Early-life exercise effects on Achilles tendon in mice selectively bred for high voluntary wheel-running behavior

Abstract

Exercise increases muscle and bone strength and mass, but effects on tendons are less documented. We investigated the impact of voluntary exercise (wheel running) during early-life exercise (weanling; 3 weeks old) compared to post-skeletal maturity (young adult; 9 weeks old) on tendon morphology and material properties. We utilized a selectively bred High Runner (HR, N = 40) mouse line and a control line (N = 40). Mice underwent 8 weeks in cages either with or without wheels. HR mice ran ~3-fold more and were smaller than controls, but exercise reduced body mass in both lines. Tendon cross-sectional area was unaffected, but tendon length showed a line*exercise interaction (p = 0.0410) and a near-significant line*age interaction (p = 0.0866). HR mice broadly had greater yield stress (p = 0.0262) and tended toward higher failure stress (p = 0.0676) than controls. Work to failure was greater in younger cohort mice (p = 0.0435), and marginal age-related interactions were observed for modulus (line*exercise, p = 0.0632) and yield strain (line*age, p = 0.0535). HR mice were more responsive to exercise; older exercised HR mice had shorter tendons (p = 0.0282), and younger exercised HR mice showed lower yield and failure strains than sedentary counterparts (p = 0.0445, 0.0246). Exercise and its relative timing produced slight but complex effects on tendon properties, with HR mice showing the strongest structural and mechanical responses.

Keywords: artificial selection; biomechanics; locomotion; maturation; tendon; wheel running.

FIGURE 1

Experimental design and sample sizes for two lines of mice (HR and control) exposed to exercise at either weanling (3 weeks old) or young adult (9 weeks old) and compared to sedentary mice. Mice from control and high‐runner lines (normal‐/high‐activity) were separated into 2 age cohorts including young (3 weeks) and adult (9 weeks) groups. These cohorts were then separated into two exercise groups (sedentary/wheel) for 8 weeks. Illustration was created with BioRender.com.

FIGURE 2

Tendon testing apparatus with custom‐made wooden rig to hold tendon without direct clamping. (a) With the Achilles tendon insertion intact, the calcaneus was wedged within the conical slot of the wooden rig (see Section 2). The proximal tendon was bound by a silk thread that was attached to the ergometer. (b) Two dots of India ink are visible; the most distal dot was used to calculate tendon length change. The suture knot was tied distally to the muscle‐tendon junction to estimate the proximal end of the tendon.

FIGURE 3

Pooled average weekly wheel running for young (left) and young adult (right) HR (red) and control (blue) mice during 8‐week training. Mean distance run per each training week for young cohort (3‐week‐old training start; a, open bars) and young adult (9‐week‐old training start; b, filled bars) mice from Control (blue) and HR (red) lines. Error bars are ±SEM.

FIGURE 4

Least squares means (LSM) for morphological measures taken from HR and Control mice after the training period beginning as either young (3 weeks old, left side of graphs) or young adult (9 weeks old, right side of graphs). Least squares means (LSM) and standard errors of (a) body mass, (b) gastrocnemius muscle mass, (c) tendon cross‐sectional area, and (d) tendon length from mixed model analyses with Age, Exercise, Line as fixed effects, and body mass as a covariate where appropriate. Control mice are indicated in blue, HR mice in red, and wheel exercised mice are indicated with a filled bar, whereas sedentary cohorts have bars with no fill. Tendon length (d) was significantly affected by the interaction of linetype*exercise (p = 0.0410) such that sedentary HR mice had longer tendons than control line mice and HR wheel access mice. Corresponding statistical analyses are listed within Table 1.

FIGURE 5

Least squares means (LSM) for tendon material properties taken from HR and Control mice after the training period beginning at either young (3 weeks old, left side of graphs) or young adult (9 weeks old, right side of graphs) stages. Least squares means (LSM) of yield strain (a), yield stress (b), failure strain (c), failure stress (d), modulus (e), and work to failure (f) from mixed model analyses with Age, Exercise, Line as fixed effects, and body mass as a covariate. Control mice are indicated in blue, HR mice in red, and wheel exercised mice are indicated with a filled bar while sedentary cohorts have bars with no fill. Yield (a) and failure (b) strains tended to be shorter in older cohort HR mice, and within younger HR mice, wheel access mice had >30% shorter yield and failure strains than sedentary HR mice (Table 1, differences of LSM). Yield stress was significantly greater in HR mice (p = 0.0262), but only approached significance for failure stress (p = 0.0676). Likewise, modulus (e) appeared to have reverse effects of exercise depending on the age cohort, but this interaction did not reach the significance threshold (p = 0.0632). Work to failure (f) was significantly greater in the younger cohort overall (p = 0.0435).