Intracellular Tension of Osteoblasts in Collagen Matrix Under Uniaxially-Fixed Boundary Condition Elicits Osteocyte Alignment with Induction of Osteogenesis

Osteocytes differentiated from osteoblasts play a significant role as mechanosensor in modulating the bone remodeling process. While the well-aligned osteocyte network along the trabeculae is known to facilitate the sensing of mechanical stimuli by cells and the intracellular communication in the bone matrix, understanding these mechanisms in the osteocyte network formation remain poor. Here, we developed a novel in vitro collagen system exerting a uniaxially-fixed boundary condition on which mouse osteoblast-like MC3T3-E1 cells were subcultured for 1 d, evoking cellular alignment along the uniaxial boundary condition. By using a myosin II inhibitor, blebbistatin, we showed that the intracellular tension was strengthened by stabilization of actin filaments along the uniaxially-fixed boundary condition, so that this positive tension-feedback under isometric contraction contributed to the enhanced intracellular tension. After 10 d incubation, cells actively migrated inside the collagen and exerted the promoted expression of osteoblast and osteocyte genes compared with 4 d incubation. Notably, the migrated cells became aligned along the boundary condition after 10 d cultivation, reconstructing the in vivo process of formation of osteocyte network. Collectively, we suggested that the intracellular tension of osteoblasts under uniaxially-fixed boundary condition plays a significant role to determine the osteocyte alignment inside the bone matrix.