The role of mineralization and organic matrix in the microhardness of bone tissue from controls and osteoporotic patients.

Degree of mineralization of bone (DMB) is a major intrinsic determinant of bone strength at the tissue level but its contribution to the microhardness (Vickers indentation) at the intermediary level of organization of bone tissue, i.e., Bone Structural Units (BSUs), has never been assessed. The purpose of this study was to analyze the relationship between the microhardness, the DMB and the organic matrix, measured in BSUs from human iliac bone biopsies. Iliac bone samples from controls and osteoporotic patients (men and women), embedded in methyl methacrylate, were used. Using a Vickers indenter, microhardness (kg/mm2) was measured, either globally on surfaced blocks or focally on 100 microm-thick sections from bone samples (load of 25 g applied during 10 sec; CV=5%). The Vickers indenter was more suited than the Knoop indenter for a tissue like bone in which components are diversely oriented. Quantitative microradiography performed on 100 microm-thick sections, allowed measurement of parameters reflecting the DMB (g/cm3). Assessed on the whole bone sample, both microhardness and DMB were significantly lower (-10% and -7%, respectively) in osteoporotic patients versus controls (p<0.001). When measured separately at the BSU level, there were significant positive correlations between microhardness and DMB in controls (r2=0.36, p<0.0001) and osteoporotic patients (r2=0.43, p<0.0001). Mineralization is an important determinant of the microhardness, but did not explain all of its variance. To highlight the role of the organic matrix in bone quality, microhardness of both osteoid and adjacent calcified matrix were measured in iliac samples from subjects with osteomalacia. Microhardness of organic matrix is 3-fold lower than the microhardness of calcified tissue. In human calcanei, microhardness was significantly correlated with DMB (r2=0.33, p=0.02) and apparent Young's modulus (r2=0.26, p=0.03). In conclusion, bone microhardness measured by Vickers indentation is an interesting methodology for the evaluation of bone strength and its determinants at the BSU level. Bone microhardness is linked to Young's modulus of bone and is strongly correlated to mineralization, but the organic matrix accounts for about one third of its variance.

[1]  Peter Zioupos,et al.  Mechanical properties of nacre and highly mineralized bone , 2001, Proceedings of the Royal Society of London. Series B: Biological Sciences.

[2]  Marco Viceconti,et al.  The effect of tissue condition and applied load on Vickers hardness of human trabecular bone. , 2007, Journal of biomechanics.

[3]  J. Currey,et al.  Hardness, Young's modulus and yield stress in mammalian mineralized tissues , 1990 .

[4]  G. Dalsky,et al.  Differential Effects of Teriparatide and Alendronate on Bone Remodeling in Postmenopausal Women Assessed by Histomorphometric Parameters , 2005, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[5]  A. Boskey,et al.  Bone Fragility and Collagen Cross‐Links , 2004, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[6]  J. Compston,et al.  Influence of estrogen therapy at conventional and high doses on the degree of mineralization of iliac bone tissue: a quantitative microradiographic analysis in postmenopausal women. , 2005, Bone.

[7]  G. Pharr,et al.  Variations in the individual thick lamellar properties within osteons by nanoindentation. , 1999, Bone.

[8]  P. Meunier,et al.  Effects of bisphosphonates on matrix mineralization. , 2002, Journal of musculoskeletal & neuronal interactions.

[9]  D. Burr.  The contribution of the organic matrix to bone's material properties. , 2002, Bone.

[10]  D. Burr,et al.  In situ examination of the time-course for secondary mineralization of Haversian bone using synchrotron Fourier transform infrared microspectroscopy. , 2008, Matrix biology : journal of the International Society for Matrix Biology.

[11]  G Boivin,et al.  Bone mineral density reflects bone mass but also the degree of mineralization of bone: therapeutic implications. , 1997, Bone.

[12]  Paul Roschger,et al.  From brittle to ductile fracture of bone , 2006, Nature materials.

[13]  S. Krane,et al.  Metabolic bone disease , 1977 .

[14]  P. Delmas,et al.  Bone quality--the material and structural basis of bone strength and fragility. , 2006, The New England journal of medicine.

[15]  E. Ritman,et al.  The effect of risedronate on bone mineralization as measured by micro-computed tomography with synchrotron radiation: correlation to histomorphometric indices of turnover. , 2005, Bone.

[16]  H Follet,et al.  The degree of mineralization is a determinant of bone strength: a study on human calcanei. , 2004, Bone.

[17]  S M Jorgensen,et al.  Long-term risedronate treatment normalizes mineralization and continues to preserve trabecular architecture: sequential triple biopsy studies with micro-computed tomography. , 2006, Bone.

[18]  P. Fratzl,et al.  Validation of quantitative backscattered electron imaging for the measurement of mineral density distribution in human bone biopsies. , 1998, Bone.

[19]  D. Dempster.  The Impact of Bone Turnover and Bone-Active Agents on Bone Quality: Focus on the Hip , 2002, Osteoporosis International.

[20]  J. H. Koolstra,et al.  Relationship between tissue stiffness and degree of mineralization of developing trabecular bone. , 2008, Journal of biomedical materials research. Part A.

[21]  P. Meunier,et al.  Alendronate increases bone strength by increasing the mean degree of mineralization of bone tissue in osteoporotic women. , 2000, Bone.

[22]  Hilla Peretz,et al.  Ju n 20 03 Schrödinger ’ s Cat : The rules of engagement , 2003 .

[23]  J. Currey,et al.  Microhardness and Young's modulus in cortical bone exhibiting a wide range of mineral volume fractions, and in a bone analogue , 1990 .

[24]  M. Grynpas,et al.  How Does Fluoride Affect Dentin Microhardness and Mineralization? , 2005, Journal of dental research.

[25]  A. Rapoff,et al.  Microindentation in bone: Hardness variation with five independent variables , 2007, Journal of materials science. Materials in medicine.

[26]  D. McNally,et al.  Knoop microhardness anisotropy of the ovine radius. , 2000, Journal of biomechanics.

[27]  R. Rizzoli,et al.  Bone strength and its determinants , 2003, Osteoporosis International.

[28]  P. Fratzl,et al.  Effects of intermittent parathyroid hormone administration on bone mineralization density in iliac crest biopsies from patients with osteoporosis: a paired study before and after treatment. , 2003, The Journal of clinical endocrinology and metabolism.

[29]  M. Glimcher.  The Nature of the Mineral Phase in Bone: Biological and Clinical Implications , 1998 .

[30]  F. Bauss,et al.  Effects of ibandronate on bone quality: preclinical studies. , 2007, Bone.

[31]  P. Price,et al.  The Size Exclusion Characteristics of Type I Collagen , 2007, Journal of Biological Chemistry.

[32]  D. Carlström.  Micro-hardness measurements on single haversian systems in bone , 1954, Experientia.

[33]  P. Lips,et al.  Contribution of raloxifene and calcium and vitamin D3 supplementation to the increase of the degree of mineralization of bone in postmenopausal women. , 2003, The Journal of clinical endocrinology and metabolism.

[34]  J. Currey,et al.  The effects of strain rate, reconstruction and mineral content on some mechanical properties of bovine bone. , 1975, Journal of biomechanics.

[35]  P. Fratzl,et al.  Constant mineralization density distribution in cancellous human bone. , 2003, Bone.

[36]  Robert O. Ritchie,et al.  Invited Article , 2004 .

[37]  M. Oyen,et al.  Nanoindentation hardness of mineralized tissues. , 2006, Journal of biomechanics.

[38]  R. Whitehead.  Biopsies , 1954, British medical journal.

[39]  S. Huja,et al.  Indentation Properties of Young and Old Osteons , 2006, Calcified Tissue International.

[40]  R. Amprino.  Investigations on some physical properties of bone tissue. , 1958, Acta anatomica.

[41]  P. Meunier,et al.  Methodological considerations in measurement of bone mineral content , 2003, Osteoporosis International.

[42]  P. Meunier,et al.  Histomorphometric Methods Applied to Bone , 1993 .

[43]  Françoise Peyrin,et al.  Quantification of the degree of mineralization of bone in three dimensions using synchrotron radiation microtomography. , 2002, Medical physics.

[44]  P. Meunier,et al.  The mineralization of bone tissue: a forgotten dimension in osteoporosis research , 2003, Osteoporosis International.

[45]  P. Zysset,et al.  Morphological and Mechanical Properties of Bone Structural Units: A Two-Case Study , 2002 .

[46]  P Zioupos,et al.  Exploring the Effects of Hypermineralisation in Bone Tissue by Using an Extreme Biological Example , 2000, Connective tissue research.

[47]  P. Fratzl,et al.  The bone mineralization density distribution as a fingerprint of the mineralization process. , 2007, Bone.

[48]  P. Zysset,et al.  Nanoindentation discriminates the elastic properties of individual human bone lamellae under dry and physiological conditions. , 2002, Bone.

[49]  John D Currey,et al.  How Well Are Bones Designed to Resist Fracture? , 2003, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[50]  R O Ritchie,et al.  Fatigue of mineralized tissues: cortical bone and dentin. , 2008, Journal of the mechanical behavior of biomedical materials.

[51]  D Aubry,et al.  Effect of microstructure on the mechanical properties of Haversian cortical bone. , 2006, Bone.

[52]  P. Delmas,et al.  The role of collagen in bone strength , 2005, Osteoporosis International.

[53]  D Vashishth,et al.  Bone stiffness predicts strength similarly for human vertebral cancellous bone in compression and for cortical bone in tension. , 2000, Bone.

[54]  P. Meunier,et al.  The Degree of Mineralization of Bone Tissue Measured by Computerized Quantitative Contact Microradiography , 2002, Calcified Tissue International.

[55]  H. Plenk,et al.  A new scanning electron microscopy approach to the quantification of bone mineral distribution: backscattered electron image grey-levels correlated to calcium K alpha-line intensities. , 1995, Scanning microscopy.