STUDY OF CHONDROGENIC EFFECTS OF CHONDROCYTES COCULTURED WITH MURINE BONE MARROW-DERIVED MESENCHYMAL STEM CELLS

Co-culture systems of marrow derived mesenchymal stem cells (mMSCs) with mature chondrocytes have theoretically been considered as a putative way of MSCs chondrogenic differentiation. MSCs differentiated in this system could be used for transplantation purpose without of any need to their purification since the cells with which MSCs are co cultured are native cartilage cells. Despite of the importance of this system the data is very rare in this context which is subject of the present study. The mesenchymal stem cells were isolated from the bone marrow of 4-6 weeks old NMRI mice by low-density primary culture system and expanded by performing two successive subcultures. Chondrocytes were obtained from rat costal cartilage and cultivated in 25-cm2 flasks for a period of two weeks. To prepare co culture systems passaged-2 mesenchymal cells and primary-cultured chondrocytes were used. In present study the different cell ratios of MSCs and chondrocytes including 1/1 1/2 and 2/1 were prepared and cultivated in a micro mass culture systems in a medium lacking the factors known to induce cartilage differentiation. In the end of cultivation periods (21 days) the groups were examined by toluidine blue staining technique and RT-PCR analysis for murine cartilage differentiation. Spindly–shaped murine mesenchymal cells were dominated the culture by second subculture. In the case of rat chondrocyte these cells were reached to confluency 2 weeks after culture initiation. Co culture results indicated that in the group of 1/2 ratio (when the number of chondrocyte were as twice as MSCs) the abundant of metachromatic matrix containing collagen II X and aggreacan were produced whereas in other groups this matrix were rarely observed. Chondrocyte co cultured with mesenchymal stem cells seemed to practically produce chondrogenic signal enough to promote intensive MSCs cartilage differentiation. In this system the ratio of the co cultured cells is important.