靶向干扰Kv1.3通道基因表达对大鼠调节性T细胞的影响  

Effect of RNA interfering Kv1.3 channel expression on regulatory T lymphocytes in rats

作　　者：李梦佳 徐琦 程路峰[1] LI Meng-jia;XU Qi;CHENG Lu-feng(Dept of Pharmacology,College of Pharmacy,Xinjiang Medical University,Urumqi 830011,China;Dept of Immunology,Basic Medical College,Xinjiang Medical University,Urumqi 830011,China)

机构地区：[1]新疆医科大学药学院药理学教研室,新疆乌鲁木齐830011 [2]基础医学院免疫学教研室,新疆乌鲁木齐830011

出　　处：《中国药理学通报》2019年第7期945-949,共5页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No 81360491)

摘　　要：目的探究靶向 RNA干扰(RNAi)KCNA3(Kv1 3)基因后,体外给予依普利酮(eplerenone,EPL),分析 Tregs细胞活化增殖的变化。方法将慢病毒载体转染至大鼠调节性T细胞(Tregs)后,qPCR法和全细胞膜片钳法检测敲减效率。ELISA法检测 Tregs组、Tregs+EPL组、RNAi Tregs组、RNAi Tregs+EPL组细胞因子 IL 10、TGF β水平的变化。结果慢病毒成功转染 Tregs细胞,Kv1 3通道 mRNA水平和电流密度抑制率分别为 78%和 71 3%;与 Tregs组相比,RNAi Tregs组内外液 TGF β水平明显降低(P<0 01);Tregs+EPL组内外液 TGF β水平均降低(P<0 05);与 RNAi Tregs组相比,RNAi Tregs+EPL组内外液 TGF β水平无明显变化;而 Tregs组、Tregs+EPL组、RNAi Tregs组和 RNAi Tregs+EPL组内外液 IL 10水平变化不明显。结论 Kv1 3通道介导 Tregs的活化增殖,而 EPL可通过直接抑制 TregsKv1 3通道,减少 Tregs活化增殖,进而抑制 TGF β的分泌水平,说明依普利酮是 Kv1 3通道的特异性阻断剂。Aim To investigate the changes in activation and proliferation of Tregs after targeted RNA interference of Kv1.3 channel genes and in vitro administration of eplerenone (EPL).Methods After lentivirus vector was transfected to regulatory T cells (Tregs) of rats,qPCR and whole-cell patch-clamp methods were used to detect gene knockout efficiency,and ELISA method was used to detect cytokine secretion IL-10 and TGF-β levels of Tregs group,Tregs+EPL group,RNAi-Tregs group,and RNAi-Tregs+EPL group.Results Lentivirus vector was successfully transfected into Tregs cells,and the mRNA level and current density of Kv1.3 channel was 78% and 71.3% respectively;compared with Tregs group,extracellular and intracellular TGF-β levels in RNAi-Tregs group were significantly reduced ( P <0.01),and extracellular and intracellular TGF-β levels in Tregs+EPL group were also reduced( P <0.05);compared with RNAi- Tregs group,extracellular and intracellular TGF-β levels in RNAi- Tregs+EPL group showed no change.However,IL-10 levels in Tregs group,Tregs+EPL group,RNAi-Tregs group,RNAi-Tregs+EPL group showed no significant change.Conclusions Kv1.3 channel mediates the activation and proliferation of Tregs cells,while EPL can reduce the activation and proliferation of Tregs cells by directly inhibiting Kv1.3 channel and reducing the secretion of TGF-β levels,further indicating that EPL is a specific blocker of Kv1.3 channel.

关 键 词：调节性T细胞 依普利酮 Kv1.3通道 RNA干扰 活化增殖 转化生长因子Β 

分 类 号：R-332[医药卫生] R331.125