Three-Point Bending of Rat Femur in the Mediolateral Direction: Introduction and Validation of a Novel Biomechanical Testing Protocol

Abstract

Mediolateral three-point bending of the rat midfemur was developed to enable the assessment of the mechanical competence of the elliptic bone cross-section in terms of its widest diameter, the apparent primary direction of bone adaptation to loading.Introduction: Today, the most commonly used method to characterize the biomechanical properties of appendicular long bones is the three-point bending testing of the midfemur in the anteroposterior (AP) direction. However, as the diameter of the elliptic cross-section of femoral diaphysis is widest in the orthogonal mediolateral (ML) direction, the femoral diaphysis should also show the highest resistance to bending along this direction. The objective of this study was thus to introduce and validate a mechanical testing protocol for femoral midshaft along the ML direction. Materials and Methods: To determine the repeatability of the novel testing protocol, 38 pairs of rat femora underwent a comprehensive structural analysis by pQCT followed by ML three-point bending. For comparison of the repeatability, corresponding tests were performed on the femoral neck. To validate the novel testing direction, the left hindlimb of 24 rats was neurectomized for 6 months, whereas the right limb served as an intact control. After excision, one half of these pairs of femora were randomly subjected to three-point bending test in the conventional AP direction and the remaining in the orthogonal ML direction. Results: The precision (CV rms ) of breaking load, stiffness, and energy absorption of the femoral midshaft in the ML direction was 3.8%, 6.6%, and 14.5%, respectively. The corresponding values for femoral neck compression test were 7.6%, 17.9%, and 18.7%, respectively. The loading-induced effect on the femoral midshaft (difference between the neurectomized [nonloaded] and contralateral intact [loaded] femur) was +2.2%, +1.9%, and +2.1% in the AP direction and −18.9%, −17.6%, and −20.3% in the ML direction (p < 0.01 for all comparisons), respectively. Conclusions: Our results show that testing of rat femoral midshaft in the ML direction is a precise and biologically valid method to determine the structural strength of this widely used skeletal site in experimental bone research.

Supraphysiological testosterone enanthate administration prevents bone loss and augments bone strength in gonadectomized male and female rats

“…We employed a three-point bending test on the femoral midshaft to evaluate the biomechanical characteristics of cortical bone (21). Consistent with previous literature (64), femoral strength deficits were not observed within 4 wk of GX, which may indicate that mechanical loading is sufficient to maintain cortical bone mass and femoral strength in the near absence of circulating sex hormones, at least during the relatively short duration of this study (43).…”

Mechanical Properties of Bone Due to SOST Expression: A 3-Point Bending Assessment of Murine Femurs

“…exercise) improves bone mass and strength by stimulating the addition of new bone onto surfaces experiencing high strains, whereas surfaces that experience small strains remain largely quiescent [8]. In line with the established functional bone adaptation to loading, first described as Wolff's Law, bones -as locomotive organs -adapt their structural rigidity and strength to incident loading through changes in structural particulars (mineral mass, geometry, architecture, material properties) [4,9]. Simplified, form follows function [10].…”

“…Although biomechanical testing of bones, similar to any other in vitro measurement, represents an oversimplification of the complexity of the actual in vivo situation, the intention is to test the skeletal structure of interest as closely as possible in terms of the predominant loading environment [9].…”

“…A load-deformation test is conducted and results are recorded in deformation as a function of the applied load, typically tension, compression, torsion, or bending [11]. In recent bone research, the most frequently used method for characterization of long bone biomechanical properties of the appendicular skeleton is 3-point bending of the femoral midshaft [2,9]. This test, coupled with measured geometric dimensions can be extrapolated to give the resulting stress and strain experienced by the bone.…”

“…Point A marks the yield force, point B marks the ultimate force, and the distance between points A and fracture is the post-yield displacement…………………………………...… 26 2. 9 Stress versus strain graph for left legs of 6 month SOST TG mice…………….. 27 Histogram displaying the comparison of cross sectional area for 6 month old SOST Transgenic mice. WT 1 mice were control littermates and faithfully express murine SOST.…”