剪应力和血管内皮生长因子诱导骨髓间充质干细胞向内皮细胞分化

Shear Stress Cooperate Vascular Endothelial Growth Factor To Induce Differentiation of Bone Marrow Mesenchymal Stem Cells Into Endothelial Cells

目的研究血管内皮生长因子与剪应力对骨髓间充质干细胞分化的作用。方法分离成人骨髓单个核细胞,经贴壁法获取骨髓间充质干细胞。所获细胞用血管内皮生长因子、剪应力(8和15 dyn/cm2)、血管内皮生长因子联合剪应力分别作用7天、24 h、24 h,观察细胞形态变化,并进行内皮细胞标记物假性血友病因子定性间接免疫荧光染色,进行D il标记的乙酰化低密度脂蛋白摄取实验以鉴定内皮细胞功能。结果在剪应力(8 dyn/cm2)、血管内皮生长因子分别作用24 h、7天后,骨髓间充质干细胞形态变得偏圆、呈多角形,血管内皮生长因子联合剪应力诱导组细胞平行于流体方向排列,假性血友病因子染色阳性,提示内皮细胞表型特征,同时D il标记的乙酰化低密度脂蛋白摄取实验阳性,提示具有内皮细胞功能。血管内皮生长因子诱导组、15 dyn/cm2剪应力组作用24 h的骨髓间充质干细胞,假性血友病因子染色与D il标记的乙酰化低密度脂蛋白摄取实验均阴性,提示未出现向内皮细胞分化。血管内皮生长因子和15 dyn/cm2剪应力共同作用24 h后,骨髓间充质干细胞向内皮细胞分化。结论剪应力可诱导骨髓间充质干细胞向内皮细胞分化,且与力的大小有关。血管内皮生长因子可联合剪应力诱导骨髓间充质干细胞向内皮细胞分化,且诱导效果优于单独使用剪应力。

Aim To investigate the effects of vascular endothelial growth factor （VEGF） and shear stress on the differentiation of bone marrow mesenchymal stem cells （ MSC ）. Methods MSC were isolated from adult marrow mononuclear cells by attaching growth. MSC were subjected to VEGF, different levels of shear stress （8 and 15 dyn/ cm^2 ） and VEGF ＋ shear stress respectively, for 7 d, 24 h or 24 h. Cell shape and orientation were observed at 0 h, 24 h during shear stress. Indirect IF staining was employed to identify the endothelial cell-specific marker von Willebrand factor （ vWF }. A functional marker of mature endothelial cells was examined by the uptake of Dil labled acetylated low density lipoproteins （Dil-ac-LDL）. Results After subjection to VEGF, shear stress of 8 dyn/cm^2 for 7 d, 24 h respectively, MSC tended to be a more rounder, polygon-like cell shape, and MSC of shear stress group aligned parallel to the flow, MSC were positive for vWF staining, indicating the endothelial phenotype, and also positive for the uptaking of Dil-ac-LDL, indicating the possession of-mature endothelial function. Cells treated with VEGF and shear stress of 15 dyn/ cm^2 for 24 h were negative for both the vWF staining and the uptaking of Dil-ac-LDL, indicating the lack of differentiation. After VEGF and shear stress acted 24 h together, MSC can be induced into endothelial cell. Conclusion Shear stress can induce the differentiation of MSC into endothelial cells in a stress level dependent way. Shear stress combined with VEGF not only induce the differentiation of MSC into endothelial cells, but also cause cell proliferation, which seems to have a better effect than only shear stress.