Effects of Aging, Long-Term and Lifelong Exercise on the Urinary Metabolic Footprint of Rats

Abstract

Life expectancy has risen in the past decades, resulting in an increase in the number of aged individuals. Exercise remains one of the most cost-effective treatments against disease and the physical consequences of aging. The purpose of this research was to investigate the effects of aging, long-term and lifelong exercise on the rat urinary metabolome. Thirty-six male Wistar rats were divided into four equal groups: exercise from 3 to 12 months of age (A), lifelong exercise from 3 to 21 months of age (B), no exercise (C), and exercise from 12 to 21 months of age (D). Exercise consisted in swimming for 20 min/day, 5 days/week. Urine samples collection was performed at 3, 12 and 21 months of life and their analysis was conducted by liquid chromatography-mass spectrometry. Multivariate analysis of the metabolite data did not show any discrimination between groups at any of the three aforementioned ages. However, multivariate analysis discriminated the three ages clearly when the groups were treated as one. Univariate analysis showed that training increased the levels of urinary amino acids and possibly protected against sarcopenia, as evidenced by the higher levels of creatine in the exercising groups. Aging was accompanied by decreased levels of urinary amino acids and signs of increased glycolysis. Concluding, both aging and, to a lesser degree, exercise affected the rat urinary metabolome, including metabolites related to energy metabolism, with exercise showing a potential to mitigate the consequences of aging.

Keywords: aging; exercise; liquid chromatography-mass spectrometry (LC/MS); urinary metabolites.

Figure 1

PCA scores plots of urine samples from young (3-month old, red dots), middle-aged (12-month old, green dots) and old rats (21-month old, black dots), normalized by total ion current (a) and creatinine (b). Ellipse represents 95% confidence intervals of Hotelling’s T² distribution.

Figure 2

(a) PCA scores plot between 3 (red) and 12 months (green), (b) corresponding PLS-DA scores plot, (c) corresponding permutations plot, (d) PCA scores plot between 3 (red) and 21 months (green) (e) corresponding PLS-DA scores plot, (f) corresponding permutations plot, (g) PCA scores plot between 12 (red) and 21 months (green), (h) corresponding PLS-DA scores plot, (i) corresponding permutations plot.

Figure 3

PLS-DA scores plot of urine samples at 12 months of age between the groups that had exercised (A and B, green) and the groups that had not exercised during the 1st half (C and D, red) with permutations plot as insert.

Figure 4

Map of metabolic pathways with highlighted urinary metabolites that changed with age and/or exercise training in (a) the exercising groups during the 1st half, (b) the non-exercising groups during the 1st half, (c) the exercising groups during the 2nd half, and (d) the non-exercising groups during the 2nd half of life. Metabolites in red, blue, or black increased, decreased, or did not change, respectively. Metabolites in grey were not identified.

Figure 5

Study design showing the interventions on the four groups of rats. Solid bars denote exercise training, open bars denote no exercise, and arrowheads denote urine sampling.