Skeletal unloading results in a cessation in bone formation and an initial increase in bone resorption. These changes are accompanied by resistance to the anabolic actions of insulin like growth factor 1 (IGF1) on bone. On the other hand skeletal loading increases IGF1 production, and enhances IGF1 activation of IGF1R as part of the mechanism by which it increases bone formation. In seeking a mechanism for the load induced regulation of IGF1 signaling we discovered that skeletal unloading was associated with a decrease in the expression of integrins, in particular integrins with b1 (IGTB1) and b3 (ITGB3) subunits. We discovered that in osteoblasts, IGF1 increased the binding of ITGB3 to IGF1R, and that if ITGB3 were downregulated, IGF1 could no longer activate IGF1R. Focal adhesion kinase (FAK) and/or its related family member protein tyrosine kinase 2 beta (PTK2B) provide a link between the integrin and growth factor receptor pathways including between ITGB3 and IGF1R. IGF1 activation of IGF1R results in phosphorylation (presumed activation) of FAK, whereas inhibition of FAK blocks activation of IGF1R either by IGF1 or by load. To test the role of IGF1 signaling directly in vivo we developed mice in which the IGF1R was deleted in mature osteoblasts and examined whether this mouse would respond to skeletal unloading or reloading. Mice lacking IGF1R showed an equivalent decrease in bone formation during unloading to controls, but failed to increase bone formation during reloading. The striking result, however, was that this failure to respond to reloading was found only in periosteal bone formation; endosteal and trabecular bone formation responded comparable to controls. These results focused our attention on the periosteum where the osteoprogenitors are exposed not only to load induced IGF1 emanating from both osteocytes and muscle, but to the integrin ligand periostin, a combination we hypothesize will maximize their proliferation, differentiation, and formation of new bone in response to mechanical load. In this project we will test the hypothesis that skeletal loading stimulates IGF1 production in osteocytes and muscle and formation of the IGF1R/integrin complex in periosteal osteoprogenitors (pOP) required for the IGF1 mediated anabolic response of these cells to load. This will be achieved in the following three aims:
Aim1 --Determine the components of the IGF1R complex that forms in response to load and/or IGF1 in periosteal cells in vitro, and assess their role in contributing to that response;
Aim 2 -- Determin the impact of deleting Igf1r and Itgb3 from pOP in vivo with respect to their ability to mediate th skeletal response to load;
Aim 3 --Determine the source(s) of IGF1 facilitating load induced periosteal bone formation with particular attention to osteocytes and muscle. We will be utilizing novel animal models and state of the art techniques to fulfill these aims. The results will provide new understanding of how IGF1/integrin signaling interactions regulate the skeletal response to mechanical load, open up new avenues for further investigation, and provide potential targets for preventing the bone loss of immobilization and aging.
The mechanisms underlying periosteal bone formation remain unclear. We have found that the deletion of IGF1 from mature osteoblasts/osteocytes disrupts periosteal bone formation in response to load but not that of trabecular or endosteal bone formation, a response involving integrins. Using cell specific deletion of the IGF1 receptor (IGF1R) and ITGB3, we plan to examine the contribution IGF1 and integrins make to mechanism by which periosteal cells respond to load.
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