安徽医科大学学报Issue(2)：129-134,135,7.
稳定敲低SIGIRR的人肾小管上皮细胞的构建及其初步功能的研究
Construction of SIGIRR stable knockdown human renal tubular epithelial cells and exploration of its function
摘要
Abstract
Objective To establish single immunoglobin IL-1-related receptor ( SIGIRR) stable knockdown human renal tubular epithelial cells ( HKC) and explore its function. Methods Designed the effective targeted shRNA se-quences for the SIGIRR gene, and then connected with the GV248-GFP-Puro to produce recombinant lentiviral vec-tor ( GV248-GFP-Puro-shSIGIRR ) . The corrected recombinants, identified by sequenced, were transfected into 293T cells with packaging plasmids (pMDL, pRev, pVSVG) for virus packaging. HKC cells were then infected by packaged viruses. Quantitative real-time PCR( qRT-PCR) and Western blot were used to detect the interference effi-ciency of SIGIRR in HKC cells. SIGIRR stable knockdown HKC cells ( HKC/shSIGIRR ) and the control cells were induced by IL-1β, then the downstream nuclear transcription factor, the phosphorylated NF-κB ( p65 ) were detected by Western blot. The mRNA expression levels of chemokine monocyte chemotactic protein 1 (MCP-1) and regulated upon activation normal T cell expressed and secreted factor ( RANTES) were analyzed by qRT-PCR.Results The results showed that the recombinant lentiviral vector GV248-GFP-Puro-shSIGIRR was successfully constructed. qRT-PCR and Western blot confirmed that SIGIRR was knockdown in HKC cells. Moreover, the re-sults also showed that compared with the control cells, the phosphorylated NF-κB (p65) and the mRNA levels of MCP-1 and RANTES were significantly increased in HKC/shSIGIRR cells by being stimulated with IL-1β. Conclu-sion The results suggest that SIGIRR acts as a“brake” of inflammatory reaction in Toll-like receptor/IL-1 recep-tor (TLR/IL-1R) pathway in HKC cells. The study may provide a potential new target for the treatment of Lupus nephritis.关键词
狼疮性肾炎/人肾小管上皮细胞/SIGIRR/慢病毒载体/NF-κB/趋化因子Key words
lupus nephritis/human renal tubular epithelial cells/SIGRR/lentiviral vector/NF-κB/chemokine分类
医药卫生引用本文复制引用
蒋克国,郝丽,王德光,周海胜,查晓军,张桂霞,金福泉,季爽,潘海峰,叶冬青..稳定敲低SIGIRR的人肾小管上皮细胞的构建及其初步功能的研究[J].安徽医科大学学报,2015,(2):129-134,135,7.基金项目
安徽省博士后科学基金(编号:9101019202) （编号:9101019202）
中国博士后科学基金(编号:2012M511399) （编号:2012M511399）
国家自然科学基金(编号:81101524、81372475、81172591) （编号:81101524、81372475、81172591）
