温肾醒脑方对血管性痴呆大鼠脑组织Nrf2-ARE信号通路的影响

为探讨温肾醒脑方(WXD)对血管性痴呆(vascular dementia,VD)大鼠认知功能的改善作用及对Nrf2-ARE信号通路的影响,将70只雄性SD大鼠随机分成正常组、假手术组、模型组、脑复康组(0.15 g/kg)、温肾醒脑方给药组(高剂量组5 g/kg、中剂量组2.5 g/kg、低剂量组1.25 g/kg),每组10只动物,采用双侧颈总动脉永久性结扎法建立VD模型,给予温肾醒脑方灌胃治疗后,Morris水迷宫测试治疗后VD大鼠认知功能的改善情况,检测脑组织中超氧化物歧化酶(SOD)、丙二醛(MDA)、谷胱甘肽过氧化物酶(GSH-Px)、一氧化氮合酶(NOS),RT-PCR检测大鼠脑组织醌NADH脱氢酶(NQO1)、血红素氧合酶1(HO-1)的mRNA水平,运用Western Blot检测Kelch样环氧氯丙烷相关蛋白-1(Keap1)、核因子E2相关因子2(Nrf2)、NQO1、HO-1蛋白水平的表达情况以及EMSA检测Nrf2与抗氧化反应元件(ARE)结合力。结果表明:与正常组比较,VD模型组大鼠逃避潜伏期明显较长,游泳总距离明显缩短,穿越站台总数明显减少(P0.01);大脑组织中MDA含量显著增加,SOD、GSH-Px和NOS的含量明显减少(P0.01);同时大脑组织中细胞核Nrf2蛋白表达降低、细胞质中Keap1蛋白表达增加,NQO1、HO-1 mRNA和蛋白表达降低(P0.01)。与VD模型组比较,脑复康和温肾醒脑方组大鼠的认知行为得到很大的提高,脑组织MDA、SOD、GSH-Px、NOS含量和脑组织病理结构得到很好的改善,Nrf2、Keap1、NQO1、HO-1在mRNA水平或蛋白水平上有一定程度改变,特别是温肾醒脑方组大鼠的改善非常显著。可见温肾醒脑方可以改善VD大鼠的认知行为,其机制可能与Nrf2-ARE抗氧化应激信号通路相关。

Effect of Wenshen Xingnao Decoctione on Nrf2-ARE Signaling Pathway in Brain Tissue of Vascular Dementia Rats

To study the effect of Wenshen Xingnao Decoction on cognitive function and Nrf2-ARE signaling pathway in vascular dementia (VD) rats. 70 male SD rats were randomly divided into normal group, sham-operated group, model group, Naofukang group (0.15 g/kg), Wenshen Xingnao Decoction group (5 g/kg in high-dose group, 2.5 g/kg in medium-dose group and 1.25g/kg in low-dose group), with 10 animals in each group. VD model was established by permanent ligation of bilateral common carotid artery. After treatment with Wenshen Xingnao Decoction, Morris water maze test improved the cognitive function of VD rats. SOD, MDA, GSH-Px and NOS in brain tissue were detected. The levels of NQO1 and HO-1 in brain tissue of rats were detected by RT-PCR and the expression of Keap1, Nrf2, NQO1 and HO-1 protein by Western blot. The binding ability of Nrf2 to ARE was detected by EMSA.The results show that:Compared with the normal group, the escape latency of VD model group was longer, the total distance of swimming was shorter, the total number of crossing platforms was decreased (P<0.01); the content of MDA in brain tissue increased significantly, the content of SOD, GSH-Px and NOS decreased significantly (P<0.01); meanwhile, the expression of Nrf2 protein in brain tissue decreased, the expression of Keap1 protein in cytoplasm increased, and the content of NQO1 protein decreased significantly (P<0.01). The expression of HO-1 and HO-1 decreased (P<0.01). Compared with VD model group, the cognitive behavior of rats in Naofukang and Wenshen Xingnao Decoction groups was greatly improved. The contents of MDA, SOD, GSH-Px, NOS and pathological structure of brain tissue were improved. Nrf2, Keap1, NQO1 and HO-1 had some changes at the level of gene or protein, especially in Wenshen Xingnao Decoction group. So Wenshen Xingnao Decoction can improve cognitive behavior of VD rats, and its mechanism may be related to Nrf2-ARE antioxidant stress signaling pathway.