Novel explant culture model for evaluation of oxygen signaling during mammalian digit regeneration

Abstract

One in 200 people in the U.S. is a survivor of a limb amputation. The ideal replacement for an amputated limb would be the regrowth of the patient's own limb. Mammals have the prominent ability to regenerate the digit tip (P3) including bone, surrounding connective tissue and epithelia. Bone fracture studies show that fluctuations in oxygen tension can trigger bone repair cascades such as osteoblast differentiation and osteoclast activation. Therefore, we hypothesize that changes in oxygen tension will facilitate the regenerative process of the digit tip. Because the ability to both control and measure oxygen tension in vivo presents an impediment to studying the influence of oxygen on a cellular level, the objective of this study was first to develop a reliable explant culture model that would simulate P3 regenerative bone formation, and second to evaluate the influence of oxygen on bone regeneration with this model. Characterization of the explant model revealed a stable population of active osteoblasts up to two weeks in culture. Our studies also reveal that a temporal hypoxic event prior to bone mineralization and an increase in oxygen tension following this hypoxic event can promote bone regeneration in our explant slice model. These findings show the oxygen profile of the microenvironment during regeneration is dynamic and that oxygen can be influential in enhancing bone regeneration.