Moderate-intensity treadmill exercise training decreases murine cardiomyocyte cross-sectional area

Abstract

The aim of this study was to examine the impact of moderate-intensity treadmill exercise on the structure and function of the murine heart and its associated impact on Akt-AMPK-mTOR signaling. A secondary aim was to test whether the exercise phenotype was altered following a cardiotoxic bolus dose of doxorubicin (DOX). Two-month-old C57Bl/6J female mice remained sedentary (SED, n = 12) or were progressively trained with treadmill running for 2 months up to 18 m/min; 60 min/day, 5 days/weeks (EX, n = 11) or EX + DOX (15 mg/kg/dose) (EX + DOX, n = 6). Following treadmill training, mice underwent graded exercise tolerance testing and echocardiography. Training improved graded exercise tolerance by 68 ± 5% relative to SED, and this effect was not altered with bolus DOX. There were no changes in relative heart size with EX or EX + DOX versus SED. Regional posterior wall thickening was improved in EX and abrogated in EX + DOX. EX had a reduced cardiomyocyte cross-sectional area (CSA) relative to SED, and CSA was further attenuated with DOX. Following EX, AMPK-associated phosphorylation of ULK1(ser317) tended to be lower relative to SED. Akt-associated phosphorylation of TSC2(thr1462) and mTOR(ser2448) were also decreased relative to SED. We observed an increase in AMPK activity with DOX that was not translated to downstream AMPK phosphorylation sites. We conclude that 2 months of moderate treadmill exercise training improves regional cardiac function and exercise capacity, but does not induce relative physiologic hypertrophy in female mice. Differential responses in Akt-AMPK-mTOR signaling may mediate the observed phenotype.

Keywords: Anthracycline; echocardiography; exercise training; heart; murine.

Figure 1

Maximal exercise testing. C57BL/6J female mice were exercise trained over 2 months prior to investigating cardiac function and cellular signaling. Maximal treadmill exercise tests (speed starting at 10 m/min with 3 m/min increases every 3 min until exhaustion) were performed at baseline and after training (final) in a subset of animals: sedentary (SED, n = 6), exercise trained (EX, n = 5), and EX + doxorubicin (DOX, n = 6) treated animals (15 mg/kg bolus following exercise training program). A significant increase in work was seen at final testing in EX and EX + DOX groups (A), and the percent change in work from baseline levels was also significantly (*P < 0.001) higher in EX and EX + DOX animals relative to SED animals (B).

Figure 2

Histological analysis of cardiomyocyte cross sectional area and cardiac fibrosis. Representative images of cardiomyocyte cross sectional area and quantification (A); and representative images of cardiac fibrosis and quantification (B), demonstrated that cardiomyocyte cross sectional area was significantly decreased with exercise (EX, n = 5) and doxorubicin (DOX, n = 6) exacerbated this effect (*P < 0.001) compared to sedentary (SED, n = 6) animals. However, cardiac fibrosis was not different between groups.

Figure 3

Western blotting analysis of cardiac AMPK-mTOR-autophagy signaling. Sedentary (SED, n = 3), exercise trained (EX, n = 3), and EX mice given a bolus injection of doxorubicin (DOX, n = 3) revealed hypo-phosphorylation patterns of AMPK phosphorylation sites following EX training. Additionally, increased AMPK activity with DOX did not result in downstream increased phosphorylation patterns of AMPK phosphorylation sites following DOX treatment. *P < 0.05.

Figure 4

Western blotting analysis of cardiac Akt-mTOR-autophagy signaling. Sedentary (SED, n = 3), exercise trained (EX, n = 3), and EX mice given a bolus injection of doxorubicin (DOX, n = 3) mice revealed while Akt signaling was not altered, there was significant hypo-phosphorylation at TSC2 and mTOR phosphorylation sites specific for Akt. DOX did not cause any significant differences in Akt mediated signaling. *P < 0.05.