Elsevier

Bone

Volume 103, October 2017, Pages 334-346
Bone

Full Length Article
Tissue-specific mineralization defects in the periodontium of the Hyp mouse model of X-linked hypophosphatemia

https://doi.org/10.1016/j.bone.2017.07.026Get rights and content

Highlights

  • A biological explanation is proposed for the higher susceptibility of XLH patients to develop periodontitis.

  • PHEX deficiency severely affects periodontal tissues, resulting in an altered tooth attachment complex.

  • Hyp mouse oral bone is less capable of repair.

  • Hyp tooth cementum is less able to adapt to tooth movement.

  • Although mineralization appears similarly disturbed in Hyp bone and cementum, osteopontin accumulation is tissue-specific.

Abstract

X-linked hypophosphatemia (XLH) is a dento-osseous disorder caused by inactivating mutations in the PHEX gene, leading to renal phosphate wasting and hypophosphatemia, and impaired mineralization of bones and teeth. In the oral cavity, recent reports suggest a higher susceptibility of XLH patients to periodontitis, where patients present with impaired tooth cementum – a bone-like tissue involved in tooth attachment to the jaw bones and post-eruption tooth positioning – and a higher frequency of intrabony defects. In the present study, the pathobiology of alveolar bone and tooth cementum was investigated in the Hyp mouse, the murine analog of XLH. PHEX deficiency in XLH/Hyp dramatically alters the periodontal phenotype, with hypoplasia of tooth root cementum associated with a lack of periodontal ligament attachment and the presence of an immature apatitic mineral phase of all periodontal mineralized tissues. Challenging the Hyp periodontium in two surgical experimental models – ligature-induced periodontal breakdown and repair, and a model of tooth movement adaptation inducing cementum formation – we show that bone and cementum formation, and their healing, are altered. Bone and cementum mineralization appear similarly disturbed, where hypomineralized pericellular matrix surrounds cells, and where the protein osteopontin (OPN, a mineralization inhibitor) accumulates in a tissue-specific manner, most notably in the perilacunar matrix surrounding osteocytes. Although the pathobiology is different between XLH/Hyp bone and cementum, our results show a major XLH phenotype in oral mineralized tissues consistent with variations in patient susceptibility to periodontal disorders.

Introduction

Disorders of mineral metabolism result in both skeletal and extraskeletal consequences. Skeletal disorders have to be considered not only with regard to the bone itself, but also in the context of disturbed mineral ion metabolism occurring at extraskeletal sites, where alterations can include any one, or a combination, of the following: abnormalities in i) calcium, phosphorus, parathyroid hormone (PTH), and vitamin D metabolism, ii) bone turnover, mineralization, volume, linear growth, and strength, and iii) vascular or soft tissue calcification [1]. In bones and teeth, these abnormalities lead to deficient mineralization known as rickets, osteomalacia and odonto/dentinomalacia.

Although rickets appeared in epidemic form during the industrial revolution from deficiency of vitamin D, in the 1930s, a familial inheritance pattern was proposed for patients displaying vitamin D-resistant rickets [2]. In the past four decades, different mechanisms involved in the initiation and maintenance of the disturbances in bone and mineral metabolism have been uncovered, and X-linked hypophosphatemia (XLH) has been identified as the most prevalent heritable rickets/osteomalacia (~ 1:20,000) [3], [4]. This genetic disorder of calcium and phosphate metabolism is caused by inactivating PHEX (phosphate-regulating gene with homologies to endopeptidases on the X chromosome) gene mutations, with XLH being characterized by renal phosphate wasting and defective mineralization of the skeleton and dentition [5], [6], [7]. Loss-of-function mutations in the PHEX gene in XLH cause reduced (or absent) enzymatic activity of PHEX, a zinc metallo-endopeptidase [8], [9], [10] highly expressed by bone (osteoblasts and osteocytes) and tooth (odontoblasts) cells [11]. Skeletal complications in XLH patients include rickets and osteo/odontomalacia, with clinical manifestations including delayed walking, leg bowing (genu varum) or knock knees (genu valgum), growth failure and dental issues such as the occurrence of “spontaneous” tooth abscesses in the absence of caries or trauma [7], [12], [13]. Ultrastructural studies have revealed that the appendicular bone in XLH patients presents with hypomineralized perilacunar matrix around osteocytes often referred to as “halos” (or periosteocytic lesions) [14], and the XLH dentin appears poorly mineralized with large, unmineralized interglobular spaces surrounding unmerged mineralization foci [12]. The mineralization-inhibiting protein osteopontin (OPN) has been proposed as a key player in the pathobiology of these mineralization defects as it accumulates in XLH bone in periosteocytic lesions [15], [16], and in interglobular dentin [17], [18].

Recently, accumulating evidence suggests a higher susceptibility of XLH patients to periodontitis [6], [19], [20], with the recent report of reduced acellular cementum thickness and a higher frequency of intrabony defects displayed in patients with XLH [20]. Consistent with these observations, cementum hypoplasia has been described in the murine model of XLH [21], [22]. Dental cementum is a mineralized tissue covering the tooth root that functions in ligamentous tooth attachment to mandibular and maxillary bone, and post-eruptive adjustment of tooth position in the jaws. During formation of cellular cementum near the tips of tooth roots, some cementoblasts become embedded in the cementoid matrix and become cementocytes. Although cementum presents specific features [23], [24], [25], in some ways it is considered as a bone-like tissue with terminally differentiated cells (cementocytes) embedded in a mineralized extracellular matrix, similar to osteocytes in bone [26]. In the context of genetic disorders disturbing calcium and phosphate metabolism, it still remains to be determined whether cementum formation and function are affected, as previously reported in XLH for bone and dentin.

In the present study, we analyzed how PHEX deficiency affects matrix mineralization of cementum and alveolar bone in the mandible and maxilla under normal and experimental conditions. The periodontal phenotype associated with XLH was examined in human samples, and characterized in greater depth using observations from experiments on the Hyp mouse – the XLH mouse model harboring a Phex mutation that phenocopies the biochemical and clinical features of XLH. Two experimental manipulations (alveolar bone healing and induction of cementum apposition) were performed in Hyp mice to explore the pathobiology of XLH oral bone and cementum.

Section snippets

Human teeth

Human permanent teeth from XLH patients and from gender- and age-matched control individuals extracted prior to orthodontic treatments were collected in the dental departments of Paris Nord Val de Seine Hospitals, AP-HP, France. Informed consent was obtained from patients in agreement with French law (agreement n°DC-2009-927, Cellule Bioéthique DGRI/A5). Teeth were fixed for 7 days at 4 °C in 4% paraformaldehyde solution at pH 7.2–7.4, followed by microwave-assisted decalcification in 4.13% EDTA

Periodontal phenotype of XLH patients and Hyp mice

PHEX was found to be expressed in the periodontium, not only by odontoblasts, osteoblasts and osteocytes, but also by the cellular cementum-forming cells – the cementoblasts and cementocytes (Supplemental Fig. 1).

Observations of tooth roots revealed that XLH teeth had a thin and poorly mineralized acellular cementum (panel A in Fig. 1), consistent with what has been reported previously [20]. In Hyp mice, the acellular cementum also was thin (panel B in Fig. 1a, b). Sirius red staining of molar

Discussion

Here, we show that both cementum and bone formation and healing are altered in XLH/Hyp periodontium. We demonstrate that loss of PHEX activity dramatically alters the periodontal phenotype in humans and in the Hyp mouse (the murine model of XLH), supporting recent clinical studies showing that adult patients with XLH have a higher susceptibility to periodontitis [19], [20]. Interestingly, although cementum and bone mineralization appears similarly altered, each tissue presents a tissue-specific

Conflict of interest statement

BRC, GF, BB, LS, JS, GP, MB, TS, MDM, CC and CB have declared that no conflict of interest exists.

AL: Research grant and/or consulting from Ultragenyx Pharmaceutical, Novato, CA.

Fundings sources

This work was supported by a grant from "La Fondation pour la Recherche Médicale" for Life Imaging Facility of Paris Descartes University (Plateforme d'Imageries du Vivant - PIV) (FRM DGE20111123012). BRC was supported by "La Fondation pour la Recherche Médicale" (PhD scholarship FDM20140731354) (France).

Authors' contribution

BRC, CC and CB designed the study. BRC and GF performed the Raman spectroscopy experiments. BRC and CB performed the surgeries. BRC, LS and JS performed the in vivo radiographic follow-up of all samples. BRC and BB performed other experiments of the study. TS provided the mouse model. BRC, MBD, AL and CC followed the patients and collected the bone and tooth specimens. BRC, MDM, CC and CB contributed to the analysis of the data and the drafting of the manuscript. All authors reviewed and

Acknowledgements

The authors thank Jean-Marc Masse and Alain Schmitt (Institut Cochin, Paris, France) and Annie Llorens (EA2496, Montrouge, France) for their help with the microscopy methods. MDM is a member of the FRQ-S Network for Oral and Bone Health Research, and the McGill Centre for Bone and Periodontal Research.

References (55)

  • S. Schwartz et al.

    Oral findings in patients with autosomal dominant hypophosphatemic bone disease and X-linked hypophosphatemia: further evidence that they are different diseases

    Oral surgery, Oral Medicine, and Oral Pathology

    (1988)
  • C. Chaussain-Miller et al.

    Dental abnormalities in patients with familial hypophosphatemic vitamin D-resistant rickets: prevention by early treatment with 1-hydroxyvitamin D

    J. Pediatr.

    (2003)
  • G.R. Naveh et al.

    Tooth movements are guided by specific contact areas between the tooth root and the jaw bone: a dynamic 3D microCT study of the rat molar

    J. Struct. Biol.

    (2012)
  • G.R. Naveh et al.

    Tooth periodontal ligament: direct 3D microCT visualization of the collagen network and how the network changes when the tooth is loaded

    J. Struct. Biol.

    (2013)
  • G.R. Naveh et al.

    Tooth-PDL-bone complex: response to compressive loads encountered during mastication — a review

    Arch. Oral Biol.

    (2012)
  • M. McClung et al.

    An update on osteoporosis pathogenesis, diagnosis, and treatment

    Bone

    (2017)
  • O.I. Kolek et al.

    1α, 25-Dihydroxyvitamin D3 upregulates FGF23 gene expression in bone: the final link in a renal-gastrointestinal-skeletal axis that controls phosphate transport

    American Journal of Physiology-Gastrointestinal and Liver Physiology

    (2005)
  • F. Albright et al.

    Rickets resistant to vitamin d therapy

    Am. J. Dis. Child.

    (1937)
  • A. Linglart et al.

    Therapeutic management of hypophosphatemic rickets from infancy to adulthood

    Endocrine Connections

    (2014)
  • M.D. McKee et al.

    Extracellular matrix mineralization in periodontal tissues: noncollagenous matrix proteins, enzymes, and relationship to hypophosphatasia and X-linked hypophosphatemia

    Periodontology 2000

    (2013)
  • T.O. Carpenter

    The expanding family of hypophosphatemic syndromes

    J. Bone Miner. Metab.

    (2012)
  • P.S. Rowe

    The PEX gene: its role in X-linked rickets, osteomalacia, and bone mineral metabolism

    Exp. Nephrol.

    (1997)
  • F. Francis et al.

    Genomic organization of the human PEX gene mutated in X-linked dominant hypophosphatemic rickets

    Genome Res.

    (1997)
  • H.S. Tenenhouse

    X-linked hypophosphataemia: a homologous disorder in humans and mice

    Nephrology, Dialysis, Transplantation: Official Publication of the European Dialysis and Transplant Association - European Renal Association

    (1999)
  • A.F. Ruchon et al.

    Pex mRNA is localized in developing mouse osteoblasts and odontoblasts

    The Journal of Histochemistry and Cytochemistry: Official Journal of the Histochemistry Society

    (1998)
  • T. Boukpessi et al.

    Dentin alteration of deciduous teeth in human hypophosphatemic rickets

    Calcif. Tissue Int.

    (2006)
  • C. Chaussain-Miller et al.

    Dentin structure in familial hypophosphatemic rickets: benefits of vitamin D and phosphate treatment

    Oral Dis.

    (2007)
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