IN-VITRO CHONDROGENESIS OF BONE-MARROW-DERIVED MESENCHYMAL PROGENITORCELLS

A culture system that facilitates the chondrogenic differentiation of rabbit bone marrow-derived mesenchymal progenitor cells has been devel oped. Cells obtained in bone marrow aspirates were first isolated by m onolayer culture and then transferred into tubes and allowed to form t hree-dimensional aggregates in a chemically defined medium. The inclus ion of 10(-7) M dexamethasone in the medium induced chondrogenic diffe rentiation of cells within the aggregate as evidenced by the appearanc e of toluidine blue metachromasia and the immunohistochemical detectio n of type II collagen as early as 7 days after beginning three-dimensi onal culture. After 21 days, the matrix of the entire aggregate contai ned type II collagen. By 14 days of culture, there was also evidence f or type X collagen present in the matrix and the cells morphologically resembled hypertrophic chondrocytes. However, chondrogenic differenti ation was achieved in only approximately 25% of the marrow cell prepar ations used. In contrast, with the addition of transforming growth fac tor-beta 1 (TGF-beta 1), chondrogenesis was induced in all marrow cell preparations, with or without the presence of 10(-7) M dexamethasone. The induction of chondrogenesis was accompanied by an increase in the alkaline phosphatase activity of the aggregated cells, The results of RT-PCR experiments indicated that both type IIA and IIB collagen mRNA s were detected by 7 days postaggregation as was mRNA for type X colla gen. Conversely, the expression of the type I collagen mRNA was detect ed in the preaggregate cells but was no longer detectable at 7 days af ter aggregation, These results provide histological, immunohistochemic al, and molecular evidence for the in vitro chondrogenic differentiati on of adult mammalian progenitor cells derived from bone marrow. (C) 1 998 Academic Press.