诱导脂肪干细胞向血管内皮细胞分化的实验研究

Cellular phenotype of adipose-derived stem cells into endothelial cells

目的 探讨诱导脂肪干细胞（ADSCs）向血管内皮细胞（ECs）分化的可能性,为ADSCs应用于创伤修复的理论提供实验依据.方法 切取大鼠脂肪组织,用酶消化法分离、培养ADSCs,流式细胞仪检测第3代ADSCs的CD49d和CD106的表达以鉴定ADSCs.实验组用含30%大鼠血管匀浆液的条件培养液诱导大鼠ADSCs,空白对照组用含10%胎牛血清DMEM培养液培养大鼠ADSCs,各组诱导3 d后经免疫组化和流式细胞仪检测ADSCs中CD34和血管性假血友病因子（vWF）相关抗原的表达变化.结果 流式细胞仪检测ADSCs的CD49d和CD106阳性率分别为（98.32±0.37）%和（1.67±0.61）%;血管条件培养液诱导组细胞CD34和vWF阳性率分别为（77.14±0.76）%和（75.46±0.37）%,较空白对照组（1.38±0.31）%和（1.70±0.23）%均升高,差异有统计学意义（P〈0.01）.结论 ADSCs可以被诱导向ECs表型分化,提示ADSCs具有参与创伤后血管形成的潜能,可以应用于创伤修复的治疗.

Objective To investigate the possibility of adipose derived stem cells （ ADSCs） for wound healing by detecting cellular phenotype conversion of ADSCs into endothelial cells （ECs）. Methods ADSCs were isolated and cultured from adipose tissue derived from SD rats （n=8）, and maintained in Dulbecco＇s modified Eagle＇s medium （DMEM） containing 10% fetal bovine serum （FBS） in vitro. The marker antigen of P3 ADSCs was detected by analysis CD49d and CD106 antigens expression using flow cytometry, and the multipotential differentiation of P3 ADSCs were identified by specific medium inducing to differentiate into osteoblasts and adipocytes. And then, the ADSCs were cultured and induced for 3 days by condition culture medium （containing 30% superior of homogenating rat blood vessels in 10%FBS DMEM） as experimental group, and were cultured by 10% FBS DMEM as control group, and the expression of CD34 and von Willebrand factor （vWF） in ADSCs were analyzed by flow cytometry. Results Flow cytometry analysis showed that the expression of CD49d and CD106 in ADSCs were positive（98.32 ±0.37）% and negative（1.67 ±0.61）% , respectively. The multipotential differentiation experiment demonstrated that the cultured P3 ADSCs can be induced to differentiate into osteoblasts and adipocytes in vitro. The positive rate of CD34 and vWF were （77.14 ± 0.76） % and （75.46 ± 0.37） % in condition medium group, higher than （1.38 ±0.31）% and （1.70±0.23）% in 10% FBS DMEM control group, respectively （P〈0.01）. Conclusion The ADSCs can be induced to differentiated into ECs, suggesting that ADSCs have potential to take part in wound repair and angiogenesis.