Osteocalcin (OC) and osteopontin (OPN) are major non-collagenous proteins (NCPs) involved in bone matrix organization and deposition. In spite of this, it is currently unknown whether OC and OPN alter bone morphology and consequently affect bone fracture resistance. The goal of this study is to establish the role of OC and OPN in the determination of cortical bone size, shape, and mechanical properties. Our results show that Oc−/− and Opn−/− mice were no different from each other or wild type (WT) with respect to bone morphology (P > 0.1). Bones from mice lacking both NCPs (Oc−/− Opn−/−) were shorter with thicker cortices and larger cortical areas compared to the WT, Oc−/−, and Opn−/− groups (P < 0.05), suggesting a synergistic role for NCPs in the determination of bone morphology. Maximum bending load was significantly different among the groups (P = 0.024), while tissue mineral density (TMD) or measures of stiffness and strength were not different (P > 0.1). We conclude that the removal of both OC and OPN from bone matrix induces morphological adaptation at the structural level to maintain bone strength.

Non-collagenous proteins are a vital component of bone matrix. Amongst them, osteocalcin (OC) and osteopontin (OPN) hold special significance due to their intimate interaction with the mineral and collagenous matrix in bone. Both proteins have been associated with microdamage and fracture, but their structural role in energy dissipation is unclear. This study used bone tissue from genetic deficient mice lacking OC and/or OPN and subjected them to a series of creep-fatigue-creep tests. To this end, whole tibiae were loaded in four-point bending to 70% stiffness loss which captured the three characteristic phases of fatigue associated with initiation, propagation, and coalescence of microdamage. Fatigue loading preceded and followed creep tests to determine creep and dampening parameters. Microdamage in the form of linear microcracks and diffuse damage were analyzed by histology. It was shown that OC and OPN were 'activated' following stiffness loss associated with fatigue damage where they facilitated creep and dampening parameters (i.e. increased energy dissipation). More specifically, post-fatigue creep rate and dampening were significantly greater in wild-types (WTs) than genetic deficient mice (p < 0.05). These results were supported by microdamage analysis which showed significant increase in creep-associated diffuse damage formation in WTs compared to genetic deficient groups (p < 0.05). Based on these findings, we propose that during local yield events, OC and OPN rely on ionic interactions of their charged side chains and on hydrogen bonding to dissipate energy in bone.

Quasi-static tests are the most commonly used for determining the resistance to structural failure by a single load at the organ (whole bone) level. The resistance to crack initiation or growth by fracture toughness testing and fatigue loading offers additional and more direct characterization of tissue material properties. Non-traditional indentation techniques and in situ testing are being increasingly used to probe the material properties of bone ultrastructure. Destructive ex vivo testing or clinical surrogate measures are considered to be the gold standard for estimating fracture risk. The type of mechanical test used for a particular investigation depends on the length scale of interest, where the outcome variables are influenced by the interrelationship between bone structure and composition. Advancement in the sensitivity of mechanical characterization techniques to detect changes in bone at the levels subjected to modifications by aging, disease, and/or pharmaceutical treatment is required. As such, a number of techniques are now available to aid our understanding of the factors that contribute to fracture risk.

Protein phosphorylation, critical for cellular regulatory mechanisms, is implicated in various diseases. However, it remains unknown whether heterogeneity in phosphorylation of key structural proteins alters tissue integrity and organ function. Here, osteopontin phosphorylation level declined in hypo- and hyper- phosphatemia mouse models exhibiting skeletal deformities. Phosphorylation increased cohesion between osteopontin polymers, and adhesion of osteopontin to hydroxyapatite, enhancing energy dissipation. Fracture toughness, a measure of bone’s mechanical competence, increased with ex-vivo phosphorylation of wildtype mouse bones and declined with ex-vivo dephosphorylation. In osteopontin-deficient mice, global matrix phosphorylation level was not associated with toughness. Our findings suggest that phosphorylated osteopontin promotes fracture toughness in a dose-dependent manner through increased interfacial bond formation. In the absence of osteopontin, phosphorylation increases electrostatic repulsion, and likely protein alignment and interfilament distance leading to decreased fracture resistance. These mechanisms may be of importance in other connective tissues, and the key to unraveling cell–matrix interactions in diseases.

Objective: We sought to determine the degree and possible causes of variability in admission practices among individual emergency physicians (EPs) at 1 emergency department (ED) using a Canadian Emergency Department Triage Acuity Scale (CTAS)-matched ED patient population. Methods: We distributed a survey measuring attitudes and demographics to all EPs (n = 30) at a large regional hospital. Hospital admissions data from 1 calendar year were matched to individual EP survey results. Emergency physicians were ranked as "lower," "average" or "higher" admitters and, using these categorical variables, the data set was analyzed for correlations and trends. Results: Overall, 97.0% of the EPs responded to the survey. Admissions by EPs ranged from 8.7% to 17.0%, (mean 12.52, standard deviation [SD] 2.21) of all patients seen. CTAS category-specific admission data demonstrated variability in the admission ranking of individual EPs. No EPs consistently performed at any 1 admission ranking across all CTAS categories. More years of emergency medicine experience was significantly correlated with higher admissions in the CTAS-2 ranking (r = 0.4, p < 0.05). Whether a physician worked full-time, part-time or as a locum was not associated with patterns of admission, nor was any particular postgraduate certification (e.g., CCFP, CCFP EM, FRCPC) or any of the surveyed attitudinal traits. Conclusion: Individual EPs' overall and CTAS-specific admissions varied substantially, and followed an approximately normal distribution curve. Emergency physicians with more years of experience had a statistically higher CTAS-2 admission rate; however, other variables, including postgraduate certification status, decision-related attitudes toward admission, and reported practices were not associated with admission proportions. Emergency physicians tend to have uniquely individual admission ranking profiles across all the CTAS categories.

Obesity is a common comorbidity of type 2 diabetes (T2D). Therefore, increased risk of fragility fractures in T2D is often confounded by the effects of obesity. This study was conducted to elucidate the mechanistic basis by which T2D alone leads to skeletal fragility. We hypothesized that obesity independent T2D would deteriorate bone's material quality by accumulating defects in the mineral matrix and undesired modifications in its organic matrix associated with increased oxidative stress and hyperglycemia. To test this hypothesis, we used 15-week-old male non-obese mice with engineered muscle creatine kinase promoter/human dominant negative insulin growth factor 1 (IGF-I) receptor (MKR) and FVB/N wild-type (WT) controls (n = 12/group). MKR mice exhibit reduced insulin production and loss of glycemic control leading to diabetic hyperglycemia, verified by fasting blood glucose measurements (>250 mg/dL), without an increase in body weight. MKR mice showed a significant decrease in femoral radial geometry (cortical area, moment of inertia, cortical thickness, endosteal diameter, and periosteal diameter). Bone mineral density (BMD), as assessed by micro-computed tomography (μCT), remained unchanged; however, the quality of bone mineral was altered. In contrast to controls, MKR mice had significantly increased hydroxyapatite crystal thickness, measured by small-angle X-ray scattering, and elongated c-axis length of the crystals evaluated by confocal Raman spectroscopy. There was an increase in changes in the organic matrix of MKR mice, associated with enhanced glycoxidation (carboxymethyl-lysine [CML] and pentosidine) and overall glycation (fluorescent advanced glycation end products), both of which were associated with various measures of bone fragility. Moreover, increased CML formation positively correlated with elongated mineral crystal length, supporting the role of this negatively charged side chain to attract calcium ions, promote growth of hydroxyapatite, and build a physical link between mineral and collagen. Collectively, our results show, for the first time, changes in bone matrix in a non-obese T2D model in which skeletal fragility is attributable to alterations in the mineral quality and undesired organic matrix modifications.

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