摘要
目的观察外源碱性成纤维细胞生长因子-2(basic fibroblast growth factor-2,FGF-2)对高糖环境下内皮祖细胞(endothelial progenitor cells,EPCs)功能的影响。方法体外分离和培养小鼠EPCs,并采用流式细胞分析和免疫荧光染色进行细胞鉴定,分别在高糖(30 mmol/L)和高糖(30 mol/L)+FGF-2(30、50、100、150、300 ng/m L)条件下干预培养,正常EPCs作为空白对照组,TUNEL检测细胞凋亡,Transwell小室和基质胶在体外观察EPCs的迁移和成管能力;Western印迹法检测活化β-catenin和总β-catenin的表达,构建小鼠后肢缺血模型,分别注射空白组、正常组、高糖组及高糖+FGF2(150 ng/m L)组EPCs,使用激光多普勒血流仪观察小鼠缺血后肢血流恢复情况。结果高糖组EPCs比空白对照组凋亡率显著增高(P<0.01),迁移和血管形成能力显著降低(P<0.01);高糖+FGF-2组EPCs与高糖组相比,细胞凋亡显著减少(P<0.05),迁移和血管形成能力显著增强(P<0.05),且在150 ng/m L FGF-2时效果最强(P<0.01)。Western印迹法分析表明,高糖组较空白对照组活化β-catenin/总β-catenin显著下降(P<0.01),而高糖+FGF-2组与高糖组相比则显著升高(P<0.05)。体内实验显示,高糖组小鼠缺血后肢较正常组血流恢复显著减少,而高糖+FGF-2组较高糖组显著增加。结论外源性FGF-2处理可能通过激活Wnt/β-catenin信号通路发挥对高糖下受损EPCs的功能修复作用。
Objective To investigate the effects of exogenous basic fibroblast growth factor-2(FGF-2)on the damaged biological activity of endothelial progenitor cells(EPCs)induced by high glucose.Methods Bone marrow endothelial progenitor cells from C57BL/6 mice was isolated,then cultured in vitro and identified by flow cytometry and immunofluorescence staining.EPCs were treated with high glucose(30 mmol/L)and high glucose+FGF-2(30,50,100,150 and 300 ng/mL)respectively,and normal EPCs were taken as a blank control.The cell apoptosis was detected by TUNEL,the migration and tube formation ability of EPCs in vitro were observed by Transwell chamber and Matrigel reagent;the expression of activatedβ-catenin and totalβ-catenin was detected by Western blot.Mouse hind limb ischemia model was constructed;and the ischemic hind limb mice were injected with EPCs obtained from blank,normal high glucose and high glucose+FGF2(150 ng/mL)groups,respectively.The blood flow recovery was observed by laser Doppler flowmetry in mice with ischemic hind limbs.Results EPCs in high glucose group were significantly higher than those in blank control group(P<0.01),and migration and tube formation ability significantly decreased(P<0.01).Compared with high glucose group,cell apoptosis significantly decreased and the migration and tube formation ability significantly increased in the high glucose+FGF-2 group;and the effect of 150 ng/mL FGF-2 was the strongest(P<0.05,P<0.01).Western blotting analysis showed that the ratio of activatedβ-catenin/totalβ-catenin in the high glucose group was significantly lower than that in the control group,while the ratio in high glucose+FGF-2 groups was significantly higher than that in the high glucose group.In vivo experiments showed that the blood flow recovery of the ischemic hind limbs of mice in the high glucose group was significantly lower than that in the normal group,while the blood flow recovery in high glucose+FGF-2 group was significantly higher than that in the high glucose group.Conclusion Exogenous FGF-2 can repair the damaged function of EPCs induced by high glucose,which may be associated with activating Wnt/β-catenin signaling pathway.
作者
李阳
陆晨晖
陆骊工
张海军
滕皋军
李茂全
LI Yang;LU Chen-hui;LU Li-gong;ZHANG Hai-jun;TENG Gao-jun;LI Mao-quan(Dept.of Vascular Intervention and surgery,Tenth People s Hospital,Tongji University,Shanghai 200072,China;Institute of Vascular Intervention,Tongji University School of Medicine,Shanghai 200072,China)
出处
《同济大学学报（医学版）》
CAS
2018年第1期34-40,共7页
Journal of Tongji University(Medical Science)
基金
国家自然科学基金(8157070468)
