Roles of the Phosphate-regulating Proteins, PHEX and DMP1, in the Dentin Matrix of XLH

X-linked hypophosphatemia (XLH) is the most common hereditary hypophosphatemic disorder resulting from loss-of-function of the phosphate-regulating endopeptidase, PHEX. Although the mineralization defects observed in XLH patients have been solely attributed to decreased serum phosphate levels, the local disruption in the organization of the dentin matrix that precedes and sustains mineralization has not been studied. Nevertheless, disruption of extracellular matrix (ECM) factors necessary for the maturation and mineralization of cartilage, bone and cementum have been reported, suggesting that PHEX mutations and/or hypophosphatemia may indeed disrupt ECM deposition. The studies herein were undertaken to elucidate the local effects of PHEX mutations on the deposition and mineralization of dentin matrix. By manipulating in vitro conditions, we evaluated the effect of phosphate donors on the differentiation of XLH dental pulp cells. We found that XLH dental pulp cells are sensitive to the phosphate source used during in vitro differentiation experiments and that XLH cells have an impaired differentiation response when compared to healthy control cells. By generating cell lines with constitutive DMP1 expression we showed that full-length DMP1(FL-DMP1) was able to reverse the impaired mineralization in XLH cells. Furthermore, induction of differentiation resulted in the upregulation of inhibitors of the canonical Wnt pathway, altered ECM protein processing and elevated levels of MMP2 and MMP3 in XLH cells. DMP1 expression was sufficient to reverse these patterns. We generated a tooth-specific DMP1-overexpressing PHEX-deficient mouse. DMP1 expression resulted in decreased Col.I protein expression in the predentin and periodontal ligament space relative to their wild-type(WT) and XLH/Hyp counterparts. Although DMP1 expression was associated with decreased WNT5A expression in DMP1-overxpressing mice, DMP1 was unable to effect the same response in the Phex-deficient background. Likewise, DMP1 overexpression failed to rescue the dentin phenotype of Hyp mice. In summary, our studies suggest that deregulation of the differentiation process may be occurring in XLH cells. DMP1 has the potential to modulate these differentiation pathways but requires a functional PHEX protein to function. These findings suggest that PHEX and DMP1 may interact and/or that DMP1 may function upstream of PHEX in many of these pathways.