Objectives: To explore the effect mechanism of moxibustion with wheat-grain size cone at "Zusanli" (ST 36) on vascular injury and oxidative stress in hyperlipidemia through mammalian target of rapamycin (mTOR)/hypoxia inducible factor-1α (HIF-1α)/vascular endothelial growth factor (VEGF) signaling pathway.
Methods: Forty healthy male SD rats with SPF grade were randomly divided into a normal group, a model group, a moxibustion group, and an inhibitor group, with 10 rats in each one. The hyperlipidemia model was established by feeding a high-fat diet for 8 weeks in rats of the model group, the moxibustion group and the inhibitor group. The moxibustion with wheat-grain size cone was delivered at bilateral "Zusanli" (ST 36) of each rat in the moxibustion group and the inhibitor group, with 3 cones on each acupoint in each intervention, once daily for 4 weeks. In the inhibitor group, before each intervention with moxibustion, rapamycin solution was injected intraperitoneally, 2.0 mg/kg. After modeling and intervention, using ELISA, the levels of total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) in the serum of rats were determined. After intervention, with HE staining and oil red O staining adopted, the abdominal aortic morphology and peripheral lipid deposition were observed. Separately, using WST-1, TBA and micro-plate method, the superoxide dismutase (SOD) activity and the levels of malondialdehyde (MDA) and nitric oxide (NO) in the serum were detected. The protein expression of mTOR, HIF-1α and VEGF in abdominal aorta were measured by Western blot method.
Results: Compared with those in the normal group, the levels of TC, TG and LDL-C increased (P<0.01) and HDL-C decreased (P<0.01) in the serum of the rats in the model group, the moxibustion group and the inhibitor group after model establishment. When compared with the normal group after intervention, in the model group, the serum levels of TC, TG, LDL-C and MDA increased (P<0.01), HDL-C level, SOD activity and NO level were reduced (P<0.01); the cell structure of the abdominal arota was abnormal, the peripheral lipids deposited seriously; and the protein expression of mTOR, HIF-1α and VEGF of abdominal aorta was elevated (P<0.01, P<0.05). In comparison with the model group, the levels of TC, TG, LDL-C and MDA were reduced (P<0.01), HDL-C levels, SOD activities and NO levels elevated (P<0.01, P<0.05), as well as the protein expression of mTOR, HIF-1α and VEGF of abdominal aorta (P<0.01, P<0.05) in the moxibustion group and the inhibitor group; besides, the vascular structure was ameliorated and the lipid deposition reduced in the moxibustion group, while, the vascular structure was still abnormal and the lipid deposition declined in the inhibitor group. When compared with the inhibitor group, the serum SOD activity and NO level increased (P<0.05) and MDA decreased (P<0.05); and the protein expression of mTOR, HIF-1α and VEGF of abdominal aorta was elevated (P<0.01, P<0.05) in the moxibustion group.
Conclusions: The vascular injury due to hyperlipidemia is repaired by moxibustion with wheat-grain size cone at "Zusanli" (ST 36) through ameliorating oxidative stress, which is associated potentially with the modulation of mTOR/HIF-1α/VEGF signaling pathway.
目的: 基于哺乳动物雷帕霉素靶蛋白（mTOR）/缺氧诱导因子-1α（HIF-1α）/血管内皮生长因子（VEGF）信号通路探讨麦粒灸“足三里”改善高脂血症血管损伤及氧化应激的效应机制。方法: 将40只健康SPF级雄性SD大鼠随机分为正常组、模型组、艾灸组和抑制剂组，每组10只。模型组、艾灸组、抑制剂组大鼠采用高脂饲料喂养8周建立高脂血症模型。艾灸组与抑制剂组于双侧“足三里”行麦粒灸干预，每次每穴灸3壮，每日1次，共4周。抑制剂组在每次麦粒灸干预前腹腔注射雷帕霉素溶液（2.0 mg/kg）。分别于造模后及干预后采用ELISA法检测大鼠血清总胆固醇（TC）、三酰甘油（TG）、高密度脂蛋白胆固醇（HDL-C）和低密度脂蛋白胆固醇（LDL-C）含量。于干预后采用HE染色、油红O染色观察大鼠腹主动脉形态及周围脂质沉积；分别采用WST-1、TBA和微板法检测血清超氧化物歧化酶（SOD）活性及丙二醛（MDA）、一氧化氮（NO）含量；Western blot法检测大鼠腹主动脉mTOR、HIF-1α、VEGF蛋白表达。结果: 造模后，与正常组比较，模型组、艾灸组、抑制剂组大鼠血清TC、TG、LDL-C含量升高（P<0.01），HDL-C含量降低（P<0.01）。干预后，与正常组比较，模型组大鼠血清TC、TG、LDL-C、MDA含量升高（P<0.01），HDL-C含量、SOD活性及NO含量降低（P<0.01）；腹主动脉组织细胞结构异常、周围脂质沉积较多；腹主动脉mTOR、HIF-1α、VEGF蛋白表达升高（P<0.01，P<0.05）。与模型组比较，艾灸组和抑制剂组大鼠血清TC、TG、LDL-C、MDA含量降低（P<0.01），HDL-C含量、SOD活性及NO含量升高（P<0.01，P<0.05）；腹主动脉mTOR、HIF-1α、VEGF蛋白表达升高（P<0.01，P<0.05）；艾灸组血管组织结构改善、脂质沉积减少，抑制剂组血管组织结构异常、脂质沉积减少。与抑制剂组比较，艾灸组大鼠血清SOD活性、NO含量升高（P<0.05），MDA含量降低（P<0.05）；腹主动脉mTOR、HIF-1α、VEGF蛋白表达升高（P<0.01，P<0.05）。结论: 麦粒灸“足三里”可通过改善氧化应激进而修复高脂饮食导致的血管损伤，其机制与调控mTOR/HIF-1α/VEGF信号通路有关。.
Keywords: Point ST 36 (Zusanli); hyperlipidemia; mTOR/ HIF-1α/VEGF signaling pathway; moxibustion with wheat-grain size cone; oxidative stress; vascular injury.