Hypophosphatasia-associated deficiencies in mineralization and gene expression in cultured dental pulp cells obtained from human teeth.
BACKGROUND
Mutations in the gene ALPL in hypophosphatasia (HPP) reduce the function of tissue nonspecific alkaline phosphatase, and the resulting increase in pyrophosphate (PP(i)) contributes to bone and tooth mineralization defects by inhibiting physiologic calcium-phosphate (P(i)) precipitation. Although periodontal phenotypes are well documented, pulp/dentin abnormalities have been suggested in the clinical literature although reports are variable and underlying mechanisms remains unclear. In vitro analyses were used to identify mechanisms involved in HPP-associated pulp/dentin phenotypes.


METHODS
Primary pulp cells cultured from HPP subjects were established to assay alkaline phosphatase (ALP) activity, mineralization, and gene expression compared with cells from healthy controls. Exogenous P(i) was provided to the correct P(i)/PP(i) ratio in cell culture.


RESULTS
HPP cells exhibited significantly reduced ALP activity (by 50%) and mineral nodule formation (by 60%) compared with the controls. The expression of PP(i) regulatory genes was altered in HPP pulp cells, including reduction in the progressive ankylosis gene (ANKH) and increased ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1). Odontoblast marker gene expression was disrupted in HPP cells, including reduced osteopontin (OPN), dentin matrix protein 1 (DMP1), dentin sialophosphoprotein (DSPP), and matrix extracellular phosphoprotein (MEPE). The addition of P(i) provided a corrective measure for mineralization and partially rescued the expression of some genes although cells retained altered messenger RNA levels for PP(i)-associated genes.


CONCLUSIONS
These studies suggest that under HPP conditions pulp cells have the compromised ability to mineralize and feature a disrupted odontoblast profile, providing a first step toward understanding the molecular mechanisms for dentin phenotypes observed in HPP.
Department of Prosthodontics and Periodontics, Division of Periodontics, School of Dentistry, Campinas State University, Piracicaba, São Paulo, Brazil.
Thaisângela L
TL
Rodrigues
Brian L
BL
Foster
Karina G
KG
Silverio
Luciane
L
Martins
Marcio Z
MZ
Casati
Enilson A
EA
Sallum
Martha J
MJ
Somerman
Francisco H
FH
Nociti
D000293:Adolescent; D000469:Alkaline Phosphatase / Q000235:genetics*; D019943:Amino Acid Substitution; D000704:Analysis of Variance; D002118:Calcium / Q000378:metabolism; D016022:Case-Control Studies; D003782:Dental Pulp / Q000166:cytology / Q000503:physiopathology*; D003804:Dentin / Q000473:pathology*; D011756:Diphosphates / Q000378:metabolism*; D004200:Diseases in Twins / Q000235:genetics; D015536:Down-Regulation; D016326:Extracellular Matrix Proteins / Q000096:biosynthesis / Q000235:genetics; D005260:Female; D015870:Gene Expression; D006023:Glycoproteins / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D007014:Hypophosphatasia / Q000235:genetics* / Q000473:pathology / Q000503:physiopathology; D008297:Male; D020125:Mutation, Missense; D009804:Odontoblasts / Q000378:metabolism / Q000473:pathology*; D053495:Osteopontin / Q000096:biosynthesis / Q000235:genetics; D028061:Phosphate Transport Proteins / Q000096:biosynthesis / Q000235:genetics; D010750:Phosphoproteins / Q000096:biosynthesis / Q000235:genetics; D010727:Phosphoric Diester Hydrolases / Q000096:biosynthesis / Q000235:genetics; D061251:Primary Cell Culture; D011755:Pyrophosphatases / Q000096:biosynthesis / Q000235:genetics; D012795:Sialoglycoproteins / Q000096:biosynthesis / Q000235:genetics; D018709:Statistics, Nonparametric; D014074:Tooth Calcification / Q000235:genetics*; D055815:Young Adult; D000099311:Dentin Sialophosphoprotein
D000469:Alkaline Phosphatase; D002118:Calcium; D011756:Diphosphates; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D053495:Osteopontin; D028061:Phosphate Transport Proteins; D010750:Phosphoproteins; D010727:Phosphoric Diester Hydrolases; D011755:Pyrophosphatases; D012795:Sialoglycoproteins; D000099311:Dentin Sialophosphoprotein; C498770:ANKH protein, human; C513309:DMP1 protein, human; C486579:MEPE protein, human; C107241:diphosphoric acid; C548267:ALPL protein, human; C065962:ectonucleotide pyrophosphatase phosphodiesterase 1