Regular exercise increases ET zebrafish’s muscle fiber CSA and improves skeletal muscle arrangement.

(A) Zebrafish body weight (g), n = 25. (B) Skeletal muscle mass to body weight (%), n = 25. (C) HE staining and TEM imaging of skeletal muscle cross-section. (D) Skeletal muscle fiber’s CSA statistics were produced using ImageJ, n = 3. (E) Frequency distribution of muscle fiber area (%). Data are presented as mean ± SD; *P<0.05, **P<0.01, ***P<0.001.

Regular exercise improves locomotion ability in ET zebrafish.

(A) Zebrafish Ucrit (cm/s). (B) Zebrafish ucrit-r (bl/s). (C) Zebrafish MO₂max (mmol/kg/h). Data are presented as mean ± SD; n = 25; *P<0.05, ***P<0.001.

Regular exercise suppresses elevated UPS and ALS in the skeletal muscle of ET zebrafish.

(A) mRNA expression of UPS and ALS-related genes in skeletal muscle, n = 6. (B) Murf1 and Fbxo32 protein blotting and quantitative analysis, n = 3. (C) Beclin1 and P62 protein levels in skeletal muscle, n = 3. Data are presented as mean ± SD, *P<0.05, **P<0.01, ***P<0.001.

Regular exercise improves activity and content in MRC complexes of skeletal muscle in ET zebrafish.

(A) Oxygen consumption curves and mitochondrial oxygen flux as they vary over time following the introduction of different substrates or inhibitors. (B) Oxygen consumption rates (OCR) of mitochondrial complex I, as well as the combined rates of complex I and II, and the maximal activity of the electron transport chain, n = 6. (C) mRNA expression levels of subunits of MRC complexes I (ndufa9a), II (sdha), III (uqcrc2b), IV (cox5aa), and V (atp5f1b), n = 6. (D) Protein levels of subunits of MRC complexes I (Ndufb8), II (Sdhb), III (Uqcrc2), and V (Atp5a1), n = 3. Data are presented as mean ± SD; *P<0.05, **P<0.01, ***P<0.001.

Regular exercise improves ethanol-induced oxidative stress and UPRmt.

(A) DHE staining the muscle tissue. (B) Quantitative analysis of skeletal muscle ROS, n = 3. (C) The mRNA expression of skeletal muscle oxidase nox2, nox4, and antioxidant enzyme sod2, n = 6. (D) Protein levels of oxidase Nox2, Nox4, n = 3. (E) UPRmt-related genes mRNA expression levels of lonp1, clpp, hspd1, and hspa9, n = 6. Data are presented as mean ± SD; *P<0.05, **P<0.01, ***P<0.001.

Regular exercise improves mitochondrial morphology and regulates mitochondrial dynamics in ET zebrafish.

(A) TEM imaging of muscle tissue mitochondria. (B) mRNA expression levels of genes related to mitochondrial dynamics, n = 6. (C) Western blot of mitochondrial fission-related proteins’ banding and quantitative analysis, n = 3. (D) Western blot of banding and quantitative analysis of mitochondrial fusion-related proteins, n = 3. Data are presented as mean ± SD;*P<0.05, **P<0.01, ***P<0.001.

Regular exercise upregulates PGC1α/NRF2 signaling in ET zebrafish.

(A) Expression levels of pgc1α and nrf2 mRNA in zebrafish, n = 6. (B) Western blot and quantitative analysis of Pgc1α and Nrf2 in skeletal muscle, n = 3. Data are presented as mean ± SD; *P<0.05, **P<0.01, ***P<0.001.