The Study Of The Isolation, Cultivation And Differentiation To Chondrocyte Type Of Adipose Derived Stem Cells

ObjectiveTo investigate the methods of isolation, cultivation and identification of Adipose derived stem cells (ADSCs) and the feasibility of inducting ADSCs to differentiate to chondrocyte by monolayer culture with exogenous Transforming growth factor-β1 (TGF-β1).Materials and methodsThe adipose tissue was obtained from rats' inguinal area and adipose curshion of testis, and isolated, cultivated by enzyme digesting. The expression of CD29 was detected by immunofiuorescence; The passaged cells were exposed to two kinds of defined mediums for chrondrogenic phenotype differentiation. One medium was with TGF-β1, and the other was blank. After two weeks of induction, the identification of chondrogenic phenotype was detected by immunofiuorescence.Results1,The fearure of the ADSCs: The primary cultured ADSCs could adhere to the culture flask within the first 24 hours, which represented small sphere shape with high refraction ratio mostly. The term of nuclear splitting may occur after 3 days, and it may form the colony after 5 days, and get 90% confluence within 7 days. The offspring cells was basically fibroblast-shaped when cultred in vitro.2,The identifiction of ADSCs: The result of immunofiuorescence showed that surface antigen CD29 was positively expressed on ADSCs.3,The differention of ADSCs to chondrocyte: After chondrogenic induction, the growth velocity of the TGF-β1 group was slower than before. In the next day, we can see some cell became polygonal, and pseudopodia and black particles around the nucleus when observed in microscope. After 14 days chondrogenic induction, the cells became buninoid and aggregate into cell nodules.4,The detection of Collagen II by immunohistochemical method: TheTGF-β1 group shows positive expression but negative expression in the blank group.Conclusion1,The surface antigen CD29 was positively expressed on ADSCs.2,ADSCs can be isolated from rats' adipose tissue and their biological characteristics were stable in vitro.3,After 14 days cultivation with exogenous TGF-β, the detection of collagen II was positively expression; To confirm the differentiation ability of ADSCs to chondrocyte.