Comparative Neural Differentiation of Orofacial and Iliac Marrow Stromal Cells

Objectives: Multipotent properties of orofacial (maxilla/mandible) bone marrow stromal cells (OFMSCs) make them ideal donor tissues for craniofacial stem cell-based therapies. Although OFMSCs arise embryologically from neural crest, they display more superior osteogenic properties than iliac crest bone marrow stromal cells (ICMSCs) that arise from mesoderm. Since skeletal site-specific neural differentiation of bone marrow stromal cells is unclear, we hypothesized that neural crest-derived OFMSCs can readily differentiate into neural tissues and are superior donor cells for neural tissue regeneration than ICMSCs.

Methods: Using an institutional review board-approved protocol, multicolony human OFMSCs and ICMSCs from 25 year-old female volunteer were expanded in a-MEM (Minimum Essential Medium) supplemented with 100 U/ml penicillin, 100 mg/ml streptomycin, 2 mM glutamine and 20% fetal calf serum. Early passage cells sub-cultured in poly-D-lysine coated dishes were pre-induced with10 ng/ml bFGF for 24 hours followed by 12 hour culture in neural induction medium composed of a-MEM, 2% DMSO, 10 ng/ml bFGF, 100 mM butylated hydroxylanisole, 10 mM forskolin, 25 mM KCl, 2 nM valproic acid and 5 mg/ml insulin. Parallel control cells were not induced. Markers of neural differentiation were assessed by immunostaining with antibodies to SOX-2, nestin, musashi, b3-tubulin, glial fibrilar acidic protein (GFAP), and oligodendrocyte (O-1). After 24hr neural induction, mRNA was isolated to assess gene transcripts of musashi and b3-tubulin

Results: Relative to control, SOX-2 and GFAP were undetectable in all three cell types after neural induction. Immunostaining of early (nestin, musashi) and late (b3- tubulin, O-1) neural markers were comparatively (2-3x) higher in OFMSCs than ICMSCs. OFMSCs gene transcripts of musashi and b3-tubulin were triple and double those of ICMSCs respectively.

Conclusions: OFMSCs can differentiate much more readily to neural tissue than ICMSCs. OFMSCs are potentially good donor cells for neural tissue regeneration compared to ICMSCs. (Supported by DHHS/NIH/NCI grant 5K08CA120875-02)