人脂肪干细胞结合微载体在生物反应器中向软骨细胞分化

目的]探索在旋转生物反应器内,应用微载体技术快速扩增并向软骨分化人脂肪干细胞。方法]将人脂肪干细胞结合Cytodex3微载体在旋转的生物反应器(RCSS)内进行动态培养,应用倒置显微镜和扫描电镜对微载体表面的脂肪干细胞进行动态观察,并对收获的脂肪干细胞进行Safran in-O、tolu id ine b lue染色等组织化学染色及Ⅱ型胶原的免疫化学染色分析。结果]脂肪干细胞于24 h内贴附于Cytodex3微载体表面,细胞形态为短梭形,随时间的延长,贴附于微载体的细胞逐渐增多,到培养后期,细胞密度可达最初接种的19倍左右,在微载体上收获的细胞进行番红花O、阿利新蓝染色呈阳性,Ⅱ型胶原染色阳性,均强于对照组。结论]利用微载体细胞培养技术可简便快速地在体外扩增脂肪干细胞,并成功实现向软骨细胞分化。

In vitro chondrogenesis of human adipose derived stem cells with microcarrier in RCSS

Objective]To investigate the possibility of quick proliferation and differentiation of human adipose derived stem cells(ADSCs) into chondrocytes with microcarriers in rotary cell culture system(RCCS).Method]The ADSCs were obtained and then co-culture with microcarriers in the RCCS which contained chondrogenic culture media for 7 day. Growth of ADSCs on Cytodex3 microcarriers was observed dynamically under phase contrast microscope and SEM.The differentiation of ADSCs into chondrocyte was identified by histochemical stain including Safranin-O,tolidine blue and Immunocytochemical analysis for type II collagen.Result]The ADSCs attached rapidly to the surface of Cytodex3 microcarriers in 24h.Quick growth of these cells was observed after they fully spread onto the microcarriers.The density of ADSCs increased about 20 times in later stage of culture as compared with the initial density.Histochemical and Immunocytochemical analysis revealed that Safranin-O,tolidine blue and II collagen stain were obviously positive.Conclusion]Microcarrier culture of ADSCs can yield a large quantity of cells within a short time and a successful differentiation into chondrocyte at the same time.