草酸钙结石晶体粘附对人肾上皮细胞lncRNA表达谱的影响  

Long non-coding RNA expression profile in calcium oxalate monohydrate attached HK-2 cells

作　　者：王铸 张建文 张颖 张圣平 孙睿 植凡 胡七一 邓琼 梁辉 

机构地区：[1]深圳市龙华区人民医院泌尿外科,广东深圳518109

出　　处：《现代泌尿外科杂志》2018年第1期52-56,76,共6页Journal of Modern Urology

基　　金：深圳市知识创新计划项目(No.JCYJ20150402144905865);深圳市龙华区科技创新资金项目(No.20150925A0410013)

摘　　要：目的比较分析肾结石草酸钙晶体对人肾上皮细胞lncRNA的表达谱的影响,筛选差异表达的lncRNA并初步探讨其与肾结石的关系。方法选取临床经结石成分分析为草酸钙的肾结石标本,充分研磨后配制草酸钙晶体悬液处理人肾上皮细胞HK-2细胞,通过MTT法测定草酸钙晶体对HK-2细胞生长的影响;提取总RNA并通过高通量转录组测序平台HiSeq 2500进行lncRNA表达谱差异性分析。对筛选获得的差异lncRNA进行Real-Time PCR验证,并通过Gene Ontology数据库和KEGG信号通路初步分析其功能。结果草酸钙晶体在数分钟内便粘附于HK-2细胞表面,并且显示出明显的细胞毒性,细胞生长速度减慢且引起细胞形态以及培养基pH值发生显著改变。RNA seq结果显示经草酸钙晶体处理后的HK-2细胞中差异表达的lncRNA有25个,其中上调表达的9个,下调表达的16个。通过实时定量RT-PCR进行验证,差异均具有统计学意义(P<0.05)。结论草酸钙结晶对人肾上皮细胞lncRNA以及mRNA表达有显著影响,并且这些lncRNA的异常表达可能与肾结石发病有关。ObjectiveTo explore the expression profiles of Long noncoding RNA （lncRNA） in calcium oxalate monohydrate attached HK2 cells,and to investigate the association between critical lncRNA expression and renal injury.MethodsThe HK2 cells were treated with crystal suspension of calcium oxalate which was grinded form clinical kidney sample and characterized by stone composition analysis.The effects of calcium oxalate crystals on the growth of HK2 cells were determined by MTT assay.Total RNA was extracted and the lncRNA expression profile was analyzed by highthroughput transcriptase sequencing platform HiSeq 2500.The profile of identified lncRNA was verified by realtime PCR and their function was analyzed by Gene Ontology database and KEGG signal pathway analysis.ResultsCalcium oxalate crystals adhered to the surface of HK2 cells in a few minutes and showed obvious cytotoxicity.RNA seq results showed that there were 25 differentially expressed lncRNAs in HK2 cells treated with calcium oxalate crystals,of which 9 were upregulated and 16 were downregulated.The difference verified by realtime PCR were statistically significant （P〈0.05）.ConclusionCalcium oxalate crystals have a significant effect on lncRNA expression in human renal epithelial cells,which may play critical roles in kidney stonemediated renal injury.

关 键 词：草酸钙结晶 肾结石 IncRNA RNA seq 

分 类 号：R692[医药卫生—泌尿科学]