Refining Retinoic Acid Stimulation for Osteogenic Differentiation of Murine Adipose-Derived Adult Stromal Cells

Abstract

Murine adipose-derived adult stromal cells (ADAS) seeded onto appropriate scaffolds and pre-incubated with retinoic acid have been shown to generate in vivo bone rapidly. Prompt resorption ensues, however, as a result of osteoclastogenesis, likely secondary to retinoic acid carryover. In this study, we determined the effects of abbreviated retinoic acid exposure on ADAS osteogenic differentiation. Histological staining and gene expression analysis revealed that longer retinoic acid exposure resulted in better in vitro bone differentiation. However, significant osteogenesis was observed in ADAS after just 15 days of retinoic acid supplementation, suggesting that continual culture with retinoic acid is unnecessary for initiation of the osteogenic program. This was confirmed using ADAS pre-incubated in monolayer with an abbreviated 15 days of retinoic acid exposure before implantation into critical-sized calvarial defects. Similar rates of regeneration were observed between ADAS exposed to for 15 days or for a full 25-day course of retinoic acid before defect repair. Furthermore, by limiting retinoic acid exposure to ADAS in monolayer without scaffold, accelerated bone formation was observed without concomitant osteoclastic resorption. These data suggest that skeletal regeneration may be improved by modulating retinoic acid exposure before implantation, markedly accelerating the repair of bone defects using ADAS.