DESCRIPTION (provided by applicant): Long bone growth in children is dependent on the constant and consistent proliferation and differentiation of chondrocytes at the growth plate. The orderly pattern of growth is regulated by a long list of both systemic and locally derived factors, including growth hormone (GH) and insulin-like growth factor-l (IGF-I). We have isolated chondrocytes from juvenile bovine metacarpals because we believe the bovine growth plate physiology approximates the human better than rodent models. The chondrocytes have been separated by density gradient centrifugation into fractions that correspond to the recognized zones of the growth plate, based on cell size, glycogen content and expression patterns of genes known to be differentially expressed in histologic growth plate zones (collagen X, alkaline phosphatase, Indian hedgehog, and IGFBP5). In short term primary cultures, all fractions exhibited significant baseline proliferative activity, but no response to GH. Also, IGF-I levels in bovine chondrocytes are extremely low, and are not increased by GH. All fractions responded to IGF-I with proliferation, and the degree of proliferation correlated with cell density and IGFBP5 expression; the fully differentiated hypertrophic cells responded the least to IGF-I. In addition, while in the high density cells IGF-I readily activated MAPK, IGF-I did not increase MAPK activity in the hypertrophic cells, suggesting that IGF-I had become de-coupled from MAPK with progressive differentiation. In other cell models, IGF-I has been proposed to provide divergent signals for both proliferation and differentiation, with the particular cellular response determined by the relative amounts of signaling molecules downstream of the IGF-I receptor. The overall aim of this project is to support our claim that GH has little direct effect at the growth plate, that systemic IGF-I is the main signal for proliferation, differentiation and hypertrophy in growth plate chondrocytes, and that the different signals mediated by IGF-I are managed by the relative levels of signaling molecules downstream of IGF-IR within the chondrocyte fractions. The relevance to public health is to better understand mechanisms regulating bone growth in children, by using cells from young non-rodent animals for in vitro studies.