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Central role of pyrophosphate in acellular cementum formation.

Foster BL, Nagatomo KJ, Nociti FH, Fong H, Dunn D, Tran AB, Wang W, Narisawa S, Millán JL, Somerman MJ - PLoS ONE (2012)

Bottom Line: Though PP(i) regulators are widely expressed, cementoblasts selectively expressed greater ANK and NPP1 along the root surface, and dramatically increased ANK or NPP1 in models of reduced PP(i) output, in compensatory fashion.In vitro mechanistic studies confirmed that under low PP(i) mineralizing conditions, cementoblasts increased Ank (5-fold) and Enpp1 (20-fold), while increasing PP(i) inhibited mineralization and associated increases in Ank and Enpp1 mRNA.These findings underscore developmental differences in acellular versus cellular cementum, and suggest new approaches for cementum regeneration.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Oral Connective Tissue Biology, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, Maryland, United States of America. brian.foster@nih.gov

ABSTRACT

Background: Inorganic pyrophosphate (PP(i)) is a physiologic inhibitor of hydroxyapatite mineral precipitation involved in regulating mineralized tissue development and pathologic calcification. Local levels of PP(i) are controlled by antagonistic functions of factors that decrease PP(i) and promote mineralization (tissue-nonspecific alkaline phosphatase, Alpl/TNAP), and those that increase local PP(i) and restrict mineralization (progressive ankylosis protein, ANK; ectonucleotide pyrophosphatase phosphodiesterase-1, NPP1). The cementum enveloping the tooth root is essential for tooth function by providing attachment to the surrounding bone via the nonmineralized periodontal ligament. At present, the developmental regulation of cementum remains poorly understood, hampering efforts for regeneration. To elucidate the role of PP(i) in cementum formation, we analyzed root development in knock-out ((-/-)) mice featuring PP(i) dysregulation.

Results: Excess PP(i) in the Alpl(-/-) mouse inhibited cementum formation, causing root detachment consistent with premature tooth loss in the human condition hypophosphatasia, though cementoblast phenotype was unperturbed. Deficient PP(i) in both Ank and Enpp1(-/-) mice significantly increased cementum apposition and overall thickness more than 12-fold vs. controls, while dentin and cellular cementum were unaltered. Though PP(i) regulators are widely expressed, cementoblasts selectively expressed greater ANK and NPP1 along the root surface, and dramatically increased ANK or NPP1 in models of reduced PP(i) output, in compensatory fashion. In vitro mechanistic studies confirmed that under low PP(i) mineralizing conditions, cementoblasts increased Ank (5-fold) and Enpp1 (20-fold), while increasing PP(i) inhibited mineralization and associated increases in Ank and Enpp1 mRNA.

Conclusions: Results from these studies demonstrate a novel developmental regulation of acellular cementum, wherein cementoblasts tune cementogenesis by modulating local levels of PP(i), directing and regulating mineral apposition. These findings underscore developmental differences in acellular versus cellular cementum, and suggest new approaches for cementum regeneration.

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Reduced pyrophosphate alters gene expression in cervical cementoblasts.Opn mRNA is markedly increased in root-lining cementoblasts in both (B) Ank and (C) Enpp1−/−, compared to (A) Ank and Enpp1+/+ controls. Increased numbers of cells associated with the thick cervical cementum express Dmp1 in (E) Ank and (F) Enpp1−/− molars, compared to (D) +/+ controls. Bsp gene expression was not different in cementoblasts in (H) Ank and (I) Enpp1−/− vs. (G) +/+ controls. Black arrowheads indicate regions of positively stained cells. All panels are samples from mice at 14 dpn. Abbreviations: d = dentin, c = (cervical) cementum; p = periodontal ligament; b = bone. Scale bar = 100 µm.
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pone-0038393-g008: Reduced pyrophosphate alters gene expression in cervical cementoblasts.Opn mRNA is markedly increased in root-lining cementoblasts in both (B) Ank and (C) Enpp1−/−, compared to (A) Ank and Enpp1+/+ controls. Increased numbers of cells associated with the thick cervical cementum express Dmp1 in (E) Ank and (F) Enpp1−/− molars, compared to (D) +/+ controls. Bsp gene expression was not different in cementoblasts in (H) Ank and (I) Enpp1−/− vs. (G) +/+ controls. Black arrowheads indicate regions of positively stained cells. All panels are samples from mice at 14 dpn. Abbreviations: d = dentin, c = (cervical) cementum; p = periodontal ligament; b = bone. Scale bar = 100 µm.

Mentions: Cementum, bone, and dentin are also characterized by their extracellular matrix (ECM) protein composition, and these ECM proteins contribute to crystal growth and regulation, and affect mechanical properties of these tissues. Because of the dramatic changes in cementum apposition, we investigated the ECM profile in PPi deficient mice. In the low PPi environment of the Ank and Enpp1−/− mice, the thick cervical cementum was marked by increased OPN and dentin matrix protein 1 (DMP1), proteins of the SIBLING family (Figure 7A–F and G–L). OPN staining strongly labeled control acellular cementum, and was intensely expressed in the corresponding Ank and Enpp1−/− cervical cementum and associated cementoblast cells. DMP1, a marker for osteocytes, odontoblasts, and cementocytes, was present at low or undetectable levels in acellular cementum in controls, in contrast to intense localization in expanded Ank and Enpp1−/− cementum. OPN and DMP1 levels were not changed in Ank or Enpp1−/− apical cementum, as well as in other dentoalveolar locations. The source of the increased OPN and DMP1 protein was confirmed, as cementoblast gene expression for both Opn and Dmp1 mRNA was increased in Ank and Enpp−/− mice (Figure 8A–C and D–F). OPN and DMP1 expression changes were not observed in other cell populations in the dentoalveolar complex in these mice. Another characteristic marker for cementum, BSP, was present in control and cementum (Figure 7M–R), and where protein concentration was diluted in the larger cementum volume of the Ank and Enpp1−/− mice, mRNA levels in cementoblasts were unaltered (Figure 8G–I).


Central role of pyrophosphate in acellular cementum formation.

Foster BL, Nagatomo KJ, Nociti FH, Fong H, Dunn D, Tran AB, Wang W, Narisawa S, Millán JL, Somerman MJ - PLoS ONE (2012)

Reduced pyrophosphate alters gene expression in cervical cementoblasts.Opn mRNA is markedly increased in root-lining cementoblasts in both (B) Ank and (C) Enpp1−/−, compared to (A) Ank and Enpp1+/+ controls. Increased numbers of cells associated with the thick cervical cementum express Dmp1 in (E) Ank and (F) Enpp1−/− molars, compared to (D) +/+ controls. Bsp gene expression was not different in cementoblasts in (H) Ank and (I) Enpp1−/− vs. (G) +/+ controls. Black arrowheads indicate regions of positively stained cells. All panels are samples from mice at 14 dpn. Abbreviations: d = dentin, c = (cervical) cementum; p = periodontal ligament; b = bone. Scale bar = 100 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3366957&req=5

pone-0038393-g008: Reduced pyrophosphate alters gene expression in cervical cementoblasts.Opn mRNA is markedly increased in root-lining cementoblasts in both (B) Ank and (C) Enpp1−/−, compared to (A) Ank and Enpp1+/+ controls. Increased numbers of cells associated with the thick cervical cementum express Dmp1 in (E) Ank and (F) Enpp1−/− molars, compared to (D) +/+ controls. Bsp gene expression was not different in cementoblasts in (H) Ank and (I) Enpp1−/− vs. (G) +/+ controls. Black arrowheads indicate regions of positively stained cells. All panels are samples from mice at 14 dpn. Abbreviations: d = dentin, c = (cervical) cementum; p = periodontal ligament; b = bone. Scale bar = 100 µm.
Mentions: Cementum, bone, and dentin are also characterized by their extracellular matrix (ECM) protein composition, and these ECM proteins contribute to crystal growth and regulation, and affect mechanical properties of these tissues. Because of the dramatic changes in cementum apposition, we investigated the ECM profile in PPi deficient mice. In the low PPi environment of the Ank and Enpp1−/− mice, the thick cervical cementum was marked by increased OPN and dentin matrix protein 1 (DMP1), proteins of the SIBLING family (Figure 7A–F and G–L). OPN staining strongly labeled control acellular cementum, and was intensely expressed in the corresponding Ank and Enpp1−/− cervical cementum and associated cementoblast cells. DMP1, a marker for osteocytes, odontoblasts, and cementocytes, was present at low or undetectable levels in acellular cementum in controls, in contrast to intense localization in expanded Ank and Enpp1−/− cementum. OPN and DMP1 levels were not changed in Ank or Enpp1−/− apical cementum, as well as in other dentoalveolar locations. The source of the increased OPN and DMP1 protein was confirmed, as cementoblast gene expression for both Opn and Dmp1 mRNA was increased in Ank and Enpp−/− mice (Figure 8A–C and D–F). OPN and DMP1 expression changes were not observed in other cell populations in the dentoalveolar complex in these mice. Another characteristic marker for cementum, BSP, was present in control and cementum (Figure 7M–R), and where protein concentration was diluted in the larger cementum volume of the Ank and Enpp1−/− mice, mRNA levels in cementoblasts were unaltered (Figure 8G–I).

Bottom Line: Though PP(i) regulators are widely expressed, cementoblasts selectively expressed greater ANK and NPP1 along the root surface, and dramatically increased ANK or NPP1 in models of reduced PP(i) output, in compensatory fashion.In vitro mechanistic studies confirmed that under low PP(i) mineralizing conditions, cementoblasts increased Ank (5-fold) and Enpp1 (20-fold), while increasing PP(i) inhibited mineralization and associated increases in Ank and Enpp1 mRNA.These findings underscore developmental differences in acellular versus cellular cementum, and suggest new approaches for cementum regeneration.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Oral Connective Tissue Biology, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, Maryland, United States of America. brian.foster@nih.gov

ABSTRACT

Background: Inorganic pyrophosphate (PP(i)) is a physiologic inhibitor of hydroxyapatite mineral precipitation involved in regulating mineralized tissue development and pathologic calcification. Local levels of PP(i) are controlled by antagonistic functions of factors that decrease PP(i) and promote mineralization (tissue-nonspecific alkaline phosphatase, Alpl/TNAP), and those that increase local PP(i) and restrict mineralization (progressive ankylosis protein, ANK; ectonucleotide pyrophosphatase phosphodiesterase-1, NPP1). The cementum enveloping the tooth root is essential for tooth function by providing attachment to the surrounding bone via the nonmineralized periodontal ligament. At present, the developmental regulation of cementum remains poorly understood, hampering efforts for regeneration. To elucidate the role of PP(i) in cementum formation, we analyzed root development in knock-out ((-/-)) mice featuring PP(i) dysregulation.

Results: Excess PP(i) in the Alpl(-/-) mouse inhibited cementum formation, causing root detachment consistent with premature tooth loss in the human condition hypophosphatasia, though cementoblast phenotype was unperturbed. Deficient PP(i) in both Ank and Enpp1(-/-) mice significantly increased cementum apposition and overall thickness more than 12-fold vs. controls, while dentin and cellular cementum were unaltered. Though PP(i) regulators are widely expressed, cementoblasts selectively expressed greater ANK and NPP1 along the root surface, and dramatically increased ANK or NPP1 in models of reduced PP(i) output, in compensatory fashion. In vitro mechanistic studies confirmed that under low PP(i) mineralizing conditions, cementoblasts increased Ank (5-fold) and Enpp1 (20-fold), while increasing PP(i) inhibited mineralization and associated increases in Ank and Enpp1 mRNA.

Conclusions: Results from these studies demonstrate a novel developmental regulation of acellular cementum, wherein cementoblasts tune cementogenesis by modulating local levels of PP(i), directing and regulating mineral apposition. These findings underscore developmental differences in acellular versus cellular cementum, and suggest new approaches for cementum regeneration.

Show MeSH
Related in: MedlinePlus