黄芪甲苷介导外泌体miR-126-5p对糖尿病皮肤溃疡创面血管新生的影响*

作者：胡清睿1，曾楚若2，宋小琴3，广可颜1，刘怡萍4，陈奕仁1，宾马赢2，熊 武2，韩 芳3，张 熙1

单位：1.湖南中医药大学，湖南 长沙 410208； 2.湖南中医药大学第一附属医院，湖南 长沙 410007； 3.湖南省脑科医院，湖南 长沙 410007； 4.湖南中医药大学针灸推拿与康复学院，湖南 长沙 410208

引用：引用：胡清睿，曾楚若，宋小琴，广可颜，刘怡萍，陈奕仁，宾马赢，熊武，韩芳，张熙.黄芪甲苷介导外泌体miR-126-5p对糖尿病皮肤溃疡创面血管新生的影响[J].中医药导报,2026,32(1):14-20.

DOI：10.13862/j.cn43-1446/r.2026.01.003

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摘要：

目的：探讨黄芪甲苷（AS-Ⅳ）介导内皮祖细胞（EPCs）外泌体miR-126-5p对糖尿病皮肤溃疡（DCU）创面血管新生的影响。方法：分离培养人外周血EPCs，通过观察细胞形态、DAPI核染及FITC-UEA-I联合Dil-ac-LDL双荧光染色法鉴定EPCs。将鉴定成功的EPCs分别用PBS、AS-Ⅳ处理，培养24 h收集上清液中EPCexos和ASIV-EPCexos，用透射电镜观察两者形态。另用miR-126-5p inhibitor和inhibitor-NC脂质体转染ASIV-EPCexos，检测上述4种外泌体中miR-126-5p水平。建立2型糖尿病SD大鼠DCU模型。造模成功的大鼠随机分为EPCexos组、ASIV-EPCexos组、ASIV-EPCexos+miR-126-5p inhibitor组、ASIV-EPCexos+inhibitor-NC组、模型组，另设空白组。第10天采集创面新生组织，用蛋白质印迹法（Western blotting）与RT-qPCR法检测VEGFa、VEGFb、VEGFc、FGF、Ang-1血管生长因子表达，免疫荧光和PAS染色检测血管新生。结果：培养24 h后，两组外泌体呈圆形或椭圆形微囊泡结构。与EPCexos组大鼠相比，ASIV-EPCexos组miR-126-5p表达上调（P＜0.05）；与ASIV-EPCexos组大鼠相比，ASIV-EPCexos+miR-126-5p inhibitor组miR-126-5p表达下调（P＜0.05）。与空白组大鼠相比，模型组血管生长因子表达量、VEGF荧光强度、BrdU阳性率和内皮细胞数均显著降低（P＜0.05）；与模型组大鼠相比，ASIV-EPCexo组血管生长因子表达量、VEGF荧光强度、BrdU阳性率和内皮细胞数均增加（P＜0.05）；与ASIV-EPCexo组大鼠相比，ASIV-EPCexo+miR-126-5pinhibitor组血管生长因子表达量、VEGF荧光强度、BrdU阳性率和内皮细胞数均显著降低（P＜0.05）。结论：AS-Ⅳ可通过介导EPCs表达负载miR-126-5p的外泌体促进DCU创面血管新生。

关键词：糖尿病皮肤溃疡；黄芪甲苷；外泌体；miR-126-5p；血管新生

Abstract：

Objective: To investigate the effects of Astragaloside IV (AS-IV) on angiogenesis in diabetic cutaneous ulcers (DCU) and its potential mechanism via mediating exosomal miR-126-5p derived from endothelial progenitor cells (EPCs). Methods: Human peripheral blood EPCs were isolated, cultured, and identified based on morphology, DAPI nuclear staining, and dual-fluorescence staining with FITC-UEA-I and Dil-ac-LDL. Identified EPCs were treated with PBS or AS-IV for 24 h. Exosomes were isolated from the culture supernatants (named EPC-exos and AS-IV-EPC-exos, respectively) and characterized by transmission electron microscopy (TEM). Furthermore, EPCs were transfected with a miR-126-5p inhibitor or inhibitor-NC liposomes, and exosomes were collected from AS-IV-treated transfected cells. The level of miR-126-5p in exosomes from different groups was quantified. A type 2 diabetic SD rat DCU model was established. Successfully modeled rats were randomly divided into EPCexos, ASIV-EPCexos, ASIV-EPCexos + inhibitor-NC, ASIV-EPCexos + miR-126-5p inhibitor and model groups, with a blank group as control. On day 10, wound neotissues were harvested. The protein and mRNA expression levels of vascular growth factors (VEGFa, VEGFb, VEGFc, FGF, Ang-1) were detected by Western blotting and RT-qPCR, respectively. Angiogenesis was assessed by immunofluorescence  and PAS staining. Results: TEM showed that exosomes from both groups exhibited round or oval vesicular structures. The expression of miR-126-5p was significantly upregulated in ASIV-EPCexos group compared with EPCexos group (P<0.05), while The expression of miR-126-5p was significantly downregulated in ASIV-EPCexos+miR-126-5p inhibitor group compared with ASIV-EPCexos group (P<0.05). Compared with the blank group, the model group showed significantly decreased expression of vascular growth factors, VEGF fluorescence intensity, BrdU-positive rate, and endothelial cell number (P<0.05). Compared with the model group, the ASIV-EPCexos group exhibited increased expression of vascular growth factors, VEGF fluorescence intensity, BrdU-positive rate, and endothelial cell number (P<0.05). However, these promoting effects were significantly attenuated in the ASIV-EPCexos + miR-126-5p inhibitor group compared with the ASIV-EPCexos group (P<0.05). Conclusion: AS-IV may promote angiogenesis in DCU wounds by enhancing the enrichment of miR-126-5p in EPC-derived exosomes.

Key words：diabetic cutaneous ulcer; Astragaloside IV; exosomes; miR-126-5p; angiogenesis

发布时间：2026-01-30

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