α-胞衬蛋白siRNA对人涎腺导管细胞的影响  

Effect of α-Fodrin siRNA on human salivary gland cells

作　　者：庞春艳[1] 王蓓[1] 剧锦哲[1] 王永福[1] 

机构地区：[1]内蒙古科技大学包头医学院风湿免疫研究所/包头医学院第一附属医院风湿免疫科,内蒙古包头014010

出　　处：《重庆医学》2016年第7期880-882,887,共4页Chongqing medicine

基　　金：包头市医药卫生基金项目(2009S1001-34);内蒙古自治区科技计划项目(20120403)

摘　　要：目的观察α-胞衬(α-Fodrin)蛋白小干扰RNA(siRNA)对人涎腺导管(HSG)细胞的影响,探讨α-Fodrin siRNA治疗干燥综合征(SS)的作用。方法实验分对照组、空载体组、α-Fodrin siRNA1组及α-Fodrin siRNA2组,将成功构建的10μgα-Fodrin siRNA1和α-Fodrin siRNA2表达载体分别采用脂质体法转染HSG细胞,空载体组HSG细胞转染等剂量的pGFP-V-RS空载体。转染后24、48、72和96h观察细胞转染效率,实时荧光定量(RT)-PCR法检测4组HSG细胞中α-Fodrin mRNA的表达水平;细胞荧光免疫法检测4组细胞中α-Fodrin蛋白的表达水平;ELISA法检测4组细胞上清液中IFN-γ、IL-10的表达水平。结果转染后48hHSG细胞转染效率最高;转染后48hα-Fodrin siRNA1组和α-Fodrin siRNA2组HSG细胞中α-Fodrin mRNA和蛋白的表达水平明显低于对照组和空载体组(P<0.05),且α-Fodrin siRNA2组中α-Fodrin mRNA的表达水平低于α-Fodrin siRNA1组的表达水平(P<0.05);α-Fodrin siRNA1组和α-Fodrin siRNA2组细胞上清液中IL-10的水平明显升高,较对照组和空载体组差异有统计学意义(P<0.05),α-Fodrin siRNA1组和α-Fodrin siRNA2组比较差异无统计学意义(P>0.05);α-Fodrin siRNA1组和α-Fodrin siRNA2组细胞上清中IFN-γ的水平低于对照组和空载体组,但4组之间比较差异无统计学意义(P>0.05)。结论 HSG细胞转染α-Fodrin siRNA载体后,α-Fodrin mRNA和蛋白的表达水平下降,同时细胞上清中IL-10的水平升高、IFN-γ水平下降,提示α-Fodrin siRNA可以减轻炎性反应,为SS的治疗提供实验依据。Objective To observe the effect ofα-Fodrin siRNA on human salivary gland（HSG）cells and to discuss its therapy on sjgren＇s syndrome（SS）.Methods The vectors expressing siRNA againstα-Fodrin of human were transfected into HSG cells of 10μgα-fodrin siRNA1 group andα-Fodrin siRNA2 group,while pGFP-V-RS vector were transfected into the cells of empty vector group,there was no handling in HSG cell of control group.The efficiency was observed by fluorescence microscope after transfection of 24,48,72 and 96hby lipofectamine 2000.The expression levels ofα-Fodrin mRNA and protein of HSG were detected by real-time（RT）-PCR and immunohistochemistry method respectively.The expression levels of IFN-γand IL-10 in supernatant of cells were detected by ELISA.Results The efficiency was highest on 48 hafter transfection.The level ofα-Fodrin mRNA and protein was lower inα-fodrin siRNA1 group andα-fodrin siRNA2 group than control group and empty vector group on 48 hafter transfection（P〈0.05）,the level ofα-Fodrin mRNA inα-fodrin siRNA2 group was significant higher thanα-fodrin siRNA1group（P〈0.05）.The levels of IL-10 were higher inα-fodrin siRNA1 group andα-fodrin siRNA2 group than control group and empty vector group on 48 hafter transfection（P〈0.05）,α-fodrin siRNA1 group andα-fodrin siRNA2 group had no statistical significance（P〉0.05）.The levels IFN-γinα-fodrin siRNA1 group andα-fodrin siRNA2 group were lower than of control group and empty vector group on 48 hafter transfection,but there were no significant differences in the four groups（P〈0.05）.ConclusionTheα-fodrin siRNA1andα-fodrin siRNA2 can suppress the levels ofα-fodrin mRNA and protein of HSG cells,at the same time;they can elevate the expression of IL-10 and decrease the level of IFN-γ.Soα-Fodrin siRNA reduce the levels of inflammatory cytokines and provide experimental basis to therapy of SS.

关 键 词：RNA 小分子干扰 Α-胞衬蛋白 人涎腺导管细胞 白细胞介素-10 干扰素-Γ 

分 类 号：R593.2[医药卫生—内科学]