1996•Zhonghua guke zazhiRequires access
Culture of adipose derived stem cells from Sprague-Dawley rat and its osteochondrogenic differentiation potential in vitro
Yang Hui-lin, Geng De-chun, Wang Junhua
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Abstract
Objective To study the method of isolating and culturing stem cells from rat adipose tissue and to determine if adipose-derived stem cells(ADSCs)harvest from rat could differentiate into osteogenic and chondrogenic in vitro.Methods ADSCs were isolated from rat inguinal fat pads after extensive washing with phosphate-derived saline and digesting with collagenase.After primary culture in control medium and expanded to two passages,the cells were incubated in either an osteogenic medium or an chondrogenic medium for 2-4 weeks to induce osteogenesis and chondrogenesis,respectively.Evidences of Osteogenic differentiation,were detected by a ALP solution,and while chondrogenic differentiation was confirmed using the histologic Alcian blue staining at acidic pH.Expressions of osteocyte and chondrocyte specific genes were confirmed by RT-PCR.Results ADSCs can be isolated from rat adipose tissue and,expanded rapidly.It exhibited a heterogeneous population of fibroblast like cells morphologically.ADSCs induced to osteogenesis were stained positively for alkaline phosphatase activity after 2 weeks and formed mineralized nodular structures,as conformed by Von kossa staining.Expression of osteocyte specific genes,such as ALP,osteopontin,osteocalcin,was detected.ALP and osteopontin,were expressed constitutively in osteogenic medium after 2 and 4 weeks of culture.Expression of osteocalcin,was induced by osteogenic growth factors at 4 weeks.Induced to chondrogenesis cells were positive of Alcian blue staining under acidic conditions and expression of aggrecan and typeⅡ/Ⅹ collagen genes.Aggrecan and typeⅡcollagen genes were abundant after 2 weeks in chondrogenic medium.TypeⅩ collagen gene was detected at 4 weeks.Conclusion Adipose-derived stem cells can be isolated from rat adipose tissue.Their biological characteristics are similar with mesenchymal stem cells(MSCs),and have the potential to differentiate into osteogenic and chondrogenic lineage.It may be an idea source of ADSCs for tissue engineering.