The primary goal of these studies is to increase the understanding of signaling pathways and regulatory molecules in cells that contribute to bone healing. Casitas b-lineage lymphoma (Cbl) is an E3 ubiquitin ligase as well as an adaptor protein. It can sequester, and thereby regulate the activity of Phosphoinositide3-kinase (PI3K), which has been shown to play a central role in cell survival, proliferation, and migration. The interaction between Cbl and PI3K has been shown to have effects on the numbers and functions of both osteoblasts and osteoclasts during normal bone homeostasis. Animals in which the Cbl-PI3K interaction is abrogated have increased osteoblast numbers and more bone formation, and increased osteoclast numbers with decreased resorptive capacity, ultimately yielding a net gain of bone volume in adulthood. To investigate whether or not the Cbl-PI3K interaction played a role in bone healing, we proposed to study bone formation during fracture healing in animals lacking this interaction. We hypothesized that abrogation of the Cbl-PI3K interaction alters the osteoprogenitor response to fracture, resulting in more bone formation and enhanced fracture healing. We first found that the lack of Cbl-PI3K interaction resulted in an enhanced proliferative response in periosteal cells adjacent to the fracture site. Furthermore, we demonstrated that this proliferative population of cells had increased expression of Osterix, a master regulator of the osteoblastic lineage, and that they had an increased capacity to differentiate towards osteoblasts compared to controls in vitro. Next, we identified a polarizing effect of Cbl-PI3K on fracture healing. Lack of the interaction resulted in increased numbers of osteoblasts, increased bone deposition, and increased bone volume within the fracture callus. At the same time, we observed increased numbers of osteoclasts, but a delay in remodeling of the soft callus to hard callus, and a delay in resorption of the bony callus. Ultimately, the net effect was a delay in bone healing in animals lacking the Cbl-PI3K interaction. Taken together, these results suggest that the Cbl-PI3K interaction regulates bone repair, and loss of the interaction leads to increased bone formation, but impaired bone remodeling, resulting in delayed healing.
