Aging alters macrophage properties in human skeletal muscle both at rest and in response to acute resistance exercise

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Abstract

Macrophages are involved in skeletal muscle repair through pro-inflammatory and alternative functions. We tested the hypothesis that aging alters the abundance and properties of skeletal muscle macrophages that will influence their functional response to acute resistance exercise. Total macrophages (CD68+), as well as pro- (CD11b+) and anti-inflammatory (CD163+) subpopulations and associated cytokine mRNAs were quantified in vastus lateralis biopsies from young (N=17) and elderly (N=17) males pre- and 72 h post-exercise. Pre-exercise, young muscle tended to possess a greater number of macrophages, whereas elderly muscle possessed higher levels of IL-1β (P=0.001), IL-1RA (P=0.003), and IL-10 (P=0.028). Post-exercise, total macrophages did not change in either group, however, the number of CD11b+ (P=0.039) and CD163+ (P=0.026) cells increased 55 and 29%, respectively, but only in the young. IL-1β (P=0.006), IL-10 (P=0.016), and AMAC-1 (P=0.044) also increased, approximately two-fold, and again only in the young. Quantitation of CD11b+ and CD163+ cells suggests that the majority of resident macrophages possess alternative functions, and a small subpopulation participates in the inflammatory response. Both subpopulations increased their activity post-exercise, exclusively in the young. These findings suggest that aging results in a defective regulation of muscle macrophage function, both at baseline and in response to resistance exercise, that may limit muscle hypertrophy in older adults.

Introduction

The developing model for repair of damaged skeletal muscle includes immune cell functions, which mediate progression from an inflammatory response to the development of an environment that promotes myogenic differentiation (Tidball, 2005). After muscle damage, systemic increases occur for pro-inflammatory cytokines interleukin (IL)-1β and tumor necrosis factor (TNF)-α, the anti-inflammatory cytokines IL-1 receptor antagonist (IL-1RA) and IL-10, and pleiotropic IL-6 (Ostrowski et al., 1999; Pedersen, 2000; Smith et al., 2000). Many of these factors are likely derived from resident macrophages that initially respond according to their classical function by producing chemoattractants and other pro-inflammatory cytokines (Fielding et al., 1993; Warren et al., 2002) resulting in increased proteolysis and an influx of macrophages from the circulation to phagocytize cellular debris (Fielding and Evans, 1997; McLennan, 1996). However, not all muscle macrophages are phagocytic (St Pierre and Tidball, 1994). Subpopulations may perform alternative functions through the production of anti-inflammatory, anti-apoptotic, and growth factors necessary for an environment in which satellite cells can repair the myofiber (Chazaud et al., 2003). This model proposes that proper regulation of macrophage functions following repeated muscle damage contributes to skeletal muscle adaptation and hypertrophy.

The study of muscle aging, and associations between cytokine activities and skeletal muscle loss, illustrate the importance of proper regulation within this model of muscle regeneration. Aging is associated with progressive muscle wasting and low-grade systemic increases in cytokines such as IL-6 and TNF-α (Visser et al., 2002). Higher systemic cytokine levels are associated with functional decline and often cachectic disease (Bautmans et al., 2005; Reuben et al., 2002), however, some counter-regulatory elements, including IL-6 and possibly IL-1RA and IL-10, remain responsive in muscle during aging and can be stimulated by exercise (Petersen and Pedersen, 2005). Regulation of inflammation and the inflammatory response may also be impaired locally in aged muscle. TNF-α expression was elevated within the muscle of frail elderly at baseline, but could be modulated with exercise (Greiwe et al., 2001). Our work has shown that, unlike for young skeletal muscle, resistance exercise does not induce IL-1β gene expression in the elderly (Jozsi et al., 2001). Thus, multiple regulatory defects occur with aging in skeletal muscle that alter cytokine expression both at rest and in response to resistance exercise that could diminish the ability to maintain and/or increase muscle mass in older individuals.

Elucidation of the role of the inflammatory response in skeletal muscle repair has been predominantly derived from the results of human exercise or animal protocols specifically designed to induce extensive muscle damage. Macrophage function and the inflammatory response in skeletal muscle after less damaging hypertrophic stimuli has not been defined nor examined in the elderly. Clusters of differentiation antigens (CD), in addition to the cytokines described above, are available to classify macrophages according to their function. CD68 is a pan marker for all monocytes and macrophages (Holness and Simmons, 1993). The CD11b marker indicates classical pro-inflammatory activation, whereas expression of CD163 and the alternative macrophage activation-associated CC-chemokine (AMAC)-1 are related to anti-inflammatory properties (Boven et al., 2004; Gratchev et al., 2001; Mosser, 2003). CD115 is an indicator of proliferation and differentiation within these cells (Hume et al., 1997). We utilized all of these markers, as well as cytokine expression, to test the hypothesis that aging alters the abundance and properties of skeletal muscle macrophages that will influence their functional response to acute resistance exercise thereby altering their potential contribution to the mechanisms of skeletal muscle hypertrophy. The total number of macrophages and their pro- and anti-inflammatory subpopulations, as well as related cytokine expression, in muscle from young and elderly subjects were quantified before and after exercise. Our results indicate that the number of macrophages within skeletal muscle from the elderly is decreased and their functional properties show defects both at rest and in response to resistance exercise, which could contribute mechanistically to age-related muscle loss.

Section snippets

Subjects

Young and elderly males (Table 1) were recruited and gave a written informed consent in accordance with the declaration of Helsinki and approved by the Institutional Review Board of the University of Arkansas for Medical Sciences. The young muscle samples were from a subset of subjects that were previously analyzed based upon their interleukin-1 genotype (Dennis et al., 2004). The subset was randomly chosen from the sampling so as to best match the genetic makeup of the general population. The

Physical characteristics

The young and elderly subject groups were not different for anthropometric measurements, however, the young achieved significantly greater 1-RMs for two of three leg exercises (Table 1). There was no difference in height, weight, or body mass index between the young (31.9±7.1 years) and elderly (71.4±4.6 years). While the 1-RMs for leg press were similar (P=0.92), the young subjects had significantly higher 1-RM for leg curls (P=0.0002) and extensions (P=0.0002) (Table 2).

Markers of macrophage function

In order to, determine

Discussion

The main finding of this investigation is that the majority of macrophages resident to skeletal muscle display anti-inflammatory characteristics at rest, these numbers increase in response to resistance exercise, and that a relatively small subpopulation dictates the pro-inflammatory response. The elderly possessed diminished numbers of both macrophage subpopulations, and defects in their functional regulation were apparent both at baseline and in response to the exercise stimulus. This work

Acknowledgements

This research was supported in part by funds provided to the UAMS Microarray Facility through Act 1 of The Arkansas Tobacco Settlement Proceeds Act of 2000; by National Center for Research Resources grants through the BRIN Program (P20 RR-16460) and the General Clinical Research Center at the University of Arkansas for Medical Sciences (M01-RR14288); and by grants from the National Institute on Aging, to CAP (AG012411).

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