天麻钩藤饮对自发性高血压大鼠血管内皮细胞 自噬相关信号通路Ca2+/AMPK/mTOR 的影响

Effect of Tianma Gouteng Decoction on Ca2+/AMPK/mTOR signaling pathway of vascular endothelial cells in spontaneous hypertensive rats

摘要:

目的 探讨天麻钩藤饮对自发性高血压大鼠（SHR）血管内皮细胞自噬相关信号通路Ca2+/AMPK/ mTOR 的影响。方法 将30 只雄性SHR 大鼠随机分为SHR 组、尼莫地平组（5.4 mg/kg）及天麻钩藤饮组 （10.26 g/kg），另设京都种大鼠（WKY）作为对照组，灌胃体积为10 ml/kg，连续给药6 周。采用无创血压仪 分别检测各组给药前、给药2 周后、给药4 周后及给药6 周后的大鼠尾动脉收缩压；采用酶联免疫吸附法 （ELISA）检测给药6 周后各组大鼠血浆血管紧张素Ⅱ（ANG Ⅱ）、白细胞介素1（IL-1）及钙调素（CaM） 水平；采用Western blot 检测给药6 周后各组大鼠血管内皮细胞中自噬标志物（Beclin-1）、微管相关蛋白 轻链（LC3- Ⅱ）及自噬通路相关腺苷酸活化蛋白激酶（AMPK），以及哺乳动物雷帕霉素靶蛋白（mTOR） 的表达情况；采用逆转录聚合酶链反应（RT-PCR）检测给药6 周后各组大鼠血管内皮细胞中自噬通路相 关AMPK、mTOR 基因的表达情况。结果 与WKY 组比较，SHR 组大鼠尾动脉收缩压增高，血浆ANG Ⅱ、 IL-1 及CaM 含量增加，血管内皮细胞中自噬标志物Beclin-1、LC3- Ⅱ和自噬通路相关AMPK、mTOR 蛋白 及mRNA 表达水平均上调（P <0.05）；与SHR 组比较，尼莫地平组和天麻钩藤饮组大鼠血压均降低（P <0.05）， 血浆ANG Ⅱ、IL-1 及CaM 含量下降，且两者血管内皮细胞中自噬标志物Beclin-1、LC3- Ⅱ和自噬通路相 关AMPK、mTOR 蛋白及mRNA 表达水平均降低（P <0.05）。结论 天麻钩藤饮对自发性高血压大鼠血管内 皮细胞自噬相关信号通路Ca2+/AMPK/mTOR 具有明显的调控作用，可能是其降压的重要机制之一。

Abstract:

Objective To investigate the effect of Tianma Gouteng Decoction on the autophagy-related Ca2+/ AMPK/mTOR pathway of vascular endothelial cells in spontaneous hypertensive rats (SHR). Methods In this study 30 male rats with spontaneous hypertension were randomly divided into 3 groups, i.e. an SHR group, a Nimodipine (5.4 mg/kg) group and a Tianma Gouteng Decoction (10.26 g/kg) group. Meanwhile, 10 male WKY rats were employed for a control group. Then all the rats had intragastric administeration of corresponding drugs (10 ml/kg) for consecutive 6 weeks. The systolic blood pressure (SBP) of SHR was separately detected by noninvasive blood pressure meter before drug administration, 2, 4 and 6 weeks after administration. The plasma levels of Ang II, IL-1 and CaM in all groups of rats were detected by enzyme-linked immunosorbent assay after 6 weeks of administration. The expressions of autophagy marker protein Beclin-1, LC3-II, autophagy pathway-related protein AMP and mTOR in the vascular endothelial cells of all the rats were detected by Western blot after 6 weeks of administration. The expressions of autophagy pathway-related AMPK and mTOR mRNAs in vascular endothelial cells of all groups of rats were detected by RT-PCR after 6 weeks of administration. Results Compared with the WKY group, SBP and plasma levels of Ang II, IL-1 and CaM were markedly increased in the SHR group (P < 0.05); the expressions of Beclin-1, LC3- II, AMP and mTOR proteins and mRNAs in the vascular endothelial cells were obviously up-regulated in the SHR group (P < 0.05). Compared with the SHR group, SBP, plasma levels of Ang II, IL-1 and CaM were obviously decreased in both Nimodipine and Tianma Gouteng Decoction groups (P < 0.05); so as the expressions of Beclin-1, LC3-II, AMP and mTOR proteins and mRNAs in the vascular endothelial cells in the two groups (P < 0.05). Conclusions Tianma Gouteng Decoction can significantly regulate the autophagy-related pathway Ca2+/AMPK/mTOR of vascular endothelial cells in spontaneous hypertensive rats, which is probably one of the important mechanisms of its antihypertensive effect.