Analysis of bone remodeling in a mouse model for craniometaphyseal dysplasia (CMD)

Date of Completion

January 2009


Biology, Molecular




Mutations in the pyrophosphate (PPi) transporter ANKH cause the autosomal dominant form of craniometaphyseal dysplasia (CMD), a rare disorder characterized by hyperostosis of craniofacial bones and widened metaphyses in long bones. We have generated a knock-in (KI) mouse model expressing a deletion mutation (Phe377del) in the Ank gene, the most common mutation found in our CMD patients. AnkKI/KI mice replicate many CMD features including increased radiopacity of craniofacial bones; undertrabeculated metaphyses in club-shaped femurs; narrowing of cranial neural foramina; fusion of middle ear bones. In vivo skeletal analyses show that AnkKI/KI mice have increased osteoblast and ostseoclast numbers, increased bone turnover serum markers, however, no changes in bone formation and mineral apposition rate. Moreover, femoral bones from AnkKI/KI mice are hypomineralized. Consistently, AnkKI/KI osteoblast cultures show significantly reduced mineral nodules. The ANK Phe377del mutation does not affect expression of Tnap and Enpp1 nor TNAP activity in osteoblasts. However, PC-1 activity is increased which is likely a compensation for the dysfunctional mutant ANK to maintain the extracellular PPi level unchanged. Expression of Mmp13, Ocn, Runx2, Osx, Phex and Fgf23 genes are decreased in AnkKI/KI late osteoblasts and bones. Systemically, AnkKI/KI mice display age-dependent hypophosphatemia, hypocalcemia and normal PTH levels. We used bone marrow-derived macrophage (BMM) cultures for studying osteoclast phenotypes in vitro. AnkKI/KI BMMs formed significantly fewer TRAP + cells (nuclei ≥3) and showed decreased mineral resorption. Cell autonomous defects in AnkKI/KI osteoclasts led to delayed expression of DC-Stamp, decreased Adam8 mRNA and to disrupted actin ring formation. Moreover, less AnkKI/KI osteoclast progenitors participate in osteoclastogenesis and increased AnkKI/KI osteoclast numbers in vivo are likely to compensate for reduced functionality. From calvarial osteoblast-BMM cocultures, we conclude that Ank KI/KI osteoblasts fail to fully support Ank KI/KI osteoclastogenesis. By cross-transplantation of 4.5-day-old femurs into back muscles of 4-week-old mice, we show that the major contributor to the CMD-like phenotype is intrinsic to transplanted bone rather than to systemic regulation by the host. Finally, preliminary data from bone marrow transplantation experiments suggest that reduced function of osteoclasts is in part responsible for abnormal bone mass. We believe that further studies of the AnkKI/KI model could potentially reveal new therapeutic approaches for treating CMD. ^