Fig 1.
Lifelong spontaneous exercise (LSE) delays the decline in physical fitness attributed to aging.
(A) Endurance test. (B) Rotarod test. (C) Grip strength test. (D) Habitual spontaneous exercise test. All data are presented as the mean ± standard deviation (SD); vs. control group of same age, *p < 0.05 and ***p < 0.001; vs. control group of different age, ¶p < 0.0001. Partially modified from Fig 3 of a previous study [15].
Fig 2.
Difference in the mass of various skeletal muscles of experimental mice subjected to LSE.
(A) Relative diaphragm weight. (B) Relative gastrocnemius muscle weight. (C) Relative soleus muscle weight. (D) Relative extensor digitorum longus muscle weight. (E) Relative tibialis anterior muscle weight. (F) Body weight. All data are presented as the mean ± SD vs. control group of same age, *p < 0.05 and ***p < 0.001; vs. control group of different age, †p < 0.05, ‡p < 0.01, and ¶p < 0.0001.
Fig 3.
Difference in fiber-cross sectional area (CSA) of skeletal muscles of experimental mice subjected to LSE.
(A) Skeletal muscle tissue microscope slides stained with Hematoxylin & Eosin. (B) Comparison of skeletal muscle fiber CSA between groups. All data are presented as the mean ± SD vs. control group of different age, †p < 0.05.
Fig 4.
Expression of skeletal muscle synthesis-related proteins in mice subjected to LSE.
(A) Western blotting band images (results of skeletal muscle synthesis-related protein analysis). (B) IGF-1 expression level. (C) S6K1 expression level. (D) mTOR expression level. All data are presented as the mean ± SD vs. control group of same age, †p < 0.05 and ‡p < 0.01.
Fig 5.
Expression of angiogenesis-related genes in endothelial cells (ECs) isolated from blood vessels of mice subjected to LSE.
(A) Vegfa expression level. (B) Vegfb expression level. (C) Vegfc expression level. (D) Plgf expression level. (E) Fgf1 expression level. (F) Fgf2 expression level. (G) Tsp1 expression level. (H) Tsp2 expression level. (I) Ang1 expression level. (J) Ang2 expression level. (K) Hgf expression level. (L) Jmjd1a expression level. All data are presented as the mean ± SD vs. control group of same age, **p < 0.01 and ****p < 0.0001; vs. control group of different age, †p < 0.05, ‡p < 0.01, and ¶p < 0.0001. N. S. = no significant difference between groups.
Fig 6.
Expression of angiogenesis-related proteins in blood vessels of mice subjected to LSE.
(A) Western blotting band images (results of angiogenesis-related proteins in carotid artery). (B) VEGFR2 expression level. (C) VEGF expression level. All data are presented as the mean ± SD vs. control group of same age, *p < 0.05 and **p < 0.01; vs. control group of different age, §p < 0.001.
Fig 7.
Inhibitory effect of LSE on the reduction of angiogenic capacity confirmed by ex vivo aortic ring assay.
(A) 10Ⅹ magnification microscopic images of aortic ring assay. (B) Comparison of the number of sprouts extending after culture of isolated mouse aorta. All data are presented as the mean ± SD vs. control group of same age, ***p < 0.001 and ****p < 0.0001; vs. control group of different age, ¶p < 0.0001.
Fig 8.
Comparison of wound healing ability of vascular ECs after senescence simulated by oxidative stress and exercise simulated by laminar shear stress in vitro.
(A) 10Ⅹ magnification microscopic images of scratch wound healing assay. (B) Migratory distance. LSS: Laminar shear stress; OS: Oxidative stress. All data are presented as the mean ± SD, *p < 0.01 and ***p < 0.001.
Fig 9.
Comparison of tube formation ability of vascular ECs after senescence simulated by oxidative stress and exercise simulated by laminar shear stress in vitro.
(A) 10Ⅹ magnification microscopic images of tube formation assay. (B) Number of branch points. (C) Total tube length. (C) Total formation area. LSS: Laminar shear stress; OS: Oxidative stress. All data are presented as the mean ± SD, *p < 0.01 and ***p < 0.001.