中文摘要：

目的：通过体外分离培养脂肪干细胞,并进行传代、表型鉴定及多向诱导分化,进一步了解其生物学特性,为临床运用奠定实验基础。方法：取SD大鼠附睾近心端,肾周及腹股沟韧带皮下脂肪组织,采用密度梯度离心和贴壁培养相结合方法,对脂肪干细胞进行分离、体外培养及扩增、表型鉴定及成骨成脂诱导分化,在倒置显微镜下观察原代及传代细胞形态变化,并采用MTT法检测第3、5、7、9代细胞增殖能力,用流式细胞仪检测细胞表面抗原标记,取第3代细胞进行成骨成脂诱导分化,并采用茜素红染色、油红O染色方法进行鉴定。结果：采用密度梯度离心和贴壁培养相结合方法得到的脂肪干细胞,其细胞形态较为均一,基本呈长梭形,旋涡状生长,并形成集落样生长。MTT法检测第3、5、7、9代ADSCs呈典型的“S”形增殖曲线,符合干细胞的特性,脂肪干细胞表型鉴定结果显示CD29阳性高表达,CD34阴性表达,CD45低表达,CD105弱阳性表达,脂肪干细胞能向成骨细胞及脂肪细胞分化。结论：脂肪干细胞取材较方便,含量丰富、并能够在体外分离培养,其稳定表达特异性的表面抗原,并且经过相应诱导培养后可向成骨及成脂多向分化,可作为优良的骨组织工程的种子细胞来源。

英文摘要：

Objective ： To investigate the method of isolation, culture and identification of adipose - derived stem cells （ADSCs） in vitro. Their biological characteristics were further understood so as to provide the reliable experimental basis for clinical therapy. Methods： ADSCs were purified by density gradient centrlfugation and adhesive culture methods. The adipose tissue was from proximal part of epididymis, kidney and subcutaneous adipose tissue of inguinal ligament, ADSCs were made isolation, characterization and multidirection differentiation. The primary and passage cells were observed under an inverted microscope. The third generation cells were used for osteogenic and adipogenic differentiation, which were identified by the methods of alizarin red staining and oil red O staining. Results ： The morphology of the ADSCs was uniform, with a long spindle and spiral shaped growth. The growth curve of ADSCs appeared as a typical ＇ S - shaped curve＇ ,which had high positive expression for CD29, negative expression for CD34, low expression for CD45 and weakly positive expression for CD105 and ADSCs can be induced differentiation into osteoblasts and adipocytes. Conclusions： ADSCs were isolated from adipose tissue by using density gradient eentrifugation and adhesive culture methods in rats. They were accorded with all the characteristics of stem cells, which provided a rich source of stem cells for the clinical application of stem cells.