Tooth Defects and DSP/DSPP Processing Interference in MMP-9 Knock-out Mice

Dentin sialophosphoprotein (DSPP) is processed into dentin sialoprotein (DSP) and dentin phosphoprotein (DPP). DSP and DPP play unique roles in tooth development since DSP or DPP mutations are associated with various forms of dentinogenesis imperfecta. Previously, we found that DSP is cleaved by matrix metalloproteinase-9 (MMP-9) into NH2- and COOH-terminal fragments. Objective: To study the role of MMP-9 in tooth formation and DSP/DSPP processing. Methods: MMP-9 null mice were used in which exon 2 and intron 2 were replaced by a phaphoglycerate kinase-neomycin cassette. Histology, radiography, nuclear magnetic resonance (NMR) and immunohistochemistry were used to analyze the tooth and dentin phenotypes of MMP-9 null mice versus wild-type animals. Results: Microscopic and histological studies showed that in MMP-9 null mice the teeth displayed severe attrition, an increased number of reparative dentin sites and both the odontoblasts and ameloblasts showed abnormal morphology. In MMP-9 null mice, X-ray and micro-CT studies demonstrated that the dentin was thin with widened dental pulp chambers and lower mineral density. Further, NMR showed that teeth had larger porosity and pore size distributions, indicating a change not only in mineral content, but also abnormal dentin ECM architecture and bound water. Immunohistochemistry studies showed that fibronectin and osteopontin, known MMP-9 substrates, were expressed at higher levels in the predentin, odontoblasts and dental pulp cells in MMP-9 null teeth compared to controls. Also, DSP was highly expressed in MMP-9 knock-out teeth. Moreover, using anti-DSP NH2-terminal and anti-DSP COOH-terminal antibodies, we found that the NH2- and COOH-terminal DSP fragments cleaved by MMP-9 exist endogenously with different distribution patterns in teeth. The NH2-ternimal DSP fragment was predominantly located in odontoblasts with moderate levels in predentin whereas the COOH-terminal DSP fragment was present primarily in the ECM, dentin and predentin. Conclusions: MMP-9 is required for tooth development and DSP/DSPP processing into active forms.