人参皂苷Rg1对人脐血间充质干细胞活性和成血管分化的影响

Effects of ginsenoside Rg1 on viability and vascular differentiation of human umbilical blood mesenchymal stem cells

目的探讨人参皂苷Rg1(GS-Rg1)对人脐血间充质干细胞(MSC)体外活性和向血管内皮分化的影响。方法无菌条件下取正常剖宫产胎盘中的人脐血,经密度梯度离心法分离得到单个核细胞,细胞培养呈长梭形时,进行成骨、成软骨及成脂诱导分化鉴定。将通过鉴定的人脐血MSCs分别用0、5、10、20、40、80 mg/L的GS-Rg1进行干预,确定GS-Rg1促人脐血MSCs增殖的最佳浓度。然后将人脐血MSCs分为实验组和对照组,用最佳浓度的GS-Rg1对实验组进行处理,对照组采用等体积的磷酸盐缓冲液处理。使用CCK-8法研究两组细胞活性,Matrigel实验研究两组体外血管新生的情况。同时用免疫荧光检测内皮细胞标志物血小板-内皮细胞黏附分子-1(CD31)、血管性血友病因子(vWF)表达情况。结果显微镜下可见细胞形态均一,呈长梭形的成纤维细胞样形态,是MSCs的典型形态。成骨、成软骨和成脂诱导分化鉴定染色结果均可见大量深染色部分,显示其为MSCs。GS-Rg1浓度为40、80 mg/L时,人脐血MSCs光密度值高于其浓度为0、5、10、20 mg/L时,差异有统计学意义(P<0.05)。GS-Rg1浓度为80 mg/L时,人脐血MSCs光密度值与其浓度为40 mg/L时比较,差异无统计学意义(P>0.05)。实验组细胞活性、管腔长度及CD31、vWF荧光强度高于对照组,差异有统计学意义(P<0.05)。结论GS-Rg1具有促进人脐血MSCs体外增殖和诱导其向内皮细胞分化及体外血管生成的潜能。

Objective To investigate the effects of ginsenoside Rg1(GS-Rg1)on the activity and endothelial differentiation of human umbilical cord blood mesenchymal stem cell(MSC)in vitro.Methods Human umbilical blood was collected from normal cesarean placenta under aseptic conditions,and mononuclear cells were isolated by density gradient centrifugation.When the cells were fusiform,osteogenesis,chondrogenesis,and adipogenic differentiation were identified.The identified human umbilical blood MSCs were treated with 0,5,10,20,40,and 80 mg/L GS-Rg1,respectively,to determine the optimal concentration of GS-Rg1 to promote the proliferation of human umbilical blood MSCs.Then human umbilical blood MSCs were divided into experimental group and control group.The experimental group was treated with optimal concentration of GS-Rg1,and the control group was treated with equal volume of phosphate buffer.CCK-8 method was used to study the cell activity of the two groups,and Matrigel experiment was used to study in vitro angiogenesis of the two groups.Meanwhile,the expression of platelet-endothelial cell adhesion molecule-1(CD31)and von Willebrand factor(vWF)were detected by immunofluorescence.Results Microscopically,the morphology of the cells was uniform,showing a long spindle fibroblast-like morphology,which was typical of MSCs.The staining results of osteogenic,chondrogenic,and adipogenic differentiation showed a large number of deep staining parts,indicating that they were MSCs.When GS-Rg1 concentration was 40 and 80 mg/L,the MSCs optical density value was higher than that when GS-Rg1 concentration were 0,5,10,and 20 mg/L,the differences were statistically significant(P<0.05).When GS-Rg1 concentration was 80 mg/L,there was no significant difference in the optical density of MSCs when GS-Rg1 concentration was 40 mg/L(P>0.05).The cell activity,lumen length,and fluorescence intensity of CD31 and vWF in experimental group were higher than those in control group,and the differences were statistically significant(P<0.05).Conclusion GS-Rg1 has the potential to promote the proliferation of human umbilical blood MSCs in vitro and induce their differentiation into endothelial cells and angiogenesis in vitro.