Cartilage differentiation is a complex, incompletely understood process. A culture system developed in our laboratory facilitates the chondrogenic differentiation of postnatal mammalian bone marrow-derived mesenchymal progenitor cells. We hypothesize that this in vitro process recapitulates that which occurs in vivo and that it can be used to define the molecular events that are important in the process of chondrogenesis. The first specific aim is to examine the cell biology of the system; including the number and type of cells that take part in the process, quantification of the proliferation and cell death that occurs and the commitment of cells in the later stages of the process. The second specific aim is to define the temporal sequence for the expression of specific extracellular matrix molecules produced during differentiation, as a means to staging the process. Most studies of skeletogenesis in vitro involve the addition of cytokines as a means to defining their functions. We hypothesize that by identifying the cytokines that are intrinsic to the cells in the culture system, and defining the sequence of their up- or down- regulation, we will be able to more accurately define the functions of the cytokines in chondrocyte differentiation. Therefore, the third specific aim is to define the temporal sequence of endogenous cytokine induction and repression during the process of chondrogenesis. The use of targeted RNA fingerprinting will allow screening and identification of specific factors which are within cytokine families known to be involved in chondrogenesis. The information gained may then be used for applications such as developing strategies for manipulating marrow mesenchymal cells to facilitate cartilage repair. The system may also be used for examining the effects of human genetic mutations in molecules that play a role in chondrogenesis.
