蛋白酶体抑制剂MG132对大鼠胶原诱导性关节炎的干预机制

Intervention effect of proteasome inhibitor MG132 in rats with collagen-induced arthritis

目的:探讨蛋白酶体抑制剂MG132对大鼠胶原诱导性关节炎(Collagen induced arthritis,CIA)的干预效果及作用机制。方法:48只雌性SD大鼠被随机分为空白对照组、CIA模型组、MG132干预模型组,每组16只CIA模型组和MG132干预模型组注射牛Ⅱ型胶原建立CIA模型大鼠,初次免疫后第21天,干预组大鼠以1 mg/kg的剂量每天1次,连续14天皮下注射MG132。建模起每周观察大鼠关节肿胀程度,计算关节炎指数(Arthritis index,AI),第42天后称重并处死大鼠;HE染色观察关节滑膜组织的病理改变;荧光底物测定法检测滑膜组织20S蛋白酶体的活性;蛋白质印迹法(Western blot)检测大鼠关节滑膜组织NF-κB/p65、IκBα蛋白的表达情况。结果:与CIA模型组比较,MG132干预模型组大鼠关节炎指数在注射MG132后一周明显降低(P<0.05),关节滑膜组织未见明显增生,只伴有少量炎性细胞浸润。与空白对照组大鼠比较,CIA模型组大鼠关节滑膜组织20S蛋白酶体活性增高;与CIA模型组大鼠比较,MG132皮下注射干预后关节滑膜组织20S蛋白酶体活性降低(P<0.05)。与空白对照组比较,CIA模型组大鼠关节滑膜组织高表达NF-κB/p65蛋白,其中胞核NF-κB/p65表达增高更为明显(P<0.01),注射蛋白酶体抑制剂MG132干预后,其滑膜组织胞浆及胞核NF-κB/p65蛋白表达均显著减少(P<0.01)。与空白对照组比较,CIA模型组大鼠关节滑膜低表达IκBα蛋白(P<0.01),注射MG132干预后关节滑膜IκBα蛋白表达显著增多(P<0.01)。结论:大鼠CIA体内实验显示,蛋白酶体抑制剂MG132经皮下注射可明显改善大鼠关节炎症状,其作用机制可能与MG132降低大鼠CIA关节滑膜组织20S蛋白酶体活性,减少其底物蛋白IκBα的表达,从而抑制NF-κB活性有关。

Objective ：To explore the intervention effect of proteasome inhibitor MG132 in rats with collagen-induced arthritis （CIA）,which resembles human rheumatoid arthritis（RA）. Methods ： Forty-eight female SD rats were randomly divided into three groups,including blank control group,CIA model group and MG132-treated group. There were sixteen rats in each group. Rats in CIA model group and MG132-treated model group were injected with type Ⅱ collagen to established CIA rats. 21 days after the initial immunization,the rats in the MG132-treated model group were injected subcutaneously with 1 mg/kg MG132 once daily for 2 weeks. 42 days after the initial immunization,the change of paw-swelling and the arthritis scores were determined. The synovial pathology examination was performed with HE staining. The 20S proteasome activity in synovial tissue was measured by fluorescence substrate assay. The expression of NF-κB/ p65, IκBα in synovial tissue were analyzed by Western blot. Results ：Proteasome inhibitor MG132 significantly at-tenuated the severity of arthritis and histopathological changes in CIA rats. Compared with the blank control group,the 20S proteasome activity was increased significantly in the CIA model group （ P 〈 0 .0 5 ） , and decreased after injection of MG132. Compared with CIA rats,the expression of NF-κB/p65 significantly decreased in rats treated with MG132（P〈0. 01）. Compared with the blank control group,the expression of IκBα protein decreased in CIA model group. After injected with MG132, the protein was significantly increased （P〈0. 01）. Conclusion ：The proteasome inhibitor MG132 may attenuates the severity of arthritis and histopathological changes in CIA rats. These effects may be mediated through the inhibition of NF-κB activity.