An Investigation of the Mineral in Ductile and Brittle Cortical Mouse Bone

Abstract

Bone is a strong and tough material composed of apatite mineral, organic matter and water. Changes in composition and organization of these building blocks affect bone’s mechanical integrity. Skeletal disorders often affect bone’s mineral phase, either by variations in the collagen or directly altering mineralization. The aim of the current study was to explore the differences in the mineral of brittle and ductile cortical bone at the mineral (nm) and tissue (µm) levels using two mouse phenotypes. Osteogenesis imperfecta murine (oim−/−) mice were used to model brittle bone; PHOSPHO1 mutants (Phospho1−/−) had ductile bone. They were compared to their respective wild-type controls. Femora were defatted and ground to powder to measure average mineral crystal size using X-ray diffraction (XRD), and to monitor the bulk mineral to matrix ratio via thermogravimetric analysis (TGA). XRD scans were run after TGA for phase identification, to assess the fractions of hydroxyapatite and β-tricalcium phosphate. Tibiae were embedded to measure elastic properties with nanoindentation and the extent of mineralization with backscattered electron microscopy (qbSEM). Interestingly, the mineral of brittle oim−/− and ductile Phospho1−/− bones had many similar characteristics. Both pathology models had smaller apatite crystals, lower mineral to matrix ratio, and showed more thermal conversion to β-tricalcium phosphate than their wild-types, indicating deviations from stoichiometric hydroxyapatite in the original mineral. The degree of mineralization of the bone matrix was different for each strain: oim−/− were hypermineralized, while Phospho1−/− were hypomineralized. However, alterations in the mineral were associated with reduced tissue elastic moduli in both pathologies. Results revealed that despite having extremely different whole bone mechanics, the mineral of oim−/− and Phospho1−/− has several similar trends at smaller length scales. This indicates that alterations from normal crystal size, composition, and structure will reduce the mechanical integrity of bone.

Endocrine role of bone: recent and emerging perspectives beyond osteocalcin

“…Complete ablation of Phospho1 in mice results in a similar phenotype to that of fro/fro mice, with Phospho1 K/K mice having significant skeletal pathology, spontaneous fractures, bowed long bones, osteomalacia and scoliosis in early life (Huesa et al 2011, Yadav et al 2011, 2014, Rodriguez-Florez et al 2014. These results indicate that PHOSPHO1 and SMPD3 are within the same metabolic pathway required for skeletal mineralisation in the mouse (Khavandgar Z, Oldknow KJ, Murshed M & Farquharson C, unpublished observations).…”

Osteogenesis imperfecta

“…First, compared with controls, the alignment of collagen fibrils was found to be more disordered and less lamellar in both oim mouse models 124,127 and biopsy samples from patients with osteo genesis imperfecta 128 ; second, the collagenous matrix is not only reduced in amount 129 but also showed more non enzymatic crosslinks 124 ; third, unmineral ized collagen fibrils from the oim mouse model showed reduced strength under tensioning 130 and more affinity to water 131 . By contrast, less tissue water was found in fully mineralized bone of the oim mice 132 and in bone biop sies from patients with osteogenesis imperfecta 121 . Last, this is in good agreement with the increased amount of mineral in the matrix, corresponding to a higher density of thinner mineral platelets in both oim mouse mod els 124,127 and patients with osteogenesis imperfecta 120 .…”

Genetic causes and mechanisms of Osteogenesis Imperfecta

“…The reduction in bone mass is likely due to high bone turnover, as oim −/− mice show increased osteoblast and osteoclast numbers by histomorphometry [33]. Bones from oim −/− mice have altered mineral content with dense and disordered apatite crystals, which likely contribute to increased brittleness [32,34–38]. Furthermore, collagen cross-linking is altered in oim −/− mice.…”