Exercise alters SIRT1, SIRT6, NAD and NAMPT levels in skeletal muscle of aged rats

Abstract

Silent information regulators are potent NAD(+)-dependent protein deacetylases, which have been shown to regulate gene silencing, muscle differentiation and DNA damage repair. Here, changes in the level and activity of sirtuin 1 (SIRT1) in response to exercise in groups of young and old rats were studied. There was an age-related increase in SIRT1 level, while exercise training significantly increased the relative activity of SIRT1. A strong inverse correlation was found between the nuclear activity of SIRT1 and the level of acetylated proteins. Exercise training induced SIRT1 activity due to the positive effect of exercise on the activity of nicotinamide phosphoribosyltransferase (NAMPT) and thereby the production of sirtuin-fueling NAD(+). Exercise training normalized the age-associated shift in redox balance, since exercised animals had significantly lower levels of carbonylated proteins, expression of hypoxia-inducible factor-1 alpha and vascular endothelial growth factor. The age-associated increase in the level of SIRT6 was attenuated by exercise training. On the other hand, aging did not significantly increase the level of DNA damage, which was in line with the activity of 8-oxoguanine DNA glycosylase, while exercise training increased the level of this enzyme. Regular exercise decelerates the deleterious effects of the aging process via SIRT1-dependent pathways through the stimulation of NAD(+) biosynthesis by NAMPT.

Fig. 1

Changes in cytosolic levels and activities of SIRT1 as a function of age and physical exercise. (A) Aging increases the levels of SIRT1. (B) Effect of exercise on SIRT1 levels in young and aged group of animals. Densities of the bands were normalized to tubulin which served as an internal control. (C) The graphical representation of SIRT1 activity. In panels (A)–(C): YC, young control; YE, young exercised; OC, old control; OE, old exercised. Values are means ± S.E. for six animals per group; ^*p < 0.05, ^**p < 0.01.

Fig. 2

Change in nuclear SIRT1 levels (A panel) and activities (C panel) in muscles from young and aged animals. (B) Impact of exercise on nuclear levels of SIRT1. Densities of the bands were normalized to lamin A which served as an internal control of nucleus. In panels (A)–(C): YC, young control; YE, young exercised; OC, old control; OE, old exercised. Values are means ± S.E. for six animals per group; ^*p < 0.05, ^**p < 0.01.

Fig. 3

Decreases in NAD⁺ and NAMPT levels were re-established by exercise training in aged animals. (A) Changes in the levels of NAD⁺ biosynthetic enzyme NAMPT. (B) Exercise increases NAMPT level in muscles of young and aged animals. Densities of the bands were normalized to tubulin which served as an internal control. (C) Exercise training prevented the age-dependent decrease in NAD⁺ level. In panels (A)–(C): YC, young control; YE, young exercised; OC, old control; OE, old exercised. Values are means ± S.E. for six animals per group; ^*p < 0.05, ^**p < 0.01.

Fig. 4

Change in levels of acetylated proteins in nuclear lysates. (A) Graphical deceptions of acetylated protein levels in age and exercise groups. (B) Abundance of acetylated proteins in nuclear lysates from muscle. Densities of the bands were normalized to lamin A which served as an internal control of nucleus. In panels (A) and (B): YC, young control; YE, young exercised; OC, old control; OE, old exercised. Values are means ± S.E. for six animals per group; ^*p < 0.05.

Fig. 5

UCP3 levels in response to exercise in young and aged animals. (A) Graphical illustration of results from densitometry. (B) Changes at protein levels for UCP3 in muscles of control and exercised group of animals. Densities of the bands were normalized to tubulin which served as an internal control. In panel (A) and (B): YC, young control; YE, young exercised; OC, old control; OE, old exercised. Values are means ± S.E. for six animals per group; ^*p < 0.05.

Fig. 6

Change in levels of carbonylated proteins in response to age and exercise. (A) Graphical illustration of results from densitometry. (B) Representative auto radiogram of carbonylated proteins from muscles of control and exercised animals. Densities of the bands were normalized to tubulin which served as an internal control. In panels (A) and (B): YC, young control; YE, young exercised; OC, old control; OE, old exercised. Values are means ± S.E. for six animals per group; ^**p < 0.01.

Fig. 7

OGG1 and SIRT6 levels and activities in resting and exercised young and aged animals. (A) Representative auto radiogram of OGG1 level. Densities of the bands were normalized to tubulin which served as an internal control. (B) Representative auto radiogram of OGG1 activity. (C) Graphical illustration of results from densitometry of SIRT6 level. (D) Representative auto radiograms from aged and/or exercised animals. In panel (A)–(D): YC, young control; YE, young exercised; OC, old control; OE, old exercised. Values are means ± S.E. for six animals per group; ^**p < 0.01.

Fig. 8

Aging increased the level of HIF-1α (A and B) and VEGF (C and D) and these increases were significantly attenuated by exercise training. Densities of the bands were normalized to tubulin which served as an internal control. In panels (A)–(D): YC, young control; YE, young exercised; OC, old control; OE, old exercised. Values are means ± S.E. for six animals per group; ^**p < 0.01.