人参皂苷Rb1对川崎病小鼠冠状动脉损伤的作用

Value of ginsenoside Rb1 in alleviating coronary artery lesion in a mouse model of Kawasaki disease

摘要:

目的 探讨人参皂苷Rb1对川崎病（KD）小鼠模型冠状动脉损伤（CAL）的治疗作用及信号通路。方法 将BALB/C小鼠随机分为对照组、模型组、阿司匹林组、人参皂苷Rb1低剂量组（50 mg/kg）和高剂量组（100 mg/kg），每组12只。除对照组外的其他各组均采用间断性腹腔注射10%牛血清白蛋白溶液进行造模；阿司匹林组、Rb1低剂量组和高剂量组分别在造模后给予相应药物灌胃20 d。苏木精-伊红染色观察冠状动脉组织病理变化；ELISA法检测各组小鼠血清和冠状动脉组织中肿瘤坏死因子-α（TNF-α）、白细胞介素（IL）-6和IL-1β等相关炎症因子；Western blot法检测各组小鼠冠状动脉组织中调控自噬信号通路（AMPK/mTOR）和氧化应激信号通路（PI3K/AKT）相关蛋白的相对表达水平。结果 病理切片结果显示，与模型组相比，高剂量Rb1显著改善了CAL小鼠的血管壁增厚、内膜水肿、纤维断裂和内皮细胞炎性浸润等症状。和对照组相比，模型组、Rb1低剂量组小鼠血清及冠状动脉组织中TNF-α、IL-6和IL-1β水平均显著增加（P < 0.05）；模型组小鼠冠状动脉组织中P-AMPK/AMPK、P-mTOR/mTOR和P-P70S6/P70S6表达水平均显著增加（P < 0.05）；而模型组小鼠冠状动脉组织中P-PI3K/PI3K、P-AKT/AKT和P-GSK-3β/GSK-3β表达水平均显著下降（P < 0.05）。和模型组相比，阿司匹林组、Rb1高剂量组小鼠血清及冠状动脉组织中TNF-α、IL-6和IL-1β水平均显著下降（P < 0.05）；Rb1低、高剂量组小鼠冠状动脉组织中P-AMPK/AMPK、P-mTOR/mTOR和P-P70S6/P70S6表达水平均显著下降（P < 0.05），且两个剂量组之间有剂量依赖性（P < 0.05）；Rb1低剂量组小鼠冠状动脉组织中P-PI3K/PI3K表达水平差异无统计学意义（P > 0.05），P-AKT/AKT和P-GSK-3β/GSK-3β表达水平增加（P < 0.05），而Rb1高剂量组上述3种蛋白相对表达水平均显著增加（P < 0.05）。和Rb1低剂量组相比，阿司匹林组和Rb1高剂量组小鼠血清及冠状动脉组织中TNF-α、IL-6和IL-1β水平均显著下降（P < 0.05）；Rb1高剂量组的P-PI3K/PI3K和P-AKT/AKT表达水平均显著增加（P < 0.05）。结论 人参皂苷Rb1可有效减轻KD小鼠模型CAL，疗效与Rb1使用剂量有关。其作用机制可能与通过调控自噬信号通路AMPK/mTOR/P70S6抑制CAL炎症，同时调控氧化应激信号通路PI3K/AKT/GSK-3β发挥保护冠状动脉内皮细胞生物学活性有关。

Abstract:

Objective To study the effect and related signaling pathways of ginsenoside Rb1 in the treatment of coronary artery lesion (CAL) in a mouse model of Kawasaki disease (KD). Methods BALB/c mice were randomly divided into a control group, a model group, an aspirin group, a low-dose ginsenoside Rb1 group (50 mg/kg), and a high-dose ginsenoside Rb1 group (100 mg/kg), with 12 mice in each group. All mice except those in the control group were given intermittent intraperitoneal injection of 10% bovine serum albumin to establish a mouse model of KD. The mice in the aspirin group, the low-dose ginsenoside Rb1 group, and the high-dose ginsenoside Rb1 group were given the corresponding drug by gavage for 20 days after modeling. Hematoxylin and eosin staining was used to observe the pathological changes of coronary artery tissue. ELISA was used to measure the levels of the inflammatory cytokines tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β) in serum and coronary artery tissue. Western blot was used to measure the relative expression levels of proteins involved in the regulation of the AMPK/mTOR autophagy signaling pathway and the PI3K/Akt oxidative stress signaling pathway in coronary artery tissue. Results The observation of pathological sections showed that compared with the model group, the high-dose ginsenoside Rb1 group had significant improvement in the symptoms of vascular wall thickening, intimal edema, fiber rupture, and inflammatory infiltration of endothelial cells. Compared with the control group, the model and low-dose ginsenoside Rb1 groups had significant increases in the levels of TNF-α, IL-6, and IL-1β in serum and coronary artery tissue (P < 0.05); the model group had significant increases in the expression levels of P-AMPK/AMPK, P-mTOR/mTOR, and P-P70S6/P70S6 in coronary artery tissue (P < 0.05) and significant reductions in the expression levels of P-PI3K/PI3K, P-AKT/AKT, and P-GSK-3β/GSK-3β in coronary artery tissue (P < 0.05). Compared with the model group, the aspirin group and the high-dose ginsenoside Rb1 group had significant reductions in the levels of TNF-α, IL-6, and IL-1β (P < 0.05); the low- and high-dose ginsenoside Rb1 groups had significant reductions in the expression levels of P-AMPK/AMPK, P-mTOR/mTOR, and P-P70S6/P70S6 (P < 0.05) in a dose-dependent manner between the two groups (P < 0.05); the low-dose ginsenoside Rb1 group had no significant change in the expression level of P-PI3K/PI3K (P > 0.05) and had significant increases in the expression levels of P-AKT/AKT and P-GSK-3β/GSK-3β (P < 0.05), while the high-dose ginsenoside Rb1 group had significant increases in the relative protein expression levels of the above three proteins (P < 0.05). Compared with the low-dose ginsenoside Rb1 group, the aspirin group and the high-dose ginsenoside Rb1 group had significant reductions in the levels of TNF-α, IL-6, and IL-1β (P < 0.05); the high-dose ginsenoside Rb1 group had significant increases in the expression levels of P-PI3K/PI3K and P-AKT/AKT (P < 0.05). Conclusions Ginsenoside Rb1 can effectively alleviate CAL in a mouse model of KD in a dose-dependent manner, possibly by regulating the AMPK/mTOR/P70S6 autophagy signaling pathway to inhibit CAL inflammation and regulating the PI3K/AKT/GSK-3β oxidative stress signaling pathway to exert a biological activity of protection against coronary artery endothelial cell injury.