黄芩苷治疗小鼠动脉粥样硬化模型的作用与机制

Role of baicalin in the treatment of mouse models of atherosclerosis and the underlying mechanism

背景:现代实验室及临床研究表明,黄芩苷可通过抗氧化应激、抑制血管平滑肌细胞增殖和迁移等多途径发挥抗动脉粥样硬化效应,但其作用机制尚不明确。目的:观察黄芩苷对动脉粥样硬化小鼠血脂、一氧化氮及核因子κB与其下游炎症因子水平的影响。方法:将60只雄性ApoE-/-小鼠随机分为5组,每组10只:对照组给予普通饲料喂养,其余5组给予高脂饲料喂养12周,建立动脉粥样硬化模型;随后对照组、模型组灌胃给予生理盐水,阿托伐他汀组灌胃给予阿托伐他汀5mg/(kg·d),黄芩苷低、中、高剂量组分别灌胃给予50,75,100mg/(kg·d)黄芩苷。连续给药4周后,采用全自动生化仪测定血脂水平,油红O染色观察主动脉斑块变化,ELISA法检测血清炎症因子水平,Western blot检测主动脉中核因子κB p65、血管细胞黏附分子1蛋白表达,硝酸酶还原法测量主动脉一氧化氮水平,实时荧光定量RT-PCR检测主动脉组织核因子κB p65、血管细胞黏附分子1、肿瘤坏死因子αmRNA的表达。结果与结论:①与对照组比较,模型组总胆固醇、三酰甘油、低密度脂蛋白水平升高(P <0.05),高密度脂蛋白水平降低(P <0.05),主动脉内膜厚度与斑块截面积增加(P <0.05),炎症因子肿瘤坏死因子α、白细胞介素1水平升高(P<0.05),NO水平降低(P<0.05),核因子κB p65、血管细胞黏附分子1蛋白表达升高(P <0.05),核因子κB p65、血管细胞黏附分子1、肿瘤坏死因子αmRNA表达升高(P <0.05);②与模型组比较,阿托伐他汀组及黄芩苷低、中、高剂量组血脂水平明显改善(P <0.05),炎症因子水平降低(P <0.05),NO水平升高(P <0.05),主动脉内膜厚度与斑块截面积减少,核因子κB p65、血管细胞黏附分子1蛋白表达降低(P <0.05),核因子κB p65、血管细胞黏附分子1、肿瘤坏死因子αmRNA表达降低(P <0.05),其中黄芩苷的改善作用存在剂量依赖性;③与阿托伐他汀组比较,黄芩苷高剂量组总胆固醇、白细胞介素1、血管细胞黏附分子1蛋白及基因表达降低(P <0.05),NO水平升高(P <0.05);④结果表明,黄芩苷可能通过改善血脂与抑制核因子κB炎症通路等途径对抗动脉粥样硬化形成。

BACKGROUND: Modern laboratory and clinical studies have shown that baicalin can exert anti-atherosclerosis effect through multiple pathways, such as antioxidant stress, inhibition of proliferation and migration of vascular smooth muscle cells, but its mechanism is still unclear. OBJECTIVE: To investigate the effect of baicalin on serum lipid, nitric oxide, nuclear factor kappa B, and its downstream inflammatory cytokines in mouse models of atherosclerosis. METHODS: Sixty male ApoE-/-mice were randomized into five groups(n=10 per group). The control group was fed with normal diet, and the other five groups were fed with high-fat diet for 12 weeks to establish the atherosclerotic model. Subsequently, the control group and model group were given saline by gavage. The atorvastatin group was given atorvastatin 5 mg/(kg·d) by gavage. The baicalin group was given baicalin 50, 75, and 100 mg/(kg·d) by gavage. After 4 weeks of administration, blood lipid level was detected by automatic biochemical analyzer. Oil red O staining was used to observe development of the atherosclerotic plaques. Serum levels of inflammatory cytokines were detected by ELISA. The protein expression levels of nuclear factor kappa B p65 and vascular cell adhesion molecule-1 in aorta were detected by western blot assay. The level of nitric oxide was detected by nitrate reductase assay. The mRNA expression levels of nuclear factor kappa B p65, vascular cell adhesion molecule-1 and tumor necrosis factor alpha in aortic tissue were detected by real-time fluorescence quantitative PCR. RESULTS AND CONCLUSION:(1) Compared with the control group, the total cholesterol, triacylglycerol and low-density lipoprotein levels were increased(P < 0.05), high-density lipoprotein cholesterol level was decreased(P < 0.05), the aortic intima thickness and plaques area were increased(P < 0.05), tumor necrosis factor alpha and interleukin 1 levels were increased(P < 0.05), nitric oxide level was decreased(P < 0.05), the protein expression of nuclear factor kappa B p65 and vascular cell adhesion molecule-1 was increased(P < 0.05), the mRNA expression levels of nuclear factor kappa B p65, vascular cell adhesion molecule-1 and tumor necrosis factor alpha were increased(P < 0.05) in the model group.(2) Compared with the model group, the lipid level in the atorvastatin and the low-, medium-and high-dose baicalin groups was improved significantly(P < 0.05), inflammatory factor levels were decreased(P < 0.05), nitric oxide level was increased(P < 0.05), the aortic intima thickness and plaques area were reduced, the protein expression of nuclear factor kappa B p65 and vascular cell adhesion molecule-1 was reduced(P < 0.05), and the mRNA expression levels of nuclear factor kappa B p65, vascular cell adhesion molecule-1 and tumor necrosis factor alpha were reduced(P < 0.05). The improvement of baicalin was in a dose-dependent manner.(3) Compared with the atorvastatin group, the expression levels of total cholesterol, interleukin 1 and vascular cell adhesion molecule-1 in the high-dose baicalin group were decreased(P < 0.05) and the level of nitric oxide was increased(P < 0.05).(4) These results show that baicalin may prevent the formation of atherosclerosis by improving blood lipid and suppressing the inflammatory pathway of nuclear factor kappa B.