骨髓基质干细胞种植体外修复内皮及对血管平滑肌细胞增生的影响

目的探讨骨髓基质干细胞(MSCs)种植体外修复内皮的可行性及对血管平滑肌细胞增生的影响。方法培养兔血管内皮、平滑肌和人MSCs,通过细胞共培养模拟血管内皮修复过程,用流式细胞仪分析MSCs分子表型特征,免疫荧光细胞化学法观察与内皮共培养的MSCsFlk1和vWF蛋白表达,根据下室内皮生长状态及是否接种MSCs将其分为对照组、单纯MSCs组、融合内皮组、对数内皮组和MSCs种植组。氚胸腺嘧啶脱氧核苷(3HTdR)掺入检测平滑肌细胞DNA合成,Westernblot检测平滑肌细胞中增殖细胞核抗原蛋白表达。结果分离的MSCs表达基质细胞标志CD105和CD166,不表达造血干祖细胞和内皮细胞标志CD34、Flk1、vWF;与内皮共培养5天时,vWF染色仍为阴性,但约25.71%MSCs开始表达Flk1;MSCs种植组平滑肌细胞3HTdR掺入虽高于融合内皮组,但与对数内皮组比较显著降低;MSCs种植组平滑肌细胞PCNA蛋白吸光度相对值虽高于融合内皮组,但与对数内皮组比较明显减少。结论MSCs种植能抑制平滑肌细胞增生,种植在成熟内皮中的MSCs具有微环境依赖向内皮分化的能力。

Marrow stromal stem cells participating in ex vivo endothelial repair and their effect on vascular smooth muscle cell proliferation

Objective To study the effect of marrow stromal stem cells (MSCs) seeding on smooth muscle cell (SMCs) proliferation and differentiation of the seeded MSCs toward endothelial cells (ECs) for ex vivo endothelial repair.Methods In the cell coculture system, rabbit ECs and human MSCs were inoculated in the lower chamber and rabbit SMCs in the upper chamber. Flow cytometry was used to determine the phenotypic character of MSCs. Fluorescence immunocytochemistry was employed to observe the Flk-1 and vWF protein expression. 3H-TdR incorporation and proliferating cell nuclear antigen (PCNA) protein expression were used to evaluate SMCs proliferation.Results The isolated MSCs expressed CD105 and CD166, but did not express CD34,Flk-1 and vWF. About 25.71% of MSCs began to express Flk-1 protein after cocultured with mature ECs for 5 days. The 3H-TdR incorporation into SMCs in the MSCs seeding group was significantly lower than that in the proliferating ECs group, but higher than that in the confluent ECs group. The PCNA protein expression of SMCs in the MSCs seeding group was also lower than that in the proliferating ECs group, but higher than that in the confluent ECs group.Conclusion MSCs seeding can inhibit SMCs proliferation. MSCs cocultured with mature ECs have the ability to undergo milieu-dependent differentiation toward ECs.