长链非编码RNAs调控骨桥蛋白在草酸钠结晶肾损伤的机制研究

发布时间：2018-03-22 23:17

本文选题：长链非编码RNAs　切入点：骨桥蛋白　出处：《第二军医大学》2017年硕士论文　论文类型：学位论文

【摘要】：【目的】运用生物信息学方法检测与骨桥蛋白相关的Lnc RNAs在草酸钠诱导结晶肾损伤中的差异变化,初步探讨骨桥蛋白相关Lnc RNAs在结晶肾损伤中的作用机制。【方法】利用草酸钠刺激肾小管上皮细胞(HK-2)构建结晶肾损伤模型。采用实时定量PCR(RT-PCR)技术,在通过草酸钠刺激肾小管上皮细胞(HK-2)构建的细胞模型中验证骨桥蛋白表达水平。运用RNA干扰技术构建骨桥蛋白敲低细胞模型。选用Arraystar公司提供的人类全基因组lnc RNA芯片(V4.0)对3组骨桥蛋白干扰组和3组对照组肾小管上皮细胞结晶模型进行骨桥蛋白(OPN)相关lnc RNAs以及m RNAs差异基因表达水平检测。其中lnc RNAs来源于权威公共转录组数据库(包括Refseq、UCSC knowngenes、Gencode等)。差异基因筛选条件是差异倍数1.5,P0.05。对差异基因进行基因本体论(GO)及信号通路分析(KEGG Pathway)等生物信息学分析。【结果】1.通过草酸钠刺激肾小管上皮细胞成功构建结晶肾损伤细胞模型以及通过RNA干扰技术构建骨桥蛋白干扰模型。2.全基因组lnc RNA芯片提示在肾小管上皮细胞结晶肾损伤模型中存在骨桥蛋白相关差异表达的lnc RNAs,共有583个lnc RNAs存在差异表达,其中骨桥蛋白干扰组表达上调354个,下调229个;共有235个m RNAs存在差异表达,其中干扰组上调139个,下调96个。3.通过基因本体论(GO)及信号通路分析(KEGG Pathway)等生物信息学分析表明,差异表达的lnc RNAs与细胞生长、酶活性调节、PI3k/Akt/NF-κB信号通路、Wnt信号通路等相关联。KEGG信号通路分析结果提示,上调lnc RNAs与14条通路相关,而下调lnc RNAs与5条通路相关。综合分析提示NOD样受体信号通路、PI3k/Akt/NF-κB信号通路、Wnt信号通路等炎症相关通路显著富集。【结论】体外建立人肾小管上皮细胞结晶肾损伤模型中存在骨桥蛋白相关差异Lnc RNAs表达。骨桥蛋白相关lnc RNAs参与草酸盐结晶肾损伤发病机制,未来可作为候选生物标记物为临床上结晶所致肾损伤提供理论及实验依据。
[Abstract]:[objective] to detect the differential changes of Lnc RNAs associated with osteopontin in sodium oxalate induced renal injury by bioinformatics. To explore the mechanism of osteopontin associated Lnc RNAs in renal injury. [methods] the model of renal injury was established by sodium oxalate stimulation of renal tubular epithelial cells (HK-2). The expression level of osteopontin was verified in a cell model constructed by sodium oxalate stimulation of renal tubular epithelial cells (HK-2). Osteopontin knockdown cell model was constructed by using RNA interference technique. The whole human genome lnc provided by Arraystar was selected. RNA microarray V4.0) was used to detect the differential gene expression levels of lnc RNAs and m RNAs related to osteopontin interference group and renal tubular epithelial cell crystallization model of 3 groups and 3 control groups. Lnc RNAs originated from authoritative common transcription. Group database (including RefseqUCSC knowngenesl Gencode et al. The screening condition of differential gene is 1.5mv P0.05.The differential gene is analyzed by gene ontology and signal pathway analysis KEGG Pathway. [results] 1. Stimulating renal tubules by sodium oxalate. The successful construction of crystalline renal injury cell model by skin cells and the construction of osteopontin interference model by RNA interference technology. 2. The whole genome lnc RNA chip indicates that there is osteopontin correlation in renal tubular epithelial cell crystal kidney injury model. Among the differentially expressed lnc RNASs, 583 lnc RNAs were differentially expressed. There were 354 up-regulated and 229 down-regulated osteopontin interference groups, and 235 m RNAs were differentially expressed, in which 139 m RNAs were up-regulated and 96 路3.The results of bioinformatics analysis, such as gene ontology and signal pathway analysis, showed that, The differential expression of lnc RNAs was associated with cell growth, enzyme activity regulation, PI3k / NF- 魏 B signaling pathway and Wnt signaling pathway. The results showed that the up-regulation of lnc RNAs was associated with 14 pathways. However, down-regulation of lnc RNAs was associated with 5 pathways. Comprehensive analysis showed that the PI3k / Akt / NF- 魏 B signaling pathway and Wnt signaling pathway were significantly enriched. [conclusion] the model of renal injury induced by crystallization of human renal tubular epithelial cells was established in vitro. Osteopontin related lnc RNAs was involved in the pathogenesis of oxalate crystal kidney injury. It can be used as a candidate biomarker to provide theoretical and experimental evidence for renal injury induced by clinical crystallization.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R692

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