水蛭对ApoE^(-/-)动脉粥样硬化小鼠自噬的影响

Effect of Hirudo on autophagy in atherosclerotic mice

目的 探讨水蛭对动脉粥样硬化小鼠自噬的影响。方法 将40只健康雄性ApoE^(-/-)小鼠随机均分为模型组、对照组(3×10^(-3) g·kg^(-1)·d^(-1)辛伐他汀)及低、中、高剂量实验组(0.45、0.9、1.8 g·kg^(-1)·d^(-1)脉血康胶囊),8只健康雄性C57BL/6J小鼠为空白组。给予普通饲料适应性喂养1周后,除空白组外,其余各组采用高脂饲料喂养8周建立动脉粥样硬化小鼠模型,之后按照设定剂量灌胃给药,每日1次,连续给药12周,期间继续给予高脂饲料饮食。用酶联免疫吸附测定(ELISA)法检测小鼠血清中肿瘤坏死因子(TNF-α)、白细胞介素-6(IL-6)水平;用蛋白质印迹法检测Beclin-1、LC3自噬蛋白水平。结果 高、中、低剂量实验组和对照组、模型组、空白组的IL-6含量分别为(107.59±3.03)、(99.31±5.12)、(103.52±2.28)、(98.68±4.68)、(112.66±6.08)和(93.98±3.43)pg·mL^(-1),TNF-α含量分别为(538.41±30.26)、(504.49±21.51)、(538.51±19.05)、(494.05±25.08)、(578.53±26.32)和(467.35±21.53)pg·mL^(-1),模型组的上述指标与实验组、对照组和空白组相比,在统计学上差异均有统计学意义(均P<0.05)。高、中、低剂量实验组和对照组、模型组的Beclin-1蛋白表达水平分别为1.48±0.05、1.72±0.05、1.19±0.02、1.51±0.04和0.66±0.03,LC3Ⅱ蛋白表达水平分别为1.53±0.01、1.83±0.02、1.16±0.01、1.90±0.01和0.49±0.01,模型组的上述指标与低、中、高实验组和对照组相比,在统计学上差异均有统计学意义(均P<0.05)。结论 水蛭可能通过调节斑块内细胞自噬水平,从而达到抗动脉粥样硬化的作用。

Objective To explore the anti-atherosclerosis mechanism of Hirudo and its effect on autophagy in mice.Methods Forty healthy male ApoE~(-/-) mice were randomly divided into model group,control group(3×10~(-3) g·kg~(-1)·d~(-1) simvastatin) and experimental-L,experimental-M,experimental-H groups(0.45,0.9,1.8 g·kg~(-1)·d~(-1),Maixuekang capsule).Eight healthy male C57BL/6J mice were divided into blank group.The mice were fed with common diet for 1 week.Then,except blank group,other groups were fed with high-fat diet.After 8 weeks of modeling,the atherosclerosis(AS) mice were given drugs orally once a day for 12 weeks,and fed with high-fat diet in the meantime.The levels of interleukin-6(IL-6),tumor necrosis factor-α(TNF-α)in serum were determined by enzyme-linked immunosorbent assay(ELISA).The levels of Beclin-1,LC3 autophagy protein were detected by Western blot method.Results The IL-6 contents in the experimental-H,experimental-M,experimental-L,control,model and blank groups were(107.59±3.03),(99.31±5.12),(103.52±2.28),(98.68±4.68),(112.66±6.08),(93.98±3.43)pg·mL~(-1);the TNF-α contents were(538.41±30.26),(504.49±21.51),(538.51±19.05),(494.05±25.08),(578.53±26.32),(467.35±21.53) pg · mL~(-1).For the above indexes,the differences between model group and experimental-H group,experimental-M group,experimental-L group,control group,blank group were all statistically significant(allP< 0.05).The Beclin-1 protein expression levels in the experimental-H,experimental-M,experimental-L,control and model groups were 1.48±0.05,1.72±0.05,1.19±0.02,1.51±0.04,0.66±0.03;the LC3 Ⅱ protein expression levels were 1.53±0.01,1.83±0.02,1.16±0.01,1.90±0.01,0.49±0.01,and the differences between model group and experimental-H group,experimental-M group,experimental-L group,control group were all statistically significant(allP< 0.05).Conclusion Hirudo can significantly reduce the area of atherosclerotic plaque by regulating the level of autophagy.