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CCR5 siRNA对类风湿关节炎大鼠滑膜细胞凋亡的影响
作者:王友强1  兰由玉2  李世勇1  郭永灿1 
单位:1. 西南医科大学附属中医医院 检验科, 四川 泸州 646000;
2. 西南医科大学附属医院 风湿免疫科, 四川 泸州 646000
关键词:类风湿关节炎 CC趋化因子受体5 小分子干扰RNA 滑膜细胞 细胞凋亡 
分类号:R593.22
出版年·卷·期(页码):2021·49·第三期(307-312)
摘要:

目的:利用小分子干扰RNA (siRNA)抑制CC趋化因子受体5(CCR5)的表达,以探讨CCR5 siRNA对类风湿关节炎(RA)大鼠滑膜细胞凋亡的影响及其可能的机制。方法:采用胶原诱导法构建RA大鼠模型,分离和培养RA大鼠滑膜细胞,并将细胞分为4组:RA组(RA大鼠滑膜细胞)、RA+NC组(加入脂质体包裹的NC siRNA)、RA+mock组(加入脂质体)、RA+siRNA组(加入脂质体包裹的CCR5 siRNA)。通过Lipofectamine 2000将CCR5 siRNA和NC siRNA载体转染入滑膜细胞;利用RNA干扰技术抑制大鼠滑膜细胞CCR5的表达后,再用qRT-PCR和Western Blot检测各组滑膜细胞CCR5、细胞凋亡相关基因[促凋亡Bcl-2相关X蛋白(Bax)、抗凋亡蛋白(Bcl-2)、Caspase-3(Cas3)]mRNA和蛋白表达情况;Annexin V-PI双染检测滑膜细胞的凋亡。结果:与RA组比较,RA+siRNA组CCR5、Bcl-2 mRNA和蛋白水平显著下降,Bax、Cas3 mRNA和蛋白水平明显上升,滑膜细胞凋亡率明显升高(均P<0.05),而RA+NC组、RA+mock组无明显变化。结论:CCR5 siRNA可抑制CCR5的表达,还可能通过上调Bax、Cas3水平及抑制Bcl-2的表达促进滑膜细胞的凋亡。

Objective: To explore the effect of C-C chemokine receptor 5(CCR5) small interfering RNA(siRNA) on the apoptosis of synovial cells in rats with rheumatoid arthritis (RA) and its possible mechanism by inhibiting the expression of CCR5 used siRNA. Methods: RA rat model was established by collagen-induced method, and then synovial cells were isolated and cultured.The synovial cells were divided into four groups:RA group (RA rat synovial cells), RA+NC group (add liposomal-coated NC siRNA), RA+mock group (add liposomes), and RA+siRNA group (add liposomal-coated CCR5 siRNA). CCR5 siRNA and NC siRNA vectors were transfected into synovial cells by Lipofectamine 2000.RNA interference was used to inhibit the expression of CCR5 in rat synovial cells,and then qRT-PCR and Western Blot were used to detect the mRNA and protein expression of CCR5 and apoptosis related genes (Bax, Bcl-2 and Cas3) in synovial cells of each group.Apoptosis of synovial cells was detected by Annexin V-PI double staining. Results: Compared with RA group, the mRNA and protein levels of CCR5 and Bcl-2 in the RA+siRNA group were significantly decreased, the mRNA and protein levels of Bax and Cas3 were significantly increased, and the apoptosis rate of synovium cells was significantly increased (P<0.05), while no significant changes were observed in the RA+NC group and RA+mock group.Conclusion: CCR5 siRNA can inhibit the expression of CCR5, and also promote the apoptosis of synovial cells by upregulating the levels of Bax and Cas3 and inhibiting the expression of Bcl-2.

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