Sex differences in endurance exercise capacity and skeletal muscle lipid metabolism in mice

Abstract

Previous studies suggest that sex differences in lipid metabolism exist with females demonstrating a higher utilization of lipids during exercise, which is mediated partly by increased utilization of muscle triglycerides. However, whether these changes in lipid metabolism contribute directly to endurance exercise performance is unclear. Therefore, the objective of this study was to investigate the contribution of exercise substrate metabolism to sex differences in endurance exercise capacity (EEC) in mice. Male and female C57BL/6-NCrl mice were subjected to an EEC test until exhaustion on a motorized treadmill. The treadmill was set at a 10% incline, and the speed gradually increased from 10.2 m/min to 22.2 m/min at fixed intervals for up to 2.5 h. Tissues and blood were harvested in mice immediately following the EEC. A cohort of sedentary, non-exercised male and female mice were used as controls. Females outperformed males by ~25% on the EEC. Serum levels of both fatty acids and ketone bodies were ~50% higher in females at the end of the EEC. In sedentary female mice, skeletal muscle triglyceride content was significantly greater compared to sedentary males. Gene expression analysis demonstrated that genes involved in skeletal muscle fatty acid oxidation were significantly higher in females with no changes in genes associated with glucose uptake or ketone body oxidation. The findings suggest that female mice have a higher endurance exercise capacity and a greater ability to mobilize and utilize fatty acids for energy.

Keywords: exercise metabolism; exercise physiology; fatty acid oxidation; ketosis; triglyceride metabolism.

FIGURE 1

Assessment of endurance exercise capacity in male and female mice. Mice were subjected to an endurance exercise capacity (EEC) test on a motorized treadmill. Incline was set to 10% and speeds were adjusted at regular intervals for up to 2 h. Exercise performance was assessed in male and female mice (n = 9 each group) by (a) total duration in minutes and (b) total distance in meters. (c) Serum lactate was measured in serum obtained from blood of male and female mice collected immediately at the conclusion of the EEC test (n = 5 each group). *p < 0.05 vs. males

FIGURE 2

Physical characteristics of male and female mice. (a) Body weight (BW) obtained in male (n = 18) and female (n = 23) mice that participated in the study. (b) Heart weight, (c) quadriceps mass, (d) heart weight to BW and (e) quadriceps to BW ratios in male (n = 9) and female (n = 14) sedentary (non‐exercised) mice. (f) 17‐beta‐estradiol levels measured in the serum obtained from a random sampling of male and female mice (n = 5 each group). *p < 0.05 vs. males

FIGURE 3

Analysis of glucose metabolism in male and female mice. (a) Blood glucose measured in male and female mice at the immediate conclusion of the endurance exercise capacity (EEC) test (n = 9 each group). A cohort of sedentary (non‐exercised) male (n = 9) and female (n = 14) mice were used as controls. Two‐way ANOVA revealed a significant main effect of exercise on blood glucose (F(1,38) = 76.69, p < 0.0001). (b) Glycogen content measured in liver obtained from sedentary and EEC male and female mice (n = 5–7 each group). Two‐way ANOVA revealed a significant main effect of exercise on liver glycogen content (F(1,18) = 72.20, p < 0.0001). (c) Glycogen content measured in gastrocnemius muscle obtained from sedentary and EEC male and female mice (n = 5–7 each group). Two‐way ANOVA revealed a significant main effect of exercise on gastrocnemius glycogen content (F(1,24) = 16.17, p = 0.005). Gene expression analysis of glucose metabolism in (d) Cardiac, (e) Hepatic and (f) Quadriceps muscle tissue harvested from sedentary (non‐exercised) male and female mice (n = 4–5 each group). Glut1, glucose transporter 1; Glut4, glucose transporter 4; Gk, glucokinase; Glut2, glucose transporter 2. *p < 0.05 vs. male or female sedentary group

FIGURE 4

Evaluation of ketone body metabolism in male and female mice. (a) Serum ketone bodies (acetoacetate and 3‐hydroxybutyrate) measured in male and female mice at the immediate conclusion of the endurance exercise capacity (EEC) test (n = 9 each group). Serum obtained from sedentary (non‐exercised) male (n = 9) and female (n = 14) mice were used for control comparison. Two‐way ANOVA revealed a significant main effect of exercise (F(1,36) = 59.86, p < 0.0001) and significant main effect of sex (F(1,24) = 8.563, p = 0.0059) on serum ketone bodies. Gene expression analysis of ketone body metabolism in (b) Hepatic, (c) Cardiac and (d) Quadriceps muscle tissue harvested from sedentary (non‐exercised) male and female mice (n = 4–5 each group). Bdh1, beta‐hydroxybutyrate dehydrogenase 1; Hmgcs2, Hydroxymethylglutaryl‐CoA synthase 2; Acat1, acetyl‐CoA acetyltransferase 1; Oxct1, 3‐oxoacid CoA‐transferase 1. *p < 0.05 vs. male or female sedentary group. #p < 0.05 vs. male EEC group

FIGURE 5

Assessment of lipid metabolism in male and female mice. Serum obtained from sedentary (non‐exercised) male (n = 9) and female (n = 12–13) mice were used for control comparisons. (a) Serum fatty acids in male and female mice at the immediate conclusion of the endurance exercise capacity (EEC) test (n = 9 each group). Two‐way ANOVA revealed a significant main effect of exercise (F(1,35) = 32.20, p < 0.0001) and significant main effect of sex (F(1,35) = 6.044, p = 0.0190) on serum fatty acids. (b) Serum triglycerides in male and female mice at the immediate conclusion of the endurance exercise capacity (EEC) test (n = 9 each group). Two‐way ANOVA revealed a significant interaction effect of exercise and sex (F(1,36) = 4.618, p = 0.0384) and significant main effect of sex (F(1,36) = 16.55, p = 0.0002) on serum triglycerides. (c) Serum cholesterol measured in male and female mice at the immediate conclusion of the endurance exercise capacity (EEC) test (n = 9 each group). Two‐way ANOVA revealed a significant main effect of exercise (F(1,36) = 32.02, p < 0.0001) and significant main effect of sex (F(1,36) = 21.93, p < 0.0001) on serum cholesterol. (d) Triacylglycerol (TAG) content measured in the gastrocnemius muscle obtained from sedentary and EEC male and female mice (n = 7 each group). Two‐way ANOVA revealed a significant main effect of exercise (F(1,24) = 11.70, p = 0.0022) and significant main effect of sex (F(1,24) = 8.783, p = 0.0068) on TAG content. (e) Analysis of genes involved in muscle TAG metabolism from quadriceps muscle tissue (n = 4 each group). Gene expression analysis of fatty acid oxidation in (f) Hepatic, (g) Cardiac and (h) Quadriceps muscle tissue harvested from sedentary (non‐exercised) male and female mice (n = 4–5 each group). *p < 0.05 male or female EEC vs. male or female sedentary group. #p < 0.05 vs. Male EEC group. **p < 0.05 vs. male sedentary group. Dgat1, diacylglycerol acyltransferase 1; Atgl, adipose triglyceride lipase; Pparα, peroxisome proliferator‐activated receptor alpha; Cpt1a, carnitine palmitoyltransferase 1A; Cd36, cluster of differentiation 36; Cpt1b, carnitine palmitoyltransferase 1B; Mcad, medium chain acyl‐CoA dehydrogenase; Lcad, long‐chain acyl‐CoA dehydrogenase