Bone adapts to its mechanical environment so that its form follows function, and failure of normal bone adaptation is known to play a significant role in the etiology of metabolic bone diseases such as osteoporosis and osteopetrosis, bone loss in space flight, and in the failure of total joint replacements. Osteocytes are intrinsically three-dimensional (3D) bone cells that are encased in mineralized extracellular bone matrix and interconnected with each other as well as osteoblasts through numerous intercellular processes by gap junctions. Therefore, they are ideally situated to sense and respond to mechanical events that arise from normal physiological loading of bone. Indeed, two-dimensional (2D) in vitro culture studies have shown that osteocytes respond biochemically to a variety of mechanical stimuli such as fluid shear and deformation that arise from physiologic activity. To gain the greatest insights into bone mechanotransduction, it is critical to develop an in vitro system that can allow the formation of (1) controlled osteocytic networks in 2D and eventually in 3D, in a configuration that closely resembles that in vivo; (2) spatially controlled co-culture of osteocytic networks and osteoblasts; and finally (3) application of physiologic levels of pressure-driven canalicular flow. Our approach to this challenge will be to incorporate microfabrication techniques and self- assembled monolayers (SAM) to develop novel 2D and 3D co-culture systems of osteocytic networks and osteoblasts to investigate bone cell mechanotransduction. The goals of this study are to: (1) develop and use an in vitro 2D co-cultured micropattern of osteocytic networks and osteoblasts, and to apply single-cell compressive deformation using atomic force microscopy (AFM) or regional fluid flow to study mechanotransduction between these cells; and (2) develop and use a microfluidic 3D co-culture system of osteocytic networks and osteoblasts, and study osteocyte response to canalicular flow and its subsequent intercellular communication with osteoblasts. With the advent of microfabrication techniques and microfluidics, bone cell mechanotransduction can be investigated in vitro under conditions that are more physiologically relevant than previously possible. New insights gained from this research will contribute to our general understanding of the etiology of menopausal and microgravity or age-related osteoporosis, and may lead to therapeutic interventions aimed at the mitigation or treatment of these diseases, as well as improvement in total joint replacements. ? ? ?
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