Multiscale experimental study on the effects of different weight-bearing levels during moderate treadmill exercise on bone quality in growing female rats

Abstract

Background: Bone tissue displays a hierarchical organization. Mechanical environments influence bone mass and structure. This study aimed to explore the effects of different mechanical stimuli on growing bone properties at macro-micro-nano scales.

Methods: Sixty five-week-old female Wistar rats were treadmill exercised at moderate intensity with the speed of 12 m/min, and then randomly divided into five groups according to weight-bearing level. After 8 weeks of experiment, femurs were harvested to perform multiscale tests.

Results: Bone formation was significantly increased by weight-bearing exercise, whereas bone resorption was not significantly inhibited. Trabecular and cortical bone mineral densities showed no significant increase by weight-bearing exercise. The microstructure of trabecular bone was significantly improved by 12% weight-bearing exercise. However, similar positive effects were not observed with further increase in weight-bearing levels. The nanomechanical properties of trabecular bone were not significantly changed by weight-bearing exercise. The macrostrength of whole femur and the nanomechanical properties of cortical bone significantly decreased in the 19% and 26% weight-bearing exercise groups.

Conclusion: When rats ran on the treadmill at moderate intensity during growth period, additional 12% weight-bearing level could significantly increase bone formation, improve microstructure of trabecular bone, as well as maintain the structure and mechanical properties of cortical bone. Excessive weight-bearing level caused no positive effects on the trabecular bone microstructure and properties of cortical bone at all scales. In addition, increased weight-bearing level exerted no significant influence on trabecular and cortical bone mineral densities.

Keywords: Bone quality; Moderate intensity; Treadmill exercise; Weight-bearing.

Fig. 1

The schematic of experimental grouping and the measurements performed in the femur. a Experimental grouping. b Measurement of the femur

Fig. 2

Indentation sites for nanoindentation test. Actual indented sites were marked by red cross under optical microscopy, and the sample thickness was 2 mm

Fig. 3

The body weight of each group during the experiment. Error bars represent standard deviation (SD). At T2, the 8-week exercise program started

Fig. 4

The cross-sectional areas in the middle segment of femoral shaft at the end of experiment. Error bars represent SD. ^△Statistically different from the ex group (p < 0.05); *Statistically different from the eb5 group (p < 0.05); ^◇Statistically different from the eb12 group (p < 0.05); ^#Statistically different from the eb19 group (p < 0.05); ^□Statistically different from the eb26 group (p < 0.05)

Fig. 5

The length of femur. Error bars represent SD

Fig. 6

The calf muscles weight. Error bars represent SD

Fig. 7

The serum concentrations of ALP and TRACP in each group. Error bars represent SD. ^△Statistically different from the ex group (p < 0.05); *Statistically different from the eb5 group (p < 0.05); ^◇Statistically different from the eb12 group (p < 0.05); ^#Statistically different from the eb19 group (p < 0.05); ^□Statistically different from the eb26 group (p < 0.05). a ALP concentration. b TRACP concentration

Fig. 8

The macromechanical parameters of the left femur measured by three-point bending mechanical test. Error bars represent SD. ^△Statistically different from the ex group (p < 0.05); *Statistically different from the eb5 group (p < 0.05); ^◇Statistically different from the eb12 group (p < 0.05); ^#Statistically different from the eb19 group (p < 0.05); ^□ Statistically different from the eb26 group (p < 0.05). a Elastic modulus. b Failure load. c Energy absorption. d Stiffness

Fig. 9

The MS/BS and MAR of trabecular bone in the femoral head. Error bars represent SD. MAR mineral apposition rate, MS/BS the ratio of mineralizing surface to bone surface. ^△Statistically different from the ex group (p < 0.05); *Statistically different from the eb5 group (p < 0.05); ^◇Statistically different from the eb12 group (p < 0.05); ^#Statistically different from the eb19 group (p < 0.05); ^□Statistically different from the eb26 group (p < 0.05). a MS/BS, b MAR

Fig. 10

The 3D microstructure parameters of trabecular bone in femoral head and cortical bone in femoral shaft. Error bars represent SD. Tb.BMD trabecular bone mineral density, BV/TV bone volume fraction, Tb.Th trabecular thickness, Tb.N trabecular number, Tb.Sp trabecular separation, Ct.BMD cortical bone mineral density, Ct.Th cortical bone thickness. ^△Statistically different from the ex group (p < 0.05); *Statistically different from the eb5 group (p < 0.05); ^◇Statistically different from the eb12 group (p < 0.05); ^#Statistically different from the eb19 group (p < 0.05); ^□Statistically different from the eb26 group (p < 0.05). a Tb.BMD, b BV/TV, c Tb.Th, d Tb.N, e Tb.Sp, f Ct.BMD, g Ct.Th

Fig. 11

The indentation modulus and hardness of trabecular and cortical bone sites as determined by nanoindentation. Error bars represent SD. E_Tb.L longitudinal indentation modulus of trabecular bone, H_Tb.L longitudinal hardness of trabecular bone, E_Ct.L longitudinal indentation modulus of cortical bone, H_Ct.L longitudinal hardness of cortical bone, E_Ct.T transverse indentation modulus of cortical bone, H_Ct.T transverse hardness of cortical bone. ^△Statistically different from the ex group (p < 0.05); *Statistically different from the eb5 group (p < 0.05); ^◇Statistically different from the eb12 group (p < 0.05); ^#Statistically different from the eb19 group (p < 0.05); ^□Statistically different from the eb26 group (p < 0.05). a E_Tb.L of trabecular bone, b H_Tb.L of trabecular bone, c E_Ct.L of cortical bone, d H_Ct.L of cortical bone, e E_Ct.T of cortical bone, f H_Ct.T of cortical bone