title: N-terminal Dentin Sialoprotein fragment induces type I collagen production and upregulates dentinogenesis marker expression in osteoblasts.
abstract: Bone and dentin are mineralized extracellular matrices produced by osteoblasts and odontoblasts, respectively, and their major organic portion is type I collagen. Dentinogenesis Imperfecta (DGI) is one of the most common clinically- and genetically-based disturbances of dentin formation, causing irreversible dentin defects. Among several types of DGI, patients with DGI type II exhibit opalescent dentin with partial or complete pulp obliteration. It has been previously reported that the non-sense mutation (c.133C>T) inDentin Sialophosphoprotein(DSPP) was identified in DGI type II patients at glutamine residue 45, resulting in the premature stop codon (p.Q45X). DSPP is known to be synthesized as a single gene product and further processed at Gly462-Asp463, resulting in the production of Dentin Sialoprotein (DSP) and Dentin Phosphoprotein (DPP). We hypothesized that the shorter form (Q45X) of N-terminal Dentin Sialoprotein (N-DSP) may cause over-production of type I collagen protein as obliterated pulp is occupied by dentin. To test this hypothesis, we generated mouse recombinant Glutathione-S-Transferase (GST)-N-DSP fusion protein, and the effect of GST-N-DSP was investigated in calvarial bone explant culture and MC3T3-E1 osteoblastic culture systems. Here we show that a significant increase in calvarial bone formation is observed by GST-N-DSP. GST-N-DSP accelerates MC3T3-E1 osteoblast cell growth and proliferation and subsequent osteoblast differentiation by inducing the expression of certain osteogenic markers such astype I collagen,Runx2,OsterixandATF4. Interestingly, GST-N-DSP significantly enhances dentinogenesis marker gene expression includingDsppandDmp1gene expression in non-odontogenic MC3T3-E1 cells. To rule out any artificial effect of GST-tag, we also used the synthetic peptide of N-DSP and confirmed the results of N-DSP peptide were essentially similar to those of GST-N-DSP. Taken together, our data suggest that N-DSP promotes bone formation by accelerating osteoblast cell proliferation and subsequent osteoblast differentiation accompanied by marked up-regulation of the dentin matrix markers, such asDsppandDmp1genes.
authors.affiliation: Department of Molecular and Cell Biology, Boston University, Henry M. Goldman School of Dental Medicine, Boston, MA 02118, USA.
authors.forename: Haytham
authors.initials: H
authors.lastname: Jaha
authors.affiliation: Department of Molecular and Cell Biology, Boston University, Henry M. Goldman School of Dental Medicine, Boston, MA 02118, USA.
authors.forename: Dina
authors.initials: D
authors.lastname: Husein
authors.affiliation: Department of Molecular and Cell Biology, Boston University, Henry M. Goldman School of Dental Medicine, Boston, MA 02118, USA.
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Ohyama
authors.affiliation: Department of Molecular and Cell Biology, Boston University, Henry M. Goldman School of Dental Medicine, Boston, MA 02118, USA; Henan Province People 's Hospital, 7 Weiwu Rd, Zhengzhou, 450003, China.
authors.forename: Dongliang
authors.initials: D
authors.lastname: Xu
authors.affiliation: Department of Biological Endodontics, Integrated Health Sciences, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, 734-8553, Japan.
authors.forename: Shigeki
authors.initials: S
authors.lastname: Suzuki
authors.affiliation: Department of Bioscience Research, College of Dentistry, University of Tennessee Health Science Center, Memphis, TN 38163, USA.
authors.forename: George T-J
authors.initials: GT
authors.lastname: Huang
authors.affiliation: Department of Molecular and Cell Biology, Boston University, Henry M. Goldman School of Dental Medicine, Boston, MA 02118, USA.
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Mochida
keywords: Dentin Sialophosphoprotein (DSPP); Dentin Sialoprotein (DSP); Dentinogenesis Imperfecta; MC3T3-E1 cells; type I collagen