Activation of intracellular calcium signaling in osteoblasts colocalizes with the formation of post-yield diffuse microdamage in bone matrix

Abstract

Previous studies demonstrated that extracellular calcium efflux ([Ca(2+)]E) originates from the regions of bone extracellular matrix that are undergoing microdamage. Such [Ca(2+)]E is reported to induce the activation of intracellular calcium signaling ([Ca(2+)]I) in MC3T3-E1 cells. The current study investigated the association between microdamage and local activation of intracellular calcium signaling quantifiably in MC3T3-E1 cells. Cells were seeded on devitalized notched bovine bone samples to induce damage controllably within the field of observation. A sequential staining procedure was implemented to stain for intracellular calcium activation followed by staining for microdamage on the same sample. The increase in [Ca(2+)]I fluorescence in cells of mechanically loaded samples was greater than that of unloaded negative control cells. The results showed that more than 80% of the cells with increased [Ca(2+)]I fluorescence were located within the damage zone. In conclusion, the findings demonstrate that there are spatial proximity between diffuse microdamage induction and the activation of intracellular calcium ([Ca(2+)]I) signaling in MC3T3-E1 cells. The downstream responses to the observed activation in future research may help understand how bone cells repair microdamage.