(-)-Epicatechin enhances fatigue resistance and oxidative capacity in mouse muscle

Abstract

The flavanol (-)-epicatechin, a component of cacao (cocoa), has been shown to have multiple health benefits in humans. Using 1-year-old male mice, we examined the effects of 15 days of (-)-epicatechin treatment and regular exercise on: (1) exercise performance, (2) muscle fatigue, (3) capillarity, and (4) mitochondrial biogenesis in mouse hindlimb and heart muscles. Twenty-five male mice (C57BL/6N) were randomized into four groups: (1) water, (2) water-exercise (W-Ex), (3) (-)-epicatechin ((-)-Epi), and (4) (-)-epicatechin-exercise ((-)-Epi-Ex). Animals received 1 mg kg(-1) of (-)-epicatechin or water (vehicle) via oral gavage (twice daily). Exercise groups underwent 15 days of treadmill exercise. Significant increases in treadmill performance (∼50%) and enhanced in situ muscle fatigue resistance (∼30%) were observed with (-)-epicatechin. Components of oxidative phosphorylation complexes, mitofilin, porin, nNOS, p-nNOS, and Tfam as well as mitochondrial volume and cristae abundance were significantly higher with (-)-epicatechin treatment for hindlimb and cardiac muscles than exercise alone. In addition, there were significant increases in skeletal muscle capillarity. The combination of (-)-epicatechin and exercise resulted in further increases in oxidative phosphorylation-complex proteins, mitofilin, porin and capillarity than (-)-epicatechin alone. These findings indicate that (-)-epicatechin alone or in combination with exercise induces an integrated response that includes structural and metabolic changes in skeletal and cardiac muscles resulting in greater endurance capacity. These results, therefore, warrant the further evaluation of the underlying mechanism of action of (-)-epicatechin and its potential clinical application as an exercise mimetic.

Figures

Figure 1. Plantaris muscle comparison of NCAF, C/Fi (B), capillary density (C) and capillary-to-fibre perimeter exchange (CFPE) (D) between the four groups

Data are means ± SEM. *P < 0.05; †significantly different from W-Ex group; n = 3 per group.

Figure 2. Relative isometric tension (P/P0) obtained at different frequencies of stimulation from EDL muscles from non-exercised mice (open and filled triangles) or exercised mice (open and filled circles) that were fed with (–)-epicatechin (open symbols) or in the control mice (filled symbols)

*P < 0.01, water vs. W-Ex; **P < 0.01, (–)-Epi vs. (–)-Epi-Ex. Data are means ± SEM, n = 6 muscles.

Figure 3. A, representative tension recordings, from the EDL muscle, during the fatigue run from the non-exercised group (left panel) and from the exercised group (right panel) normalized to the initial tension. B, results of fatigue run for all four groups

Data are means ± SEM, n = 6 muscles. *P < 0.05.

Figure 4. A, mitochondrial protein complexes for the quadriceps femoris muscle

*Significant differences between groups; **significant differences between W-Ex, (–)-Epi, and (–)-Epi-Ex versus W group; †significantly different from corresponding complex for (–)-Epi group. B, mitofilin and porin proteins. **Significant differences between W-Ex, (–)-Epi, and (–)-Epi-Ex versus water group for mitofilin and porin. Data are means ± SEM, n = 3 per group.

Figure 5. A, mitochondrial protein complexes for the cardiac muscle

*Significantly different from corresponding complex for W group; #significantly different with corresponding complex for water group; ‡significantly different from corresponding complex for W-Ex group; **significant differences between W-Ex, (–)-Epi, and (–)-Epi-Ex versus W group; †significantly different from corresponding complex of (–)-Epi group. B, mitofilin and porin protein. **Significant differences between W-Ex, (–)-Epi, and (–)-Epi-Ex versus water group for mitofilin and porin. Data are means ± SEM, n = 3 per group.

Figure 6. Representative Western blot results for nNOS, p-nNOS and p-nNOS/nNOS ratio (A) and Tfam (B) from quadriceps femoris muscle

*P < 0.05; †significantly different from water only group for p-nNOS and nNOS.

Figure 7. Mitochondrial volume density for the plantaris muscle (A), and cristae membrane surface area normalized to the outer membrane surface area (B)

Data are means and SEM. The representative electron microscopy images of control (left side) and (–)-epicatechin (right side) treated muscle above each graph. For panel A, the scale bar represents 2 μm, whereas for panel B, the scale bar represents 200 nm. *P < 0.05.

Figure 8. Mitochondrial volume density for the heart muscle (A), and cristae membrane surface area normalized to the outer membrane surface area (B)

Data are means and SEM. The representative electron microscopy images of control (left side) and (–)-epicatechin (right side) treated muscle above each graph. For panel A, the scale bar represents 4 μm, whereas for panel B, the scale bar represents 400 nm. *P < 0.05.