目的 研究黄芪甲苷聚乙二醇衍生化磷脂酰乙醇胺(PEG-PE)纳米胶束的制备、细胞内分布及抗心肌细胞凋亡。方法 采用薄膜水化法制备黄芪甲苷PEG-PE(AST-Ⅳ@PEG-PE)纳米胶束,并进行表征观察;采用稳定性实验和体外释药对载药系统进行评价;以香豆素-6作为荧光探针,评价PEG-PE纳米胶束的细胞摄取及细胞内分布;采用10 μmol·L-1异丙肾上腺素诱导H9c2心肌细胞凋亡,将等剂量(50 μmol·L-1)黄芪甲苷和黄芪甲苷PEG-PE纳米胶束在造模前预处理2 h,测定caspase 3活性、活性氧(reactive oxygen species,ROS)水平,Bcl-2和BAX蛋白的表达水平。结果 黄芪甲苷PEG-PE纳米胶束具有粒径小、稳定性良好、释药缓慢等特点;荧光实验表明,PEG-PE纳米胶束可以促进药物的细胞摄取,入胞后,还可将药物聚集在线粒体周围,具有良好的线粒体趋向性;细胞凋亡实验结果表明,黄芪甲苷PEG-PE纳米胶束可以明显降低caspase 3蛋白活性、Bax蛋白表达、ROS水平(P<0.05),显著提高Bcl-2蛋白的表达(P<0.05),这些结果均与等剂量的黄芪甲苷存在显著性差异。结论 AST-Ⅳ@PEG-PE纳米胶束可以很大程度上提高黄芪甲苷的心肌细胞摄取量,并聚集在线粒体周围,增强药物抗心肌细胞凋亡作用。
Abstract
OBJECTIVE To investigate the preparation of astragaloside Ⅳ PEG-PE nanomicelles, its intracellular distribution and the study of its anti-apoptosis of cardiomyocytes. METHODS Astragaloside Ⅳ PEG-PE (AST-Ⅳ@PEG-PE) nanomicelles were prepared by thin film hydration method and characterized and observed. The drug delivery system was evaluated by stability test and in vitro release. Coumarin-6 as a fluorescent probe was used to evaluate the uptake and distribution of PEG-PE micelles in cells. The model of H9c2 cardiomyocyte apoptosis was induced by 10 μmol·L-1 isoproterenol. The same dose (50 μmol·L-1) of astragaloside Ⅳ and astragaloside Ⅳ PEG-PE micelles were pretreated for 2 h before modeling. The caspase 3 activity, ROS level, Bcl-2 and BAX protein expression level were measured. RESULTS Astragaloside Ⅳ PEG-PE nanomicelles had excellent characteristics such as small particle size, good stability and slow drug release. Fluorescence tests showed that PEG-PE nanomicelles could promote the cellular uptake of drugs. After entering the cells, they can also gather the drugs around mitochondria, which has a good mitochondrial tendency. The results of cell apoptosis test showed that astragaloside Ⅳ PEG-PE nanomicelles can significantly reduce caspase 3 protein activity, Bax protein expression, and ROS levels (P<0.05), and significantly increase Bcl-2 protein expression (P<0.05). CONCLUSION These results are significantly better than that of the astragaloside Ⅳ group. Astragaloside Ⅳ PEG-PE can greatly increase the myocardial uptake of astragaloside Ⅳ, aggregate around the mitochondria and enhance the anti-cardiomyocyte apoptosis effect of the drug.
关键词
黄芪甲苷 /
聚乙二醇衍生化磷脂酰乙醇胺纳米胶束 /
线粒体 /
心肌细胞凋亡
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Key words
astragaloside Ⅳ /
PEG-PE nanomicelles /
mitochondria /
cardiomyocyte apoptosis
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中图分类号：
R944
R969
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脚注
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基金
河南省医学科技攻关项目资助(2018020445)
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