The Role of Rac and Filamin A in Osteoclast Function and Osteoclast Heterogeneity

Osteoclasts are multinucleated cells that resorb bone and carry out their function via a dynamic actin cytoskeleton that is regulated by Rho GTPases. Rho GTPases function as molecular switches that regulate the actin cytoskeleton in osteoclasts and many other cell types. A Rac GTPase validation experiment was used to prove that osteoclast function is best characterized through three-dimensional (3-D) volumetric analysis of resorption pits (Rpits). The success of this method prompted us to measure Rpits in osteoclasts lacking actin binding protein Filamin A (FLNA), which interacts with Rho GTPases to regulate osteoclast formation (osteoclastogenesis (OCG)) and function. It was observed using this technique that FLNA-knock-out (KO) osteoclasts were smaller and produced shallower Rpits compared to WT osteoclasts. Using an osteoporosis model as well as in vivo techniques measuring bone material, structural and mechanical properties, it was demonstrated that FLNA knockout (KO) bones were protected from post-menopausal osteoporosis associated with estrogen depletion. FLNA-KO cortical bones were also found to be more ductile than WT cortical bones. The discrepancy between bone behavior in vertebrae vs. femora in FLNA mice triggered the question as to whether osteoclasts behave differently depending on the skeletal site from which they originated. The osteoporosis model, resorption pit method and aforementioned in vivo techniques with the exception of bone mechanical properties were used to determine whether osteoclasts and bone turnover differed between the cranial and appendicular skeletons. Calvarial bone displayed higher turnover rates compared to long bone and mandibular bone. The novel in vitro 3-D method, characterization of FLNA in vivo and the assessment of osteoclasts arising from different bone locations were executed to provide more insight into the development of pharmacotherapeutics to better manage pathological bone loss.Ph.D