A Search for the Osteogenic Factor in Dentin. Rat Incisor Dentin Contains a Factor Stimulating Rat Muscle Cells in Vitro to Incorporate Sulfate into An Altered Proteoglycan

Demineralized dentin matrix has the capacity to induce bone formation via a chondrogenic pathway when implanted into muscle, in a fashion entirely analogous to bone matrix implants. In this work we have attempted to isolate, from rat incisor dentin, the matrix factor responsible for initiating osteogenesis. Rat incisor dentin was demineralized with EDTA plus 4.0 M guanidine. HCl. The proteins in the extracts were collected and, after a CaCl2 precipitation step, fractionated on Sephacryl S-200 in 6.0 M guanidine. HCl. The primary assay for activity was the incorporation of 35S-sulfate into proteoglycan in cultures of the fibroblast-like outgrowth cells from explants of neonatal rat muscle. Two Sephacryl S-200 fractions showed enhanced sulfate incorporating activity, but only one showed enhanced incorporation without a concomitant increase in cell number. In the presence of this fraction, the cell cultures produced a larger amount of a new small proteoglycan, as compared to controls, and a significant amount of a much larger proteoglycan. The active fraction had proteins in the Mr range from 8,000 to 15,000 as the major components. These data suggest that the fraction identified may contain the factors responsible for initiating the osteogenic response to dentin matrix upon its implantation in muscle in vivo.