作 者：
谢亮亮;黄时海;汤龙;韦英明;石德顺;李湘萍
单 位：
广西大学亚热带农业生物资源保护与利用国家重点实验室;广西大学生命科学与技术学院
关键词：
脂多糖结合蛋白;RNAi;水牛;单核巨噬细胞;LPS;基因表达
摘 要：
为了探讨抑制脂多糖结合蛋白(LBP)基因表达对水牛单核巨噬细胞在内毒素(LPS)诱导下炎症相关基因的表达,首先采用三质粒慢病毒包装系统包装LBP基因shRNA重组质粒pSicoR-GFP-shLBP774.利用病毒颗粒感染水牛单核巨噬细胞,进行荧光定量qRT-PCR及ELISA检测.结果显示,单核巨噬细胞在用5×108 IU/mL滴度的LBP-shRNA774慢病毒颗粒,感染指数(MOI)为300、感染7d的条件下,感染效率超过50%.qRT-PCR检测结果显示,与LPS对照组相比,shLBP774感染组可极显著抑制LBP基因的表达(p<0.01),CD14,TLR4,TNF-α,IL-1β,IL-8等基因表达显著降低(p<0.05).ELISA检测结果发现,与对照组相比,细胞分泌的TNF-α,IL-1β蛋白量显著降低(p<0.05).以上结果表明,抑制LBP基因表达可以有效降低LPS诱导下单核巨噬细胞炎症相关基因的表达,进而调控LPS引发的炎症反应,提示可将LBP作为靶基因深入研究水牛单核巨噬细胞免疫功能和LPS致炎信号传导分子机制.
译 名：
Effects of LBP Gene Silencing on Gene Expression in Water Buffalo Monocyte-Macrophages Post-LPS Stimulation
作 者：
XIE Liang-liang;HUANG Shi-hai;TANG Long;WEI Ying-ming;SHI De-shun;LI Xiang-ping;State Key Laboratory of Subtropical Bioresource Conservation and Utilization at Guangxi University;College of Life Science and Technology,Guangxi University;
单 位：
XIE Liang-liang%HUANG Shi-hai%TANG Long%WEI Ying-ming%SHI De-shun%LI Xiang-ping%State Key Laboratory of Subtropical Bioresource Conservation and Utilization at Guangxi University%College of Life Science and Technology,Guangxi University
关键词：
lipopolysaccharide binding protein(LBP);;RNAi;;water buffalo(Bubalus bubalis);;monocyte-macrophages;;lipopolysaccharides(LPS);;gene expression
摘 要：
In order to investigate the effect of inhibition of LBP(lipopolysaccharide binding protein)gene expression on LPS-induced expression of macrophage inflammatory genes in buffalo mononuclear macrophages,LBP-shRNA 774 recombinant plasmid was used to package the virus,and the suitable conditions for target cell infection were explored.Then inflammation-related genes in virus-infected cells were detected with the qRT-PCR and ELISA methods.The results showed that the virus titer of LBP-shRNA774 was>5×108 IU/mL,MOI was 300,and after 7 days,the infection rate was more than 50%.qRT-PCR showed that compared with the LPS control group,the mRNA level of LBP gene was reduced by 50%,meanwhile CD14,TLR4,TNF-α,IL-1βand IL-8 were decreased significantly(p<0.05).ELISA demonstrated that the contents of cytokine TNF-αand of IL-1βin the LBP-shRNA774 group were significantly lower than in the LPS control group(p<0.05).Also,the content of soluble LBP(s LBP)protein was significantly lower(p<0.05),only accounting for 48.54%.The above results suggested that inhibition of LBP gene expression can effectively reduce the expression of LPS-induced inflammation-related genes of monocyte-macrophage,and regulate the LPS-induced inflammatory response.
