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NMR investigation of the role of osteocalcin and osteopontin at the organic-inorganic interface in bone.

ABSTRACT: Mechanical resilience of bone tissue decreases with age. The ability to comprehensively probe and understand bone properties could help alleviate this problem. One important aspect of bone quality that has recently been made evident is the presence of dilatational bands formed by osteocalcin (OC) and osteopontin (OPN), which contribute to fracture toughness. However, experimental evidence of the structural role of these two proteins at the organic-mineral interface in bone is still needed. Solid state nuclear magnetic resonance (SSNMR) is emerging as a useful technique in probing molecular level aspects of bone. Here, we present the first SSNMR study of bone tissue from genetically modified mice lacking OC and/or OPN. Probing the mineral phase, the organic matrix and their interface revealed that, despite the absence of OC and OPN, the organic matrix and mineral were well preserved, and the overall exposure of collagen to hydroxyapatite (HA) nanoparticles was hardly affected. However, the proximity to the HA surface was slightly increased for a number of bone components including less abundant amino acids like lysine, suggesting that this is how the tissue compensates for the lack of OC and OPN. Taken together, the NMR data supports the recently proposed model, in which the contribution of OC-OPN to fracture toughness is related to their presence at the extrafibrillar organic-mineral interfaces, where they reinforce the network of mineralized fibrils and form dilatational bands. In an effort toward further understanding the structural role of individual amino acids of low abundance in bone, we then explored the possibility of specific (13)C enrichment of mouse bone, and report the first SSNMR spectra of 97% (13)C lysine-enriched tissue. Results show that such isotopic enrichment allows valuable molecular-level structural information to be extracted, and sheds light on post-translational modifications undergone by specific amino acids in vivo.

SUBMITTER: Nikel O

PROVIDER: S-EPMC3901427 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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NMR investigation of the role of osteocalcin and osteopontin at the organic-inorganic interface in bone.

Nikel Ondřej O Laurencin Danielle D McCallum Scott A SA Gundberg Caren M CM Vashishth Deepak D

Langmuir : the ACS journal of surfaces and colloids 20131101 45

Mechanical resilience of bone tissue decreases with age. The ability to comprehensively probe and understand bone properties could help alleviate this problem. One important aspect of bone quality that has recently been made evident is the presence of dilatational bands formed by osteocalcin (OC) and osteopontin (OPN), which contribute to fracture toughness. However, experimental evidence of the structural role of these two proteins at the organic-mineral interface in bone is still needed. Solid ...[more]

PMID: 24128197

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Project description:Age-related bone fragility fractures present a significant problem for public health. Measures of bone quality are increasingly recognized to complement the conventional bone mineral density (BMD) based assessment of fracture risk. The ability to probe and understand bone quality at the molecular level is desirable in order to unravel how the structure of organic matrix and its association with mineral contribute to the overall mechanical properties. The (13)C{(31)P} REDOR MAS NMR (Rotational Echo Double Resonance Magic Angle Spinning Nuclear Magnetic Resonance) technique is uniquely suited for the study of the structure of the organic-mineral interface in bone. For the first time, we have applied it successfully to analyze the structure of intact (non-powdered) human cortical bone samples, from young healthy and old osteoporotic donors. Loading problems associated with the rapid rotation of intact bone were solved using a Finite Element Analysis (FEA) approach, and a method allowing osteoporotic samples to be balanced and spun reproducibly is described. REDOR NMR parameters were set to allow insight into the arrangement of the amino acids at the mineral interface to be accessed, and SVD (Singular Value Decomposition) was applied to enhance the signal to noise ratio and enable a better analysis of the data. From the REDOR data, it was found that carbon atoms belonging to citrate/glucosaminoglycans (GAGs) are closest to the mineral surface regardless of age or site. In contrast, the arrangement of the collagen backbone at the interface varied with site and age. The relative proximity of two of the main amino acids in bone matrix proteins, hydroxyproline and alanine, with respect to the mineral phase was analyzed in more detail, and discussed in view of glycation measurements which were carried out on the tissues. Overall, this work shows that the (13)C{(31)P} REDOR NMR approach could be used as a complementary technique to assess a novel aspect of bone quality, the organic-mineral interface structure.

Project description:Bone is the living composite biomaterial having unique structural property. Presently, there is a considerable gap in our understanding of bone structure and composition in the native state, particularly with respect to the trabecular bone, which is metabolically more active than cortical bones, and is readily lost in post-menopausal osteoporosis. We used solid-state nuclear magnetic resonance (NMR) to compare trabecular bone structure and composition in the native state between normal, bone loss and bone restoration conditions in rat. Trabecular osteopenia was induced by lactation as well as prolonged estrogen deficiency (bilateral ovariectomy, Ovx). Ovx rats with established osteopenia were administered with PTH (parathyroid hormone, trabecular restoration group), and restoration was allowed to become comparable to sham Ovx (control) group using bone mineral density (BMD) and µCT determinants. We used a technique combining (1)H NMR spectroscopy with (31)P and (13)C to measure various NMR parameters described below. Our results revealed that trabecular bones had diminished total water content, inorganic phosphorus NMR relaxation time (T1) and space between the collagen and inorganic phosphorus in the osteopenic groups compared to control, and these changes were significantly reversed in the bone restoration group. Remarkably, bound water was decreased in both osteopenic and bone restoration groups compared to control. Total water and T1 correlated strongly with trabecular bone density, volume, thickness, connectivity, spacing and resistance to compression. Bound water did not correlate with any of the microarchitectural and compression parameters. We conclude that total water, T1 and atomic space between the crystal and organic surface are altered in the trabecular bones of osteopenic rats, and PTH reverses these parameters. Furthermore, from these data, it appears that total water and T1 could serve as trabecular surrogates of micro-architecture and compression strength.

Dataset Information

NMR investigation of the role of osteocalcin and osteopontin at the organic-inorganic interface in bone.

Publications

NMR investigation of the role of osteocalcin and osteopontin at the organic-inorganic interface in bone.

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