Endurance exercise training reinforces muscular strength with improvements in mitochondrial oxidative capacity, lysosome reformation, and myogenic differentiation against doxorubicin-induced skeletal muscle wasting in mice

Abstract

Purpose: Doxorubicin (DOX) is a chemotherapeutic medication broadly used to treat diverse cancers. However, chronic DOX chemotherapy can cause myotoxicity and muscle atrophy. Endurance exercise (EXE) is used to prevent negative muscle excitation. Based on emerging evidence, this study investigated the challenges that occur in skeletal muscle quantity, quality, and metabolic determinants through autophagy, myogenic regulatory factors (MRF), antioxidant enzymes, and both the AMPK and AKT/mTOR pathways.

Methods: Male C57BL/6J adult mice were divided into four groups after one week of acclimation: sedentary (SED) plus saline (SAL)-receiving (SED-SAL), EXE plus SAL-receiving (EXE-SAL), SED plus DOX-receiving (SED-DOX), and EXE plus DOX-receiving (EXEDOX) groups. All mice were intraperitoneally inoculated with either SAL or DOX (5 mg/kg, every 2 weeks) for 8 weeks, while a treadmill running EXE was performed. Body weight, muscle weight, and muscle strength were measured, and the red portions of the gastrocnemius muscle were excised for biochemical analysis.

Results: Chronic DOX administration deteriorated body composition by decreasing body and absolute muscle weights, whereas EXE reinforced a grip strength per body weight. Although DOX inhibited BECN1 expression, EXE enhanced CS, LC3-I, LC3-II, and LAMP levels. Moreover, DOX did not interrupt MRF functions, but EXE improved MYOD without altering SOD1 or SOD2 expression. However, neither the AMPK nor the AKT/mTOR signaling pathways were associated with either DOX-receiving or EXE training.

Conclusion: DOX chemotherapy-induced muscle wasting is associated with autophagy dysregulation. However, long-term aerobic EXE training enhances muscular strength with an increase in mitochondrial oxidative capacity, lysosome formation, and myogenic differentiation.

Keywords: autophagy dysregulation; cancer chemotherapy; exercise adaptation; grip strength; myogenic regulatory factors; skeletal muscle atrophy.

Figure 1.

Skeletal muscle strength test in mice. (A) A representative image for the grip strength test, (B) quantification of grip strength in each group, and (C) The ratio of grip strength/body weight in each group. All mice were assigned seven per group. SAL, saline; DOX, doxorubicin; SED, sedentary; EXE, exercise training. Significant differences are denoted by an asterisk for p < 0.01(**) and p < 0.001(^***). Data are presented as mean ± SEM.

Figure 2.

Effects of endurance exercise training-induced adaptations during DOX chemotherapy on the key regulators of autophagy in skeletal muscle. (A) Representative western blot images are shown on the left side of the graph. Quantifications of (B) CS, (C) BECN1, (D) ATG 7, (E) p62, (F) LC3B-II, LC3B-I, and LC3B II/I ratio, (G) LAMP2, and (H) CTSL protein levels. All target proteins were normalized by Ponceau-stained total proteins (n=6-7 mice per group). SAL, saline; DOX, doxorubicin; SED, sedentary; EXE, exercise training. CS, Citrate synthase; BECN1, Beclin-1; LAMP2, Lysosome-associated membrane protein 2; CTSL, Cathepsin L.

Figure 3.

Effects of endurance exercise training-induced adaptations during DOX chemotherapy on myogenic regulatory factors (MRF) and antioxidant enzymes in skeletal muscle. (A) Representative western blot images are shown on the left side of the graph. Quantifications of (B) PAX7, MYOD, and MYOG, (C) SOD1, and (D) SOD2 protein levels. All target proteins were normalized by Ponceau-stained total proteins (n=6-7 mice per group). SAL, saline; DOX, doxorubicin; SED, sedentary; EXE, exercise training. PAX 7, paired box protein 7; MYOD, myogenic differentiation 1; MYOG, myogenin; SOD1, superoxide dismutase 1; SOD2, superoxide dismutase 2.

Figure 4.

Effects of endurance exercise training-induced adaptations during DOX chemotherapy on metabolic determinants in skeletal muscle (A) Representative western blot images are shown on the left side of the graph. Quantifications of (B) p-AMPK^Thr172, AMPK, and p-AMPK^Thr172/AMPK ratio, (C) p-AKT^Thr308, AKT, and p-AKT^Thr308/AKT ratio, (D) p-AKT^Ser473, AKT, and p-AKT^Ser473/AKT ratio, (E) p-mTOR^Ser2448, mTOR, and p-mTOR^Ser2448/ mTOR ratio. All target proteins were normalized by Ponceau-stained total proteins (n=6-7 mice per group). SAL, saline; DOX, doxorubicin; SED, sedentary; EXE, exercise training. AMP-activated protein kinase, AMPK; Mammalian target of rapamycin, mTOR.