目的 观察人参皂苷(GS-Rg1)对人内皮祖细胞(EPCs)增殖、黏附、迁移、成管能力及血小板衍生生长因子(PDGF)分泌水平的调控作用.方法 取足月健康新生儿脐带血,培养并鉴定EPCs,将EPCs随机分为实验组和对照组,实验组给予40 mg/L的GS-Rg1,对照组给予等量PBS.培养48 h后,采用CCK-8试剂盒检测细胞增殖能力,黏附能力测定实验检测细胞黏附能力,细胞划痕实验检测细胞迁移能力,Matrigel体外成血管实验检测细胞成管能力,采用ELISA法检测细胞培养液上清中的PDGF.结果 与对照组相比,实验组EPCs增殖能力增高,细胞黏附数增多,细胞绝对迁移宽度增宽,体外成管数增多,细胞培养液上清中PDGF水平增高(P均<0.05).结论 GS-Rg1能提高人EPCs的增殖、黏附、迁移、成管能力,并促进其分泌PDGF.

Objective:To investigate the effect of astragaloside IV (AS-IV) on the secretion of stromal cell-derived factor-1α (SDF-1α) and CXC chemokine receptor 4 (CXCR4) by high glucose injured human umbilical vein endothelial cells (HUVECs), so as to lay a foundation for further study on AS-IV improving angiogenesis by regulating SDF-1 α/CXCR4 axis of endothelial cells.Methods:HUVECs were isolated and cultured from the umbilical vein of full-term healthy newborns and identified by von Willebrand factor (vWF) combined with 4-diamino-2-phenylindole (DAPI) nuclear staining. The obtained HUVECs was cultured in EGM-2 medium with 30 mmol/L glucose for 120 h to obtain high glucose damaged HUVECs. After intervention with different concentration gradients (25 mg/L, 50 mg/L, 100 mg/L, 200 mg/L, 400 mg/L) AS-IV for 72 hours, the contents of SDF-1α and CXCR4 were detected by enzyme linked immunosorbent assay (ELISA) method to determine the best concentration of AS-IV. The supernatant of damaged HUVECs were collected at 6, 12, 24, 48 and 72 hours after intervention with the best concentration of AS-IV, and the contents of SDF-1α and CXCR4 were detected by ELISA method to determine the best action time of AS-IV. The damaged HUVECs was randomly divided into experimental group and control group, and the blank group was set up at the same time. The experimental group was treated with the best concentration of AS-IV and the best time, the control group and the blank group were treated with the same volume of phosphate buffered saline (PBS) solution, and the contents of SDF-1α and CXCR4 in each group were detected by ELISA method.Results:The vWF factor on the cell membrane was green fluorescence, and the nucleus was blue after DAPI staining. When the fusion image showed green fluorescence, HUVECs were identified by blue fluorescence. The expression of SDF-1α in damaged HUVECs was the best when treated with AS-IV of 100 mg/L for 24 hours (1 642.87 pg/ml), and the expression of CXCR4 in damaged HUVECs was the best when treated with AS-IV of 50 mg/L for 48 hours (8.44 ng/ml). Compared with the control group, the contents of SDF-1α and CXCR4 in the experimental group were significantly increased, and the difference was statistically significant ( P<0.05). While the contents of SDF-1α and CXCR4 in the experiment group were slightly less than those in the blank group and there was no statistically significant difference ( P>0.05). Conclusions:AS-IV can promote the expression of SDF-1α and CXCR4 in HUVECs damaged by high glucose to return to normal physiological level, so as to play the role of vascular repair and neovascularization.

目的 探讨黄芪甲苷(AstragalosideⅣ,AS-Ⅳ)对人内皮祖细胞(endothelial progenitor cells,EPCs)生物学功能的影响,为深入研究AS-Ⅳ调节EPCs介导的血管新生的作用机制奠定基础.方法 取足月健康新生儿脐带血,采用密度梯度离心法分离得到单个核细胞,经传代培养,采用CD31抗体联合DAPI核染以及FITC-UEA-I和Dil-ac-LDL双荧光染色法鉴定EPCs.将鉴定成功的EPCs随机分为实验组和对照组,实验组采用100 mg/L的AS-IV干预,对照组用等容量的PBS液处理,两组细胞培养24 h后,通过CCK-8细胞增殖检测试剂盒、黏附能力测定试验、细胞划痕试验及Matrigel体外成血管试验观察AS-IV对EPCs增殖、黏附、迁移和成管功能的影响.结果 与对照组相比,实验组细胞增殖的OD值增大,细胞黏附数增多,细胞迁移宽度增加(即细胞迁移率增大),体外成管数增多,差异均有统计学意义(P<0.05).结论 黄芪甲苷能改善体外人EPCs的生物学功能,具有调节EPCs介导血管新生的潜能.

Objective The objective is to probe into the effects of astragaloside Ⅳ (AS-Ⅳ) on activity and proliferative function of endothelial progenitor cells (EPCs),which lays a foundation for further study on the effects of AS-Ⅳ on vascular neovascularization mediated by endothelial progenitor cells.Methods The mononuclear cells were isolated by the density gradient centrifugation in umbilical cord blood of full-term healthy infants,and EPCs were obtained by subculture and cell identification when the cells presented spindle shapes.The obtained EPCs were randomly divided into the experimental group and the control group.In the experimental group,EPCs were cultured by AS-Ⅳ with different concentration gradients (25 mg/L,50 mg/L,100 mg/L,200 mg/L and 400 mg/L),while in the control group,they were treated with the same amount of phosphate buffer saline (PBS) solutions.The effects of AS-Ⅳ on the proliferation of endothelial progenitor cells was studied by cell counting kit-8 (CCK-8) cell proliferation experiment,and the activity rate of EPCs cells was measured at the optimum concentration of EPCs proliferation.Results EPCs were successfully obtained after confirming nuclear staining test of CD31 antibody and 4',6-diamidi-no-2-phenylindole (DAPI).Further study showed that AS-Ⅳ can promote the proliferation of EPCs,and its optimal concentration of EPCs proliferation is 100 mg/L.Compared with the normal control group,the activity rate of endothelial progenitor cells after intervention of AS-Ⅳ was 98.7％,higher than 98.12％ in the control group,with significant difference (x2 =49.59,P ＜0.01).Conclusions AS-Ⅳ can enhance the activity of human EPCs and promote their proliferation in vitro.

目的 观察积雪草苷对兔耳创伤修复不同时期及转化生长因子β1(TGF-β1)、Smad3免疫组织化学表达的影响.方法 将24只新西兰白兔按照随机数字表法分为生理盐水1组、生理盐水2组、积雪草苷1组、积雪草苷2组4组,每组6只.积雪草苷干粉制备成1 000 mg/mL的注射液,现备现用.制作兔耳直径1 cm全层皮肤缺损模型,造模后5 d(创伤修复增殖期),生理盐水1组、积雪草苷1组每日8:00、20:00于兔耳创面处分别注射0.9％氯化钠注射液、积雪草苷液,每次0.2 mL,连用15 d.造模后30 d(创伤修复塑形期),生理盐水2组、积雪草苷2组每日8:00、20:00于兔耳创面处分别注射0.9％氯化钠注射液、积雪草苷液,每次0.2 mL,连用15 d.生理盐水1组、积雪草苷1组于造模后0、5、20 d进行拍照做肉眼观察,生理盐水2组、积雪草苷2组于造模后0、30、45 d进行拍照做肉眼观察;生理盐水1组和积雪草苷1组,生理盐水2组和积雪草苷2组分别于造模后20、45 d后处死动物,切取耳创伤面标本,进行免疫组织化学染色.结果 与生理盐水1组比较,积雪草苷1组可以明显促进创伤修复,炎性细胞明显减少,可见大量的成纤维细胞和毛细血管形成.积雪草苷2组瘢痕组织增生层明显比生理盐水2组薄,且未见丰富的血管和成纤维细胞,胶原纤维含量降低,排列稀疏.积雪草苷1组TGF-β1、Smad3的表达量均高于生理盐水1组(P＜0.05);积雪草苷2组TGF-β1、Smad3的表达量均低于生理盐水2组(P＜0.05).结论 积雪草苷在兔耳创伤修复增殖期干预促进瘢痕形成,但在兔耳创伤修复塑形期干预减少瘢痕形成.