草酸钙结石晶体对人近曲肾小管上皮细胞蛋白质表达谱的影响  

Effect of calcium oxalate stone on protein expression profiles of human proximal renal tubular epithelial cells

作　　者：张建文 徐长志[2] 张颖 孙睿 植凡 胡七一 张圣平 梁辉 王铸 Zhang Jianwen;Xu Changzhi;Zhang Ying;Sun Rui;Zhi fan;Hu Qiyi;Zhang Shengping;Liang Hui;Wang Zhu(Department of Urology,People's Hospital of Longhua,Shenzhen 518109,China;Departmentof Clinical Laboratory,Lingnan Hospital,The Third Affiliated Hospital,Sun Yat-Sen University,Guangzhou 510530,China)

机构地区：[1]深圳市龙华区人民医院泌尿外科,518109 [2]中山大学附属第三医院岭南医院检验科,广州510530

出　　处：《中华腔镜泌尿外科杂志（电子版）》2019年第4期225-229,共5页Chinese Journal of Endourology(Electronic Edition)

基　　金：深圳市科技计划项目(JCYJ20170307141840188);深圳市龙华区科技创新资金项目(2017009)

摘　　要：目的探讨肾结石草酸钙晶体对人近曲肾小管上皮细胞蛋白质表达谱的影响,筛选差异表达蛋白质并初步探讨肾结石与肾损伤的关系。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,处理人近曲肾小管上皮细胞HK-2,提取总蛋白并通过SDS-PAGE对蛋白质量进行鉴定,通过BCA法测定蛋白浓度,然后通过TMT标记定量蛋白质组学分析法筛选实验组和对照组蛋白质表达谱的差异,最后通过生物信息学的方法进行聚类分析、GO富集分析以及差异基因相关的蛋白质相互作用网络分析。结果草酸钙晶体对HK-2细胞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。TMT标记定量蛋白质组学分析结果显示经草酸钙晶体处理后的HK-2细胞中差异表达的蛋白分子共1 141个,其中上调表达的699个,下调表达的442个。生物信息学分析结果提示这些差异蛋白在细胞外复合体、细胞器以及多种细胞学过程中发挥重要作用;差异蛋白共表达网络分析发现在细胞骨架维持(ACTB,CFL1)和细胞特殊功能如分泌和加帽中起作用的蛋白(MYH9,MYH14)关系最为密切。结论草酸钙结晶对人肾上皮细胞蛋白质表达有显著影响,并且这些差异表达的蛋白质分子可能与肾结石的形成以及肾损伤有关。Objective To investigate the effect of calcium oxalate crystals on the protein expression profiles in human proximal renal tubular epithelial cells,screen differentially expressed proteins and explore the relationship between kidney stones and renal injury.Methods The clinical kidney stone samples were analyzed and chose to culture with human proximal tubular epithelial cells HK-2.The cell total protein was extracted and qualified by SDS-PAGE.The protein concentration was determined by BSA method,and the difference of protein expression profiles between the experimental group and the control group were screened by TMT-labeled quantitative proteomics analysis.Cluster analysis,GO enrichment analysis and differential gene-related interaction network analysis were performed by bioinformatics method.Results The calcium oxalate crystal showed obvious cytotoxicity to HK-2 cells,and the cell growth slowed down and caused cell shape change significantly.The TMT-labeled quantitative proteomic analysis showed that after treatment with calcium oxalate crystals,a total of 1 141 proteins were differentially expressed in HK-2 cells,of which 699 were up-regulated and 442 were down-regulated.Bioinformatics analysis indicated that these differential proteins played important roles in extracellular complexes,organelles and various cytological processes,differential protein co-expression network analysis demonstrated that proteins involved in cytoskeletal maintenance(such as ACTB,CFL1)and several specific functions such as secretion and capping(MYH9,MYH14)were most closely related.Conclusion Calcium oxalate crystal has a significant effect on protein expression profiling in human renal epithelial cells,and these differentially expressed protein molecules may be involved in the formation of kidney stones and kidney injury.

关 键 词：草酸钙结晶 肾结石 蛋白质表达谱 肾上皮细胞 聚类分析 

分 类 号：R692.4[医药卫生—泌尿科学]