世界中医药

目的：通过研究复方黄柏液对糖基化内皮细胞增殖迁移成管的影响，寻找复方黄柏液治疗糖尿病足溃疡的依据。方法：为说明晚期糖基化终产物受体（RAGE），用晚期糖基化终产物（AGEs）诱发内皮细胞功能障碍的重要受体，本研究设置了RT-PCR实验沉默了RAGE，结果发现沉默RAGE后，经AGEs处理内皮细胞，检测其内皮细胞增殖迁移成管作用出现上升，因此检测RAGE表达水平的变化能作为后续评价复方黄柏液改善内皮细胞功能的重要指标。如果RAGE水平出现下降，就证明内皮细胞功能障碍得到了修复，复方黄柏液治疗糖尿病足溃疡是有效的。为了评估内皮细胞的增殖能力，设置CCK-8实验；用划痕实验检查内皮细胞的迁移水平变化；通过成管实验来测试内皮细胞的成管作用；利用蛋白质印迹法检测RAGE蛋白表达水平。结果：RAGE-3 siRNA沉默效率最好，可用于后续实验。与空白组比较，AGEs处理后内皮细胞增殖活性、迁移水平、成管作用都下降，RAGE蛋白表达水平上升。与AGEs-BSA处理组比较，沉默RAGE或加入复方黄柏液后均较AGEs处理后内皮细胞的增殖迁移成管作用有了提高，RAGE蛋白的表达下降，差异有统计学意义（P<0.05）。结论：复方黄柏液可能通过改善AGEs诱发的内皮细胞功能障碍即促进内皮细胞增殖迁移成管来实现对糖尿病足溃疡的愈合。

English Summary:

This study aims to clarify the mechanism of Compound Fluid of Cortex Phellodendri in treating diabetic foot ulcer based on the influence of the prescription on the proliferation，migration，and tube formation of endothelial cells treated with advanced glycation end products(AGEs).Methods:In order to show that receptor for advanced glycation end products(RAGE) is an important receptor for AGEs-induced endothelial cell dysfunction，we performed RT-PCR to confirm the silencing of RAGE.We found that the proliferation，migration，and tube forming ability of endothelial cells treated by AGEs improved after the silencing of RAGE.Therefore，RAGE can serve as important index for evaluating the improvement of endothelial cell function by Compound Fluid of Cortex Phellodendri.If RAGE level dropped，it proved that endothelial cell dysfunction had been alleviated and that Compound Fluid of Cortex Phellodendri was effective for diabetic foot ulcer.Cell counting kit-8(CCK-8) assay was employed to evaluate the proliferation of endothelial cells，scratch assay to check the migration of the cells，and tube forming experiment to test the tube forming ability.Western blotting was used to detect the expression level of RAGE protein.Results:The silencing efficiency of siRNA on RAGE-3 was the highest and thus it was used for subsequent experiments.Compared with normal control group，AGEs treatment decreased the proliferation activity，migration level，and tube forming ability of endothelial cells and increased the RAGE protein level.Compared with AGES-BSA treatment group，the proliferation，migration，and tube forming ability of endothelial cells were improved and the expression of RAGE protein was decreased after the addition of Compound Fluid of Cortex Phellodendri or the silencing of RAGE(P<0.05).Conclusion:Compound Fluid of Cortex Phellodendri may improve the dysfunction of endothelial cells induced by AGEs to relieve diabetic foot ulcers.

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