title: Tissue-nonspecific alkaline phosphatase deficiency causes abnormal craniofacial bone development in the Alpl(-/-) mouse model of infantile hypophosphatasia.
abstract: Tissue-nonspecific alkaline phosphatase (TNAP) is an enzyme present on the surface of mineralizing cells and their derived matrix vesicles that promotes hydroxyapatite crystal growth. Hypophosphatasia (HPP) is an inborn-error-of-metabolism that, dependent upon age of onset, features rickets or osteomalacia due to loss-of function mutations in the gene (Alpl) encoding TNAP. Craniosynostosis is prevalent in infants with HPP and other forms of rachitic disease but how craniosynostosis develops in these disorders is unknown.
  
  
  OBJECTIVE
  Because craniosynostosis carries high morbidity, we are investigating craniofacial skeletal abnormalities in Alpl(-/-) mice to establish these mice as a model of HPP-associated craniosynostosis and determine mechanisms by which TNAP influences craniofacial skeletal development.
  
  
  METHODS
  Cranial bone, cranial suture and cranial base abnormalities were analyzed by micro-CT and histology. Craniofacial shape abnormalities were quantified using digital calipers. TNAP expression was suppressed in MC3T3E1(C4) calvarial cells by TNAP-specific shRNA. Cells were analyzed for changes in mineralization, gene expression, proliferation, apoptosis, matrix deposition and cell adhesion.
  
  
  RESULTS
  Alpl(-/-) mice feature craniofacial shape abnormalities suggestive of limited anterior-posterior growth. Craniosynostosis in the form of bony coronal suture fusion is present by three weeks after birth. Alpl(-/-) mice also exhibit marked histologic abnormalities of calvarial bones and the cranial base involving growth plates, cortical and trabecular bone within two weeks of birth. Analysis of calvarial cells in which TNAP expression was suppressed by shRNA indicates that TNAP deficiency promotes aberrant osteoblastic gene expression, diminished matrix deposition, diminished proliferation, increased apoptosis and increased cell adhesion.
  
  
  CONCLUSIONS
  These findings demonstrate that Alpl(-/-) mice exhibit a craniofacial skeletal phenotype similar to that seen in infants with HPP, including true bony craniosynostosis in the context of severely diminished bone mineralization. Future studies will be required to determine if TNAP deficiency and other forms of rickets promote craniosynostosis directly through abnormal calvarial cell behavior, or indirectly due to deficient growth of the cranial base.
authors.affiliation: Department of Orthodontics and Pediatric Dentistry, School of Dentistry, University of Michigan, Ann Arbor, MI, USA.
authors.forename: Jin
authors.initials: J
authors.lastname: Liu
authors.affiliation: Department of Orthodontics and Pediatric Dentistry, School of Dentistry, University of Michigan, Ann Arbor, MI, USA.
authors.forename: Hwa Kyung
authors.initials: HK
authors.lastname: Nam
authors.affiliation: Department of Orthodontics and Pediatric Dentistry, School of Dentistry, University of Michigan, Ann Arbor, MI, USA.
authors.forename: Cassie
authors.initials: C
authors.lastname: Campbell
authors.affiliation: Sanford-Burnham Medical Research Institute, La Jolla, CA, USA.
authors.forename: Kellen Cristina da Silva
authors.initials: KC
authors.lastname: Gasque
authors.affiliation: Sanford-Burnham Medical Research Institute, La Jolla, CA, USA.
authors.forename: José Luis
authors.initials: JL
authors.lastname: Millán
authors.affiliation: Department of Orthodontics and Pediatric Dentistry, School of Dentistry, University of Michigan, Ann Arbor, MI, USA. Electronic address: nhatch@umich.edu.
authors.forename: Nan E
authors.initials: NE
authors.lastname: Hatch
mesh_terms: D000469:Alkaline Phosphatase / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D019465:Craniofacial Abnormalities / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D003398:Craniosynostoses / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D004195:Disease Models, Animal; D007014:Hypophosphatasia / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D051379:Mice; D018345:Mice, Knockout
chemical_list: D000469:Alkaline Phosphatase
keywords: Animal models; Bone; Craniofacial; Disorders of phosphate; Micro-CT (μCT)