DMP1 depletion decreases bone mineralization in vivo: an FTIR imaging analysis - PubMed (original) (raw)

DMP1 depletion decreases bone mineralization in vivo: an FTIR imaging analysis

Yunfeng Ling et al. J Bone Miner Res. 2005 Dec.

Abstract

The role of DMP1 in mineralization was analyzed by comparing bone mineral and matrix properties in dmp1-null female mice to heterozygous and wildtype controls by FTIR imaging spectroscopy. The observed decreased mineral content in dmp1 null mice indicates a key role for dmp1 in bone mineralization. Indirect effects of DMP1 on other systems also determine the KO phenotype.

Introduction: Dentin matrix protein 1 (DMP1), an acidic phosphorylated extracellular matrix protein, is highly expressed in mineralized tissues. In vitro, DMP1 peptides can promote or inhibit mineralization depending on the extent of phosphorylation, the peptide size, and concentration. To clarify the biological function of DMP1 protein on in vivo mineralization, this study analyzed bone properties of dmp1 knockout (KO) mice compared with heterozygous (HET) and wildtype (WT) controls.

Materials and methods: Tibias from dmp1 KO and age-, sex-, and background-matched HET and WT mice at 4 and 16 weeks (N(total) = 60) were examined by Fourier transform infrared imaging (FTIRI), histology (n = 6 per genotype and age; N = 36), and geometry by muCT (n = 4 per genotype and age; N = 24). Serum ionic calcium and phosphate concentrations were also determined.

Results: The mineral-to-matrix ratios (spectroscopic parameter of relative mineral content) were significantly lower in dmp1 KO mice tibias compared with WT and HET at 4 and 16 weeks. The mineral crystallinity (crystal size/perfection) was significantly increased in dmp1 KO and HET mice relative to WT. Collagen cross-link ratios (a spectroscopic parameter related to the relative amounts of nonreducible/reducible collagen cross-links) in dmp1 KO were not significantly different from WT and HET. Based on muCT, cortical bone cross-sectional areas at 16 but not 4 weeks were significantly reduced in the KO compared with controls. Maximum, minimum, and polar cross-sectional moments of inertia were significantly lower in dmp1 KO than in HET at 16 weeks but not at 4 weeks. Histological analysis and muCT 3-D images suggested that dmp1 KO mice had osteomalacia. Dmp1 KO mice had significantly lower ionic calcium and phosphate concentrations relative to WT, whereas in the HET, values for phosphate were equivalent, and calcium values were decreased relative to WT values.

Conclusions: The findings of decreased mineral-to-matrix ratio and increased crystal size in bones of dmp1 KO mice suggest that DMP1 has multiple roles (both direct and indirect) in the regulation of postnatal mineralization. We suggest that direct effects on mineral formation, crystal growth, and indirect effects on regulation of Ca x P concentrations and matrix turnover all contribute to the dominant phenotype in the dmp1 KO mouse.

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Conflict of interest statement

The authors have no conflict of interest.

Figures

FIG. 1

FIG. 1

Photomicrographs of the proximal tibia of a 16-week-old _dmp1_-WT (left) and KO (right) mouse stained by the von Kossa (top) and Goldner’s trichrome staining (bottom) showing the morphologic changes resulted from DMP1 deficiency. Smaller and disorganized mineralized trabeculae (arrowhead) surrounded by widespread excessive osteoid, increased width of osteoid seams (arrow), irregular and expanded growth plate, enlarged metaphysis, smaller secondary ossification center, considerable cortex porosity in the dmp1 KO mice tibia compared with WT. Labels 1–3 in the first picture indicates anatomic sites examined by FTIRI: (1) primary spongiosa; (2) secondary spongiosa; (3) cortical bone.

FIG. 2

FIG. 2

Representative 3-D rendered μCT images of 16-week dmp1 (A, C, E, and G) WT and (B, D, F, and H) KO mice tibias. Dmp1 (B) KO tibia showed enlarged metaphysis, abnormal shape and shorter length compared with (A) WT tibia. In longitudinal sections of tibias, significantly enlarged flared metaphysis, expanded growth plates, and smaller secondary ossification centers were seen in dmp1 (D) KO tibia relative to (C) WT. In 5-mm-thick cross-section under the growth plate, trabeculae were sparser in the dmp1 (F) KO than in the dmp1 (E) WT. In the 1-mm cross-section of tibia midshaft, thinner cortical wall and larger marrow space were observed in the dmp1 (H) KO compared with (G) WT. All scale bars are 1 mm.

FIG. 3

FIG. 3

(A) Mineral-to-matrix ratio, spectroscopic mineral content, analyzed by FTIRI at cortex, primary spongiosa and secondary spongiosa of tibias of dmp1 WT, HET, and KO mice at 4 and 16 weeks. (B) Mineral crystal size and perfection (crystallinity; 1030/1020 intensity ratio) and (C) XLR (1660/1690 intensity ratio). *p < 0.05 vs. 4-week WT; #p < 0.05 vs. 4-week HET; € p < 0.05 vs. 16-week WT; £p < 0.05 vs. 16-week HET; ¥p < 0.05 vs. 4-week KO. All values are mean ± SD (n = 6).

FIG. 4

FIG. 4

Typical FTIRIs of various spectral parameters in dmp1 WT, HET, and KO mice at 4 and 16 weeks. Representative images of mineral-to-matrix ratio at (A) the cortex and (B) secondary spongiosa and typical images of 1030/1020 ratio (crystallinity) at (C) the cortex and (D) secondary spongiosa. x and y axes represent pixel number. The numerical scales shown on the right side represent the intensity for each parameter in the same row and were kept consistent in WT, HET, and KO.

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