Fibroblast and osteoblast differentiation characteristics of periosteum-derived cells in mouse calvaria

Abstract

Objective: Periosteum is considered to be involved in maintenance and regeneration of the bone and gingiva. Here, we investigated whether periosteum-derived cells (PDCs) display the ability of gingival fibroblast differentiation and osteoblast differentiation, and examined their characteristics of fibrogenesis compared with that of bone marrow cells (BMCs). Methods: PDCs and BMCs were prepared from skull periosteum and bone marrow of femurs in 4-week-old female ICR mice and cultured in growth medium (D) supplemented with fibrogenic (F) or osteogeneic (O) medium for 3 weeks, followed by additional subtreatments with F or O for 2 weeks. Cells were also cultured with SIS3, a Smad3 phosphorylation inhibitor, followed by TGF-β treatment. Gene and protein expressions related to fibrogenic and osteogenic differentiation were evaluated with RT-PCR and western blot and immunostaining, respectively. Results: Both cells showed ALP activity after osteogenic induction, with PDCs showing increased ALP activity by sub-treatment of D-O and F-O, but not with BMCs. PDCs showed higher gene expression of Runx2, Fgfr1, and α-SMA compared with BMCs in osteogenic culture and after sub-treatment with F. Furthermore, Fgfr1 gene expression was stronger in PDCs than in BMCs after inhibiting Smad3 phosphorylation. Notably, TGF-β treatment stimulated or recovered the expression of S100A4 and vimentin, even after inhibition of Smad3 phosphorylation in BMCs, but not in PDCs. Conclusion: PDCs might have potential of both osteogenic and fibrogenic differentiation. Furthermore, TGF-β signaling could occur without Smad3 phosphorylation in PDCs, indicating presence of a non-canonical signal transcription factor in PDCs.