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目的:探讨新生大鼠缺氧缺血性脑损伤(HIBD)后大脑皮质分子伴侣介导的自噬(CMA)活性的变化及参附注射液的干预作用。方法:参照Rice法制备HIBD模型。将新生7日龄SD大鼠随机分为假手术组(S组)、生理盐水对照组(C组)、参附注射液治疗组(SF组),每组按照观察时间依次分为6 h、12 h、1 d、3 d、5 d、7 d六个亚组,每个亚组8只大鼠。采用免疫组化法和Western Blot法检测大鼠大脑皮质溶酶体相关膜蛋白-2A(LAMP-2A)的表达情况。结果:免疫组化法和Western Blot法显示LAMP-2A在S组各时间点表达水平差异无统计学意义(P>0.05);C组和SF组LAMP-2A的表达水平在各时间点较S组明显升高(P<0.05),且在3 d达到高峰,随后其表达水平下降;SF组LAMP-2A的表达水平在6 h、12 h、1 d较C组高(P<0.05),但其3 d时峰值表达水平较C组低(P<0.05)。结论:HIBD时存在CMA现象,参附注射液在早期能促进LAMP-2A大量表达,后期则抑制其过度表达,从而对HIBD后的新生大鼠可能具有保护作用。
Objective: To investigate the changes of molecular chaperone-mediated autophagy (CMA) in neonatal rats with hypoxic-ischemic brain damage (HIBD) and the intervention effect of Shenfu injection. Methods: The HIBD model was prepared with reference to Rice method. The neonatal 7-day-old SD rats were randomly divided into sham operation group (S group), saline control group (C group) and Shenfu injection treatment group (SF group). Each group was divided into 6 h, 12 h, 1 d, 3 d, 5 d, 7 d six subgroups, each subgroup of 8 rats. Immunohistochemistry and Western Blot were used to detect the expression of lysosome-associated membrane protein-2A (LAMP-2A) in rat cerebral cortex. Results: Immunohistochemistry and Western Blot showed that there was no significant difference in the expression of LAMP-2A at each time point in S group (P> 0.05). The expression of LAMP-2A in group C and SF at each time point was higher than that of S (P <0.05), and reached a peak at 3 days. The expression level of LAMP-2A in SF group was higher at 6 h, 12 h and 1 d than that of C group (P <0.05) However, the peak expression level at 3 d was lower than that in C group (P <0.05). CONCLUSIONS: The CMA phenomenon exists in HIBD. Shenfu injection can promote the expression of LAMP-2A in early stage and inhibit the overexpression in later stage, which may have a protective effect on neonatal rats after HIBD.