Fu Xin-Ying, Sun Zheng-Ji, Long Qing-Yin, Tan Wei, Li Yan-Jun, Wu Lu, He Qing-Hu, Zhang Wei
Hunan Provincial Key Laboratory of Cardiovascular and Cerebrovascular Prevention and Treatment of Integrated Traditional Chinese and Western Medicine,Hunan University of Chinese Medicine Changsha 410208,China.
Yueyang Hospital of Traditional Chinese Medicine Yueyang 414021,China.
Zhongguo Zhong Yao Za Zhi. 2023 Jan;48(1):202-210. doi: 10.19540/j.cnki.cjcmm.20220905.703.
This study aims to explore the effect of Buyang Huanwu Decoction glycosides on the inflammatory response of apolipoprotein E(-/-)(ApoE(-/-)) mice and RAW264.7 cells through nuclear factor kappa-B(NF-κB) signaling pathway. In the in vivo experiment, ApoE(-/-) mice were fed with high-fat diets for 12 weeks to induce the animal model of atherosclerosis, and 75 μg·mL(-1) oxidized low-density lipoprotein(Ox-LDL) incubated RAW264.7 cells for 24 h to establish the atherosclerosis cell model. Automatic biochemical analyzer, hematoxylin-eosin(HE) staining, enzyme-linked immunosorbent assay(ELISA), Western blot, and droplet digital polymerase chain reaction(PCR) were used to determine the blood lipid levels, aortic intimal thickness, inflammatory factor content, NF-κB pathway-related proteins, and mRNA expression levels, and evaluate arterial atherosclerotic lesions and anti-atherosclerotic mechanisms of the drug. The model of atherosclerosis was successfully established in ApoE(-/-) mice after 12 weeks of feeding with high-fat diets. In the model group, the plasma levels of total cholesterol(TC), triglyceride(TG), and low-density lipoprotein cholesterol(LDL-C) were increased(P<0.01), the intima of the blood vessels was thickened, the levels of inflammatory factors tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) were increased, and the protein and mRNA expressions of NF-κB and inhibitor of NF-κB(IκBα) were significantly increased as compared with the control group. Compared with the model group, the high-dose Buyang Huanwu Decoction glycoside group decreased the plasma levels of TC, TG, and LDL-C, reduced the plaque area and thickness and the content of inflammatory factor TNF-α, and inhibited the protein and mRNA expressions of NF-κB and IκBα, with the effect same as Buyang Huanwu Decoction. In the in vivo experiment, 75 μg·mL(-1) Ox-LDL stimulated RAW264.7 cells for 24 h to successfully establish a foam cell model. As compared with the control group, the nuclear amount of NF-κB and the protein and mRNA expressions of IκBα in the model group increased. Compared with the model group, the middle-dose and high-dose Buyang Huanwu Decoction glycoside groups decreased the nuclear amount of NF-κB and the protein and mRNA expressions of IκBα. The above results show that the glycosides are the main effective substances of Buyang Huanwu Decoction against atherosclerosis, which inhibit the NF-κB pathway and reduce the inflammatory response, thus playing the role against atherosclerotic inflammation same as Buyang Huanwu Decoction.
本研究旨在探讨补阳还五汤总苷通过核因子κB（NF-κB）信号通路对载脂蛋白E基因敲除（ApoE-/-）小鼠及RAW264.7细胞炎症反应的影响。在体内实验中，将ApoE-/-小鼠喂饲高脂饲料12周以诱导动脉粥样硬化动物模型，并用75μg·mL-1氧化型低密度脂蛋白（Ox-LDL）孵育RAW264.7细胞24小时以建立动脉粥样硬化细胞模型。采用自动生化分析仪、苏木精-伊红（HE）染色、酶联免疫吸附测定（ELISA）、蛋白质免疫印迹法（Western blot）及液滴数字聚合酶链反应（PCR）检测血脂水平、主动脉内膜厚度、炎症因子含量、NF-κB通路相关蛋白及mRNA表达水平，评价动脉粥样硬化病变及药物的抗动脉粥样硬化机制。喂饲高脂饲料12周后，ApoE-/-小鼠成功建立动脉粥样硬化模型。模型组血浆总胆固醇（TC）、甘油三酯（TG）及低密度脂蛋白胆固醇（LDL-C）水平升高（P<0.01），血管内膜增厚，炎症因子肿瘤坏死因子-α（TNF-α）及白细胞介素-6（IL-6）水平升高，与对照组比较，NF-κB及NF-κB抑制蛋白（IκBα）的蛋白及mRNA表达显著增加。与模型组比较，高剂量补阳还五汤总苷组降低了血浆TC、TG及LDL-C水平，减小了斑块面积和厚度及炎症因子TNF-α含量，抑制了NF-κB及IκBα的蛋白及mRNA表达，其作用与补阳还五汤相同。在体外实验中，用75μg·mL-1 Ox-LDL刺激RAW264.7细胞24小时成功建立泡沫细胞模型。与对照组比较，模型组NF-κB核转位量及IκBα的蛋白及mRNA表达增加。与模型组比较，中剂量和高剂量补阳还五汤总苷组降低了NF-κB核转位量及IκBα的蛋白及mRNA表达。上述结果表明，总苷是补阳还五汤抗动脉粥样硬化的主要有效物质，其通过抑制NF-κB信号通路、减轻炎症反应，发挥与补阳还五汤相同的抗动脉粥样硬化炎症作用。
