Elevated SOCS3 and altered IL-6 signaling is associated with age-related human muscle stem cell dysfunction

Abstract

Aging is associated with increased circulating interleukin-6 (IL-6) and a reduced myogenic capacity, marked by reduced muscle stem cell [satellite cell (SC)] activity. Although IL-6 is important for normal SC function, it is unclear whether elevated IL-6 associated with aging alters SC function. We hypothesized that mild chronically elevated IL-6 would be associated with a blunted SC response through altered IL-6 signaling and elevated suppressor of cytokine signaling-3 (SOCS3) in the elderly. Nine healthy older adult men (OA; 69.6 ± 3.9 yr) and 9 young male controls (YC; 21. 3 ± 3.1 yr) completed 4 sets of 10 repetitions of unilateral leg press and knee extension (75% of 1-RM). Muscle biopsies and blood were obtained before and 3, 24, and 48 h after exercise. Basal SC number was 33% lower in OA vs. YC, and the response was blunted in OA. IL-6(+)/Pax7(+) cells demonstrated a divergent response in OA, with YC increasing to 69% at 3 h and peaking at 24 h (72%), while IL-6(+)/Pax7(+) cells were not increased until 48 h in OA (61%). Type II fiber-associated phosphorylated signal transducer and activator of transcription (pSTAT3)(+)/Pax7(+) cells demonstrated a similar delay in OA, not increasing until 48 h (vs. 3 h in YC). SOCS3 protein was 86% higher in OA. These data demonstrate an age-related impairment in normal SC function that appears to be influenced by SOCS3 protein and delayed induction of IL-6 and pSTAT3 in the SCs of OA. Collectively, these data suggest dysregulated IL-6 signaling as a consequence of aging contributes to the blunted muscle stem cell response.

Fig. 1.

Baseline muscle cross-sectional area (CSA) by fiber-type. Type I muscle fiber area and type II muscle fiber area in young controls (YC; open bar) and older adults (OA; closed bar). *P < 0.05 vs. OA; data are presented as means ± SD.

Fig. 2.

Muscle satellite cell (SC) response to acute resistance exercise is blunted with age. A: number of Pax7⁺ cells (SCs) per 100 myofibers. *Effect of age, P < 0.05 vs. YC. **Effect of time, P < 0.05 vs. baseline measurements (PRE; bar denotes both YC and OA different from PRE at 48 h). B: whole muscle mRNA expression of myogenin in YC (open bars) and OA (closed bars); a.u., arbitrary units. *Effect of time, P < 0.05 vs. PRE (bar denotes both groups are different from PRE). Type I-associated SCs (C) and type II-associated SCs (D) expressed as a percentage of total myonuclei in YC (open bars) and OA (closed bars). *Effect of age, P < 0.05 vs. YC. **Effect of time, P < 0.05 in YC only. Data are presented as baseline (PRE), 3, 24, and 48 h postexercise; means ± SE.

Fig. 3.

IL-6 is elevated in the elderly. A: serum IL-6 concentration in YC (open bars) and OA (closed bars). *Effect of age, P < 0.05 vs. YC. **Effect of time, P < 0.05 vs. PRE in YC. ***Effect of time, P < 0.05 vs. PRE and 3 h (bar denotes effect of time in both YC and OA). B: baseline whole muscle mRNA expression of IL-6 in YC (open bars) and OA (closed bars). *Effect of age, P < 0.05 vs. YC. C: whole muscle mRNA expression of IL-6 through the study time course in YC (open bars) and OA (closed bars). *Effect of age, P < 0.05 vs. YC. **Effect of time, P < 0.05 vs. PRE (bar denotes effect in both groups). ***Effect of time, P < 0.05 vs. PRE and 3 h in OA only. ****Effect of time, P < 0.05 vs. all other time points in OA only. D: whole muscle mRNA expression of IL-6 in YC only to illustrate the effect of time (fold change from PRE). *P < 0.05 vs. PRE. All data are presented as baseline (PRE), 3, 24, and 48 h postexercise; means ± SE.

Fig. 4.

SC-specific IL-6 is differentially regulated with age. A: immunofluorescent image of a muscle cross section stained for Pax7 (red), laminin (orange), IL-6 (green), and nuclei (DAPI, blue). Various panels illustrate an IL-6⁺/Pax7⁺ cell (long arrow) and an IL-6⁻/Pax7⁺ cell (arrowhead). B: quantification of the proportion of SCs staining positive for IL-6 (as a percentage of total SC number) in YC (open bars) and OA (closed bars). *Effect of age, P < 0.05 vs. YC. **Effect of time for YC, P < 0.05 vs. PRE and 48 h. ***Effect of time for OA, P < 0.05 vs. PRE, 3, and 24 h. C: proportion of total SCs (black) positive of IL-6 (grey) in OA across time. **Effect of time, P < 0.05 vs. PRE for both IL-6 and Pax7. D: proportion of total SCs (black) positive of IL-6 (grey) in YC across time. *Effect of time for IL-6 at 3 h, P < 0.05 vs. PRE. **Effect of time, P < 0.05 vs. PRE for both IL-6 and Pax7. ***Effect of time, P < 0.05 vs. PRE for Pax7 only (bar denotes effect in both groups). All data are presented as baseline (PRE), 3, 24 and 48 h postexercise; means ± SE.

Fig. 5.

Fiber-type specific phosphorylated signal transducer and activator of transcription (pSTAT3) response is altered with age. A: immunofluorescent image of a muscle cross section stained for Pax7 (red), laminin (green), myosin heavy chain type-I (MYH1, green), phosphorylated STAT3 (pSTAT3, orange), and nuclei (DAPI, blue). Various panels illustrate a type I-associated pSTAT3⁺/Pax7⁺ cell (closed arrow) and a type II-associated pSTAT3⁺/Pax7⁺ cell (open arrow). Type I SC specific pSTAT3 (B) and Type II SC specific pSTAT3 (C) response in YC (open bars) and OA (closed bars). *Effect of age, P < 0.05 vs. YC. **Effect of time, P < 0.05 vs. PRE. ***Effect of time, P < 0.05 vs. PRE and 3 h in YC only (bar denotes effect in both groups). All data are presented as baseline (PRE), 3, 24, and 48 h postexercise; means ± SE.

Fig. 6.

pSTAT3 gene targets are differentially expressed with age. Whole muscle mRNA expression of cMyc (A), suppressor of cytokine signaling-1 (SOCS1; B), baseline SOCS3 (C), and SOCS3 (D) in YC (open bars) and OA (closed bars). *Effect of age, P < 0.05 vs. YC. **Effect of time, P < 0.05 vs. PRE. ***Effect of time, P < 0.05 vs. PRE and 3 h (bar denotes effect in both groups). All data are presented as baseline (PRE), 3, 24, and 48 h postexercise; means ± SE.

Fig. 7.

SOCS1 and SOCS3 protein content is elevated in elderly muscle. SOCS1 protein (A), baseline SOCS3 protein (B), and SOCS3 protein content (C) in YC (open bars) and OA (closed bars) with the associated representative blots. *Effect of age, P < 0.05 vs. YC. **Effect of time, P < 0.05 vs. 3 h. All data are presented as baseline (PRE), 3, 24, and 48 h postexercise; means ± SE.