Establishing Biomechanical Mechanisms in Mouse Models: Practical Guidelines for Systematically Evaluating Phenotypic Changes in the Diaphyses of Long Bones

Mice are widely used in studies of skeletal biology, and assessment of their bones by mechanical testing is a critical step when evaluating the functional effects of an experimental perturbation. For example, a gene knockout may target a pathway important in bone formation and result in a “low bone mass” phenotype. But how well does the skeleton bear functional loads; eg, how much do bones deform during loading and how resistant are bones to fracture? By systematic evaluation of bone morphological, densitometric, and mechanical properties, investigators can establish the “biomechanical mechanisms” whereby an experimental perturbation alters whole‐bone mechanical function. The goal of this review is to clarify these biomechanical mechanisms and to make recommendations for systematically evaluating phenotypic changes in mouse bones, with a focus on long‐bone diaphyses and cortical bone. Further, minimum reportable standards for testing conditions and outcome variables are suggested that will improve the comparison of data across studies. Basic biomechanical principles are reviewed, followed by a description of the cross‐sectional morphological properties that best inform the net cellular effects of a given experimental perturbation and are most relevant to biomechanical function. Although morphology is critical, whole‐bone mechanical properties can only be determined accurately by a mechanical test. The functional importance of stiffness, maximum load, postyield displacement, and work‐to‐fracture are reviewed. Because bone and body size are often strongly related, strategies to adjust whole‐bone properties for body mass are detailed. Finally, a comprehensive framework is presented using real data, and several examples from the literature are reviewed to illustrate how to synthesize morphological, tissue‐level, and whole‐bone mechanical properties of mouse long bones. © 2015 American Society for Bone and Mineral ResearchPeer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/111801/1/jbmr2539.pd

Machining-induced thermal damage in cortical bone: necrosis and micro-mechanical integrity

In bone cutting, the tissue is exposed to necrosis due to temperature elevation, which can significantly influence postoperative results in orthopaedic surgeries. This damage is usually revealed through histological analysis to show the necrotic extent; however, this technique does not capture mechanical damage, which is essential for a full material integrity assessment. Here, with micro-mechanics, it is demonstrated that machining-induced damage in bone extends beyond the necrotic region. Drilling with different conditions was performed on ex-vivo bovine cortical bone, inducing different damage degrees. Micro-pillar compression tests were performed in the machined sub-surface to identify changes in properties and failure modes caused by drilling. It was revealed that at high cutting temperatures, the bone near the machined surface suffers from lower modulus (−42%), strength (−41%) and brittle behaviour, whereas the bulk bone remains undamaged with pristine properties and ductile behaviour. Histology was also performed to evaluate necrosis and, surprisingly, it was found that the brittle and weaker bone layer is more than three times larger when compared to the necrotic layer, clearly showing that the drilling thermo-mechanical effect could affect not only biologically, but also micro-mechanically. Consequently, these results reveal another kind of bone damage that has so far been neglected

Bone Material Properties in Osteogenesis Imperfecta

Osteogenesis imperfecta entrains changes at every level in bone tissue, from the disorganisation of the collagen molecules and mineral platelets within and between collagen fibrils, to the macro-architecture of the whole skeleton. Investigations using an array of sophisticated instruments at multiple scale levels have now determined many aspects of the effect of the disease on the material properties of bone tissue. The brittle nature of bone in osteogenesis imperfecta reflects both increased bone mineralisation density – the quantity of mineral in relation to the quantity of matrix within a specific bone volume – and altered matrix-matrix and matrix mineral interactions. Contributions to fracture resistance at multiple scale lengths are discussed, comparing normal and brittle bone. Integrating the available information provides both a better understanding of the effect of current approaches to treatment – largely improved architecture and possibly some macro-scale toughening - and indicates potential opportunities for alternative strategies that can influence fracture resistance at longer length scales

Biomolecular regulation, composition and nanoarchitecture of bone mineral

Abstract Tough natural nanocomposites like bone, nacre and sea sponges contain within their hierarchy, a mineral (phosphate, silicate or carbonate) phase that interacts with an organic phase. In bone, the role of mineral ultrastructure (organization, morphology, composition) is crucial to the mechanical and biological properties of the tissue. Better understanding of mineral interaction with the organic matrix, in particular non-collagenous proteins, osteocalcin (OC) and osteopontin (OPN), can lead to better design of biomimetic materials. Using small angle x-ray scattering (SAXS) and wavelength dispersive spectroscopy (WDS) on single (OC−/− and OPN−/−) and double (OC-OPN−/−;−/−) genetic knockout mice bones, we demonstrate that both osteocalcin and osteopontin have specific roles in the biomolecular regulation of mineral in bone and together they are major determinants of the quality of bone mineral. Specifically, for the first time, we show that proteins osteocalcin and osteopontin regulate bone mineral crystal size and organization in a codependent manner, while they independently determine crystal shape. We found that OC is more dominant in the regulation of the physical properties of bone mineral, while OPN is more dominant in the regulation of the mineral composition

A direct role of collagen glycation in bone fracture

Non-enzymatic glycation (NEG) is an age-related process accelerated by diseases like diabetes, and causes the accumulation of advanced glycation end-products (AGEs). NEG-mediated modification of bone's organic matrix, principally collagen type-I, has been implicated in impairing skeletal physiology and mechanics. Here, we present evidence, from in vitro and in vivo models, and establish a causal relationship between collagen glycation and alterations in bone fracture at multiple length scales. Through atomic force spectroscopy, we established that NEG impairs collagen's ability to dissipate energy. Mechanical testing of in vitro glycated human bone specimen revealed that AGE accumulation due to NEG dramatically reduces the capacity of organic and mineralized matrix to creep and caused bone to fracture under impact at low levels of strain (3000-5000;strain) typically associated with fall. Fracture mechanics tests of NEG modified human cortical bone of varying ages, and their age-matched controls revealed that NEG disrupted microcracking based toughening mechanisms and reduced bone propagation and initiation fracture toughness across all age groups. A comprehensive mechanistic model, based on experimental and modeling data, was developed to explain how NEG and AGEs are causal to, and predictive of bone fragility. Furthermore, fracture mechanics and indentation testing on diabetic mice bones revealed that diabetes mediated NEG severely disrupts bone matrix quality in vivo. Finally, we show that AGEs are predictive of bone quality in aging humans and have diagnostic applications in fracture risk

Correction: Advanced Glycation Endproducts and Bone Material Properties in Type 1 Diabetic Mice.

[This corrects the article DOI: 10.1371/journal.pone.0154700.]