黄芪甲苷通过SDF-1α/CXCR4轴促进高糖受损人内皮祖细胞迁移的研究

Study of astragaloside IV promoting migration of high glucose(HGS)-impaired human endothelial progenitor cells via SDF-1α/CXCR4 axis

目的:研究黄芪甲苷(AS-Ⅳ)通过SDF-1α/CXCR4轴促进高糖受损人内皮祖细胞(EPCs)迁移的机制,为临床开发AS-Ⅳ用于糖尿病血管新生奠定基础。方法:分别取新生儿脐带血和脐静脉分离培养出人EPCs和人脐静脉内皮细胞(HUVECs)并鉴定。AS-Ⅳ促进高糖受损EPCs和HUVECs分泌SDF-1α和CXCR4:将获得的EPCs和HUVECs分别随机分为实验组、对照组和空白组。Western Blot法检测各组细胞分泌SDF-1α、CXCR4蛋白水平,荧光定量PCR法检测SDF-1α、CXCR4 mRNA表达水平。Transwell实验观察高糖受损EPCs的迁移:Transwell上室接种用最佳浓度AS-Ⅳ预处理的高糖受损EPCs,下室接种用最佳浓度AS-Ⅳ预处理的高糖受损HUVECs,两种细胞共培养,分为实验组、阳性对照组、实验拮抗对照组、模型拮抗对照组,统计穿膜细胞数。结果:与空白组比较,对照组EPCs的SDF-1α和CXCR4蛋白及mRNA表达量显著减少(P<0.05);与对照组比较,实验组EPCs、HUVECs的SDF-1α和CXCR4蛋白及mRNA表达量显著增多(P<0.05)。实验组的穿膜细胞数显著高于实验拮抗对照组和模型拮抗对照组(P<0.05),显著低于阳性对照组(P<0.05)。结论:AS-Ⅳ通过上调SDF-1α/CXCR4轴促进高糖受损人EPCs迁移,从而为糖尿病血管新生创造条件,具有较高的临床开发前景。

Objective:To study the mechanism of astragaloside Ⅳ(AS-Ⅳ) promoting the migration of high glucoseimpaired human endothelial progenitor cells through the SDF-1α/CXCR4 axis,and to lay the foundation for clinical development of AS-Ⅳ for diabetic angiogenesis.Methods:Human endothelial progenitor cells(EPCs) and Human umbilical vein endothelial cells(HUVECs) were isolated and cultured from neonatal umbilical cord blood and umbilical vein,respectively.AS-Ⅳupregulated the secretion of SDF-1α and CXCR4 in HGS-impaired EPCs and HUVECs:EPCs and HUVECs were randomly divided into experimental group,control group and blank group,respectively.The protein levels of SDF-1α and CXCR4 secreted by cells in each group were detected by Western Blot.The expression levels of SDF-1α and CXCR4 mRNA were detected by fluorescence quantitative PCR.EPCs migration ability was tested via Transwell invasion:Transwell was inoculated with high glucose damaged EPCs pretreated with the best concentration of AS-Ⅳ in the upper chamber and HUVECs pretreated with the best concentration of AS-Ⅳ in the lower chamber.The two cells were co-cultured.They were divided into experimental group,positive control group,experimental antagonist control group and model antagonist control group.The number of transmembrane cells was counted.Results:Compared with the blank group,the protein and mRNA expressions of SDF-1α and CXCR4 in EPCs in the control group were less were significantly decreased(P<0.05);Compared with the control group,the protein and mRNA expressions of SDF-1α and CXCR4 in EPCs and HUVECs in the experimental group were significantly increased(P<0.05).The number of transmembrane cells in the experimental group was significantly higher than that in the experimental antagonist group and the model antagonist group(P<0.05);however,the number of transmembrane cells in the experimental group was lower than that in the positive control group(P<0.05).Conclusion:AS-Ⅳ promotes the migration of EPCs in HGS-impaired humans by up-regulating the SDF-1α/CXCR4 axis,thus creating conditions for diabetic angiogenesis,which has a high clinical development prospect.