （all P < 0.001）. Moreover， co-expression network analysis and bivariate correlation analysis demonstrated that lncRNA CRNDE was negatively correlated with NPHS1 （the encoded nephrin is the decisive key protein for the structural integrity and function of podocytes）. Conclusions The expression level of lncRNA CRNDE is significantly up-regulated in DN patients， which is negatively correlated with NPHS1， indicating that lncRNA CRNDE may promote the podocyte damage in DN by regulating the expression level of NPHS1.

【Key words】Diabetic nephropathy;Long non-coding RNA;CRNDE;NPHS1;Podocyte

糖尿病肾病（DN）是糖尿病全身性微血管病变常见的并发症，是我国终末期肾病（ESRD）的主要病因，并且成为目前慢性疾病防治的重要问题之一[1-2]。越来越多的研究显示，在DN早期，当系膜细胞和内皮细胞尚未改变时就已出现足细胞损伤，表现为足细胞数量减少，胞体缩小，足突增宽融合、消失，进而引起蛋白尿导致肾功能恶化进展[3-4]。因此，足细胞损伤是DN早期重要的病理基础，然而其参与DN发生发展的机制尚未完全阐明。

随着测序技术的发展及精准医学的提出，以 Illumina高通量测序为代表的二代测序技术成为当下检测已知、预测未知RNA的首选技术，为在分子水平寻找生物标志物提供了极大的便利。长链非编码 RNA（lncRNA）是一类数量庞大、转录本长度大于 200个核苷酸且不具有编码蛋白质功能的RNA。近年多项研究显示，lncRNA的异常表达与恶性肿瘤、心血管疾病、神经疾病及代谢性疾病等多种疾病相关[5-7]。……