Abstract
Osteocytes are the most abundant cells in bone yet are the most challenging to study because they are embedded in a mineralized matrix. We generated a clonal cell line called IDG-SW3 (for Immortomouse/Dmp1-GFP-SW3) from long-bone chips from mice carrying a Dmp1 promoter driving GFP crossed with the Immortomouse, which expresses a thermolabile SV40 large T antigen regulated by interferon γ (IFN-γ). Cells from these mice can be expanded at 33 °C in the presence of IFN-γ and then allowed to resume their original phenotype at 37 °C in the absence of IFN-γ. IDG-SW3 cells are Dmp1-GFP(-) and T antigen(+) under immortalizing conditions but Dmp1-GFP(+) and T antigen(-) under osteogenic conditions. Like osteoblasts, they express alkaline phosphatase and produce and mineralize a type 1 collagen matrix containing calcospherulites. Like early osteocytes, they express E11/gp38, Dmp1, MEPE, and Phex. Like late osteocytes, they develop a dendritic morphology and express SOST/sclerostin and fibroblast growth factor 23 (FGF-23), regulated by parathyroid hormone (PTH) and 1,25-dihydroxyvitamin D(3). When cultured on 3D matrices, they express Dmp1-GFP and sclerostin. When the 3D cultures are implanted in calvarial defects in vivo, they accelerate bone healing. This cell line should prove useful for studying osteoblast-to-osteocyte transition, mechanisms for biomineralization, osteocyte function, and regulation of SOST/sclerostin and FGF-23.