Reference point indentation is insufficient for detecting alterations in traditional mechanical properties of bone under common experimental conditions

Reference point indentation (RPI) was developed as a novel method to assess mechanical properties of bone in vivo, yet it remains unclear what aspects of bone dictate changes/differences in RPI-based parameters. The main RPI parameter, indentation distance increase (IDI), has been proposed to be inversely related to the ability of bone to form/tolerate damage. The goal of this work was to explore the relationshipre-intervention RPI measurebetween RPI parameters and traditional mechanical properties under varying experimental conditions (drying and ashing bones to increase brittleness, demineralizing bones and soaking in raloxifene to decrease brittleness). Beams were machined from cadaveric bone, pre-tested with RPI, subjected to experimental manipulation, post-tested with RPI, and then subjected to four-point bending to failure. Drying and ashing significantly reduced RPI's IDI, as well as ultimate load (UL), and energy absorption measured from bending tests. Demineralization increased IDI with minimal change to bending properties. Ex vivo soaking in raloxifene had no effect on IDI but tended to enhance post-yield behavior at the structural level. These data challenge the paradigm of an inverse relationship between IDI and bone toughness, both through correlation analyses and in the individual experiments where divergent patterns of altered IDI and mechanical properties were noted. Based on these results, we conclude that RPI measurements alone, as compared to bending tests, are insufficient to reach conclusions regarding mechanical properties of bone. This proves problematic for the potential clinical use of RPI measurements in determining fracture risk for a single patient, as it is not currently clear that there is an IDI, or even a trend of IDI, that can determine clinically relevant changes in tissue properties that may contribute to whole bone fracture resistance

Time course of rapid bone loss and cortical porosity formation observed by longitudinal μCT in a rat model of CKD

Background Rodent studies of bone in chronic kidney disease have primarily relied on end-point examinations of bone microarchitecture. This study used longitudinal in vivo microcomputed tomography (in vivo μCT) to characterize the onset and progression of bone loss, specifically cortical porosity, in the Cy/+ rat of model of CKD. Methods Male CKD rats and normal littermates were studied. In vivo μCT scans of the right distal tibia repeated at 25, 30, and 35 weeks were analyzed for longitudinal changes in cortical and trabecular bone morphometry. In vitro μCT scans of the tibia and femur identified spatial patterns of bone loss across distal, midshaft and proximal sites. Results CKD animals had reduced BV/TV and cortical BV at all time points but developed cortical porosity and thinning between 30 and 35 weeks. Cortical pore formation was localized near the endosteal surface. The severity of bone loss was variable across bone sites, but the distal tibia was representative of both cortical and trabecular changes. Conclusions The distal tibia was found to be a sensitive suitable site for longitudinal imaging of both cortical and trabecular bone changes in the CKD rat. CKD trabecular bone loss progressed through ~30 weeks followed by a sudden acceleration in cortical bone catabolism. These changes varied in timing and severity across individuals, and cortical bone loss and porosity progressed rapidly once initiated. The inclusion of longitudinal μCT in future studies will be important for both reducing the number of required animals and to track individual responses to treatment

Effects of combination treatment with alendronate and raloxifene on skeletal properties in a beagle dog model.

A growing number of studies have investigated combination treatment as an approach to treat bone disease. The goal of this study was to investigate the combination of alendronate and raloxifene with a particular focus on mechanical properties. To achieve this goal we utilized a large animal model, the beagle dog, used previously by our laboratory to study both alendronate and raloxifene monotherapies. Forty-eight skeletally mature female beagles (1-2 years old) received daily oral treatment: saline vehicle (VEH), alendronate (ALN), raloxifene (RAL) or both ALN and RAL. After 6 and 12 months of treatment, all animals underwent assessment of bone material properties using in vivo reference point indentation (RPI) and skeletal hydration using ultra-short echo magnetic resonance imaging (UTE-MRI). End point measures include imaging, histomorphometry, and mechanical properties. Bone formation rate was significantly lower in iliac crest trabecular bone of animals treated with ALN (-71%) and ALN+RAL (-81%) compared to VEH. In vivo assessment of properties by RPI yielded minimal differences between groups while UTE-MRI showed a RAL and RAL+ALN treatment regimens resulted in significantly higher bound water compared to VEH (+23 and +18%, respectively). There was no significant difference among groups for DXA- or CT-based measures lumbar vertebra, or femoral diaphysis. Ribs of RAL-treated animals were smaller and less dense compared to VEH and although mechanical properties were lower the material-level properties were equivalent to normal. In conclusion, we present a suite of data in a beagle dog model treated for one year with clinically-relevant doses of alendronate and raloxifene monotherapies or combination treatment with both agents. Despite the expected effects on bone remodeling, our study did not find the expected benefit of ALN to BMD or structural mechanical properties, and thus the viability of the combination therapy remains unclear

Structural and estimated material properties.

<p>Structural and estimated material properties.</p

Body mass and femoral dimensions.

<p>Body mass and femoral dimensions.</p

In vivo assessments after 6 and 12 months of treatments among the groups.

<p>(A) First cycle indentation depth from BioDent cyclic indentation device showed significant group effects but no effect of time or interaction between group and time. (B) Bone material strength index (BMSi) from Osteoprobe impact testing showed no significant effect of treatment or time. (C) Bound water from UTE-MRI measures of the proximal tibia showing higher percentages in raloxifene and combination treatments relative to vehicles. Gray bars represent 6 months; Black bars represent 12 months. * p < 0.05 when pooled across groups relative to VEH group.</p

Dynamic histomorphometry of trabecular and cortical bone.

<p>Dynamic histomorphometry of trabecular and cortical bone.</p

Bone density, architecture and geometry.

<p>Bone density, architecture and geometry.</p

In vivo tibia indentation and hydration properties after 6 and 12 months of treatment.

<p>In vivo tibia indentation and hydration properties after 6 and 12 months of treatment.</p

Iliac crest trabecular bone formation rate.

<p>* p < 0.05 versus vehicle.</p