Comparative sequential morphological analyses during in vitro chondrogenesis and osteogenesis of mesenchymal stem cells embedded in collagen gels.

Chondrogenesis and osteogenesis during fetal development and postnatal growth constitute one of the most interesting and complicated subjects in biology. In this study, bone marrow mesenchymal stem cells (MSCs) were embedded in collagen gel, cultured in chondrogenic or osteogenic medium, and compared morphologically during chondrogenic and osteogenic differentiation sequentially by light and electron microscopy and immunohistochemical examination. Before induction, the MSCs were dispersed and round in the collagen gel. At day 1, MSCs with a large number of short processes produced extracellular fibers whose immunoreactivity was positive for collagen type I. At day 3, the shape of MSCs changed from round to elongated. Gap junctions positive for connexin 43 were also observed. At day 7, remarkable morphological differences were first observed during chondrogenesis and osteogenesis. The shape of MSCs changed to polygonal without cell processes during chondrogenesis, while MSCs remained spindle shaped with long processes during osteogenesis. Concurrently, collagen type II during chondrogenesis and osteocalcin during osteogenesis were first detected. At day 21, chondrogenesis and osteogenesis of the MSC/collagen composite further progressed, respectively. In vitro chondrogenesis and osteogenesis using an MSC/collagen composite clarified the morphological differences.