NF-κB靶向性哑铃形“圈套”寡核苷酸对肾小球系膜细胞细胞因子表达的影响  

Inhibitory effect of circular dumbbell decoy ODNs targeting NF-кB on cytokines expression in mesangial cells

作　　者：张建国[1] 王惠民[1] 国华[2] 郝天智[2] 杨文军[2] 

机构地区：[1]第三军医大学附属大坪医院野战外科研究所肾内科,重庆400042 [2]第三军医大学附属大坪医院野战外科研究所第一研究室,重庆400042

出　　处：《西部医学》2004年第4期289-292,共4页Medical Journal of West China

基　　金：重庆市科委应用基础项目 (41 A1 1 537)

摘　　要：目的　设计合成靶向 NF-κB的哑铃形圈套 ODNs,并检测其对 NF-κB转录活性和肾小球系膜细胞细胞因子 (IL - 6和 TGF-β1)表达的抑制效应。方法　采用电泳迁移率改变实验 (EMSA)体外检测哑铃形圈套 ODNs对 NF-κB转录活性的抑制效应。提取大鼠肾小球系膜细胞 ,随机分为正常对照组、L PS刺激组、哑铃形圈套 ODNs处理组、无关圈套 ODNs处理组和脂质体处理组。采用阳离子脂质体将 2、4、8mg/ L 不同剂量哑铃形圈套 ODNs转染大鼠肾小球系膜细胞 ,6 h后用 L PS刺激 ,收集转染后 8、12、18上清和细胞。用酶联免疫吸附试验 (EL ISA)法检测上清细胞因子蛋白表达 ,逆转录 PCR(RT- PCR)法检测细胞因子 m RNA转录。结果　哑铃形圈套 ODNs在体外能有效地抑制 NF-κB与其顺式元件的结合 ;4、8mg/ L剂量哑铃形圈套 ODNs可明显抑制 L PS诱导的大鼠肾小球系膜细胞 IL - 6和 TGF-β1 转录与表达。结论　靶向 NF- κB的哑铃形圈套 ODNs在体外可抑制 NF- κB的转录活性 。Objective To investigate the inhibitory effects of circular dumbbell decoy oligodeoxynucleotides(ODNs) targeting NF-κB on the transcriptional activity of NF-κB binding its cis-elements and the expression of cytokines(IL-6 and TGF-β_1) in mesangial cells. Methods The inhibitory effect of circular dumbbell decoy ODNs on the transcriptional activity of NF-κB was detected by electrophoretic mobilitory shift assay (EMSA). Mesangial cells extracted from rats were randomly divided into normal control, LPS-stimulated, circular dumbbell decoy-ODNs treated, scrambled-decoy-ODNs treated and cationic liposome treated group. Circular dumbbell decoy ODNs at the doses of 2 mg/L, 4 mg/L, and 8 mg/L were transfected respectively into mesangial cells from rats using cationic liposome vector. Mesangial cell were stimulated with LPS at 6 h after circular dumbbell decoy-ODNs transfection. Then, the transfected supernatants and cells were collected at 8 h, 12 h. Cytokines mRNA expression in mesangial cells was detected by RT-PCR and cytokines protein level by ELISA. Results Circular dumbbell decoy ODNs could effectively inhibit the DNA binding activity of NF-кB to its cis-elements in vitro. Circular dumbbell decoy ODNs at the doses of 4 and 8 mg/L could effectively inhibit the expression levels of IL-6, TGF-β1 mRNA and protein in mesangial cells.Conclusion Circular dumbbell decoy ODNs targeting NF-κB could effectively inhibit the expression of cytokines in mesangial cells through the inhibition of NF-κB activity.

关 键 词：NF—κB 哑铃形圈套ODNs 细胞因子 

分 类 号：Q254[生物学—细胞生物学] Q952