摘要
目的：探讨Kr＆#252;ppel样因子2（KLF2）对氧化型低密度脂蛋白（oxidized low density lipo-protein， ox-LDL）诱导人脐静脉内皮细胞（HUVECs）微小RNA 146a（microRNA-146a， miR-146a）及细胞因子单核细胞趋化蛋白（MCP-1）和白细胞介素-6（IL-6）表达的影响。方法构建 KLF2过表达腺病毒载体（ rAAV-KLF2）及KLF2-siRNA，分别转染HUVECs及ox-LDL诱导的HUVECs。于0 h、3 h、6 h、12 h、24 h、48 h收集HUVECs，采用实时荧光定量PCR，检测HUVECs miR-146a的表达；设计合成锁核酸（the locked nucleic acid，LNA））修饰的anti-miR-146a的反义核苷酸（LNA-anti-miR-146a）转染HUVECs。收集各组细胞上清，采用双抗体夹心ABC-ELISA方法检测MCP-1及IL-6的表达。结果KLF2明显抑制静息及ox-LDL诱导的HUVECs miR-146a的表达，并显著减少ox-LDL诱导HUVECs MCP-1、IL-6的表达，且具有时间依赖性；miR-146 a 沉默可削弱ox-LDL诱导的内皮细胞炎症反应，而且部分逆转了KLF2对内皮细胞炎症反应的抑制作用。结论 KLF2通过下调miR-146 a的表达抑制ox-LDL活化的HUVECs MCP-1、IL-6的分泌。
Objective To investigate the effects of Kr ＆#252;ppel-like factor 2 （ KLF2 ） on oxidized low density lipoprotein （ox-LDL） induced expression of microRNA-146a （miR-146a） and proinflammatory cyto-kines （MCP-1 and IL-6） in human umbilical vein endothelial cells （HUVECs）.Methods Human umbili-cal vein endothelial cells （HUVECs） were cultured and then stimulated with 50μg/ml ox-LDL for 24 hours. HUVECs were infected with adenovirus vectors over-expressing human KLF2 at an appropriate multiplicity of infection.KLF2-siRNA duplexes were transfected into HUVECs to silence the gene expression .HUVECs were collected at time points of 0 h, 3 h, 6 h, 12 h, 24 h and 48 h after infection.Real-time quantitative-PCR was performed to measure the expression of miR-146a at mRNA level.Silenced endogenous miR-146a using LNA-anti-miR-146 a was transfected into HUVECs with lipofectamine 2000 .The levels of MCP-1 and IL-6 in supernatants were detected by ELISA .Results KLF2 remarkably inhibited the expression of miR-146 a in unstimulated and ox-LDL-stimulated HUVECs in a time-dependent manner .The ox-LDL induced ex-pression of miR-146a, MCP-1 and IL-6 in HUVECs were significantly decreased by KLF2.Silenced expres-sion of miR-146a downr-egulated the ox -LDL induced expression of MCP-1 and IL-6 in HUVECs.Moreover, silenced miR-146 a could partly reverse the inhibitory effects of KLF 2 on ox-LDL induced expression of MCP-1 and IL-6 in HUVECs.Conclusion KLF2 inhibited ox-LDL induced expression of MCP-1 and IL-6 in HUVECs partly through down-regulating the expression of miR-146a.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2014年第5期337-342,共6页
Chinese Journal of Microbiology and Immunology
基金
国家自然科学基金资助项目（81172781）
