Self-repair of rat cortical bone microdamage after fatigue loading in vivo

Abstract

Bone microdamage can be repaired through bone remodeling induced by loading. In this study, a loading device was developed for improved efficiency and the self-repair process of bone microdamage was studied in ovariectomized rats. First, four-point bending fixtures capable of holding two live rats simultaneously were designed. Rats were loaded and subjected to a sinusoidal wave for 10,000 cycles. They were then divided into four groups to evaluate time points from 1 to 4 weeks in the microdamage repair process. The loaded right ulna was used for microdamage parameter analysis, and the loaded right radius was tested for mechanical properties. In all groups, microdamage consisted primarily of microcracks, which were observed in bone surrounding the force-bearing point. The values of the microdamage parameters were significantly lower at 3 weeks than at 2 weeks. However, none of the differences in mechanical properties between any four groups were statistically significant. This study shows that the improved application of loading in the form of bending for double-rat simultaneous administration was practical and efficient. These results suggest that microdamage was repaired between 2 weeks to 3 weeks after fatigue damage and microdamage is a more sensitive index of bone quality than mechanical properties.

Figure 1

The four-point bending fatigue test apparatus is an open loop system consisting of the following components: (1) upper connecting rod, (2) linear compression spring, (3) upper indenter, (4) lower supports, (5) loading plates where the rat is placed, (6) T-shaped upper indenter slide, (7) C-shaped lower support slide, and (8) lower connecting rod. The picture shows the following. (a) Front view of the apparatus. (b) Lateral view. (c) Fixture with a single rat. (d) Fixture system for two rats.

Figure 2

Basic fuchsin staining showing microcracks (white arrow) in (a) OVX 2nd week group rats and (c) OVX 1st week group rats. (b, d) Red epifluorescent light microscopy showing the same microcrack (white dashed arrow) in the identical view. Diffused microdamage (black arrow) was detected in (c) cortical bone (black arrow) and under (d) red epifluorescent light microscopy (black dashed arrow). Scale bars = 100 μm.

Figure 3

Parameters and mechanical properties of microcracks and absorptive spaces versus time. Specific values are listed in Table 1. (a) Average microcrack length (Cr.Le) and average number of microcracks (Cr.N), (b) microcrack surface density (Cr.S.Dn) and microcrack density (Cr.Dn), (c) peak load and modulus of elasticity, and (d) absorptive lacunar density. *P < 0.05, **P < 0.01.

Figure 4

(a, d) Basic fuchsin staining showed resorption spaces (black arrow) in the cortical bone. Blue-violet epifluorescent light microscopy revealed area of osteogenesis (white arrow) and (b, e) green fluorescence attributable to calcein showing osteogenesis during the 4 days preceding euthanasia, (b, e) orange fluorescence and (c, f) yellow fluorescence attributed to acheomycin, which indicated osteogenesis in the 2 weeks before euthanasia. Scale bars = 50 μm.