Testosterone supplementation upregulates androgen receptor expression and translational capacity during severe energy deficit

Abstract

Testosterone supplementation during energy deficit promotes whole body lean mass accretion, but the mechanisms underlying that effect remain unclear. To elucidate those mechanisms, skeletal muscle molecular adaptations were assessed from muscle biopsies collected before, 1 h, and 6 h after exercise and a mixed meal (40 g protein, 1 h postexercise) following 14 days of weight maintenance (WM) and 28 days of an exercise- and diet-induced 55% energy deficit (ED) in 50 physically active nonobese men treated with 200 mg testosterone enanthate/wk (TEST) or placebo (PLA) during the ED. Participants (n = 10/group) exhibiting substantial increases in leg lean mass and total testosterone (TEST) were compared with those exhibiting decreases in both of these measures (PLA). Resting androgen receptor (AR) protein content was higher and fibroblast growth factor-inducible 14 (Fn14), IL-6 receptor (IL-6R), and muscle ring-finger protein-1 gene expression was lower in TEST vs. PLA during ED relative to WM (P < 0.05). Changes in inflammatory, myogenic, and proteolytic gene expression did not differ between groups after exercise and recovery feeding. Mechanistic target of rapamycin signaling (i.e., translational efficiency) was also similar between groups at rest and after exercise and the mixed meal. Muscle total RNA content (i.e., translational capacity) increased more during ED in TEST than PLA (P < 0.05). These findings indicate that attenuated proteolysis at rest, possibly downstream of AR, Fn14, and IL-6R signaling, and increased translational capacity, not efficiency, may drive lean mass accretion with testosterone administration during energy deficit.

Keywords: androgen receptor; inflammation; muscle mass; myonuclear accretion; negative energy balance; translational capacity.

Fig. 1.

Experimental design. The current analysis was part of a larger study assessing the effects of exogenous testosterone administration on changes in body composition after a 28-day exercise- and diet-induced energy deficit (ED) designed to be 45% of total energy needs (43). Biopsies were collected at the end of weight maintenance (WM) and ED before (Resting) and 1 (Post) and 6 (Recovery) h after exercise (1 h cycle ergometry), with a mixed meal (40 g protein) consumed following the first postexercise biopsy. Steady-state aerobic exercise bouts were matched between WM and ED for each participant based on power output (124 ± 22 W) and total work performed (448 ± 77 kJ). DXA, dual-energy X-ray absorptiometry.

Fig. 2.

Participant stratification according to leg lean mass and total testosterone. Changes in leg lean mass (kg) and total testosterone (ng/dL) were positively associated for all 50 participants. A subset of individuals exhibiting marked increases (TEST, n = 10) or decreases (PLA, n = 10) in both leg lean mass and total testosterone were included in all analyses.

Fig. 3.

Resting androgen receptor (AR) protein content (A), muscle total RNA content (B), fibroblast growth factor-inducible 14 (Fn14, C), IL-6 receptor (IL-6R, D), and muscle ring-finger protein-1 (MuRF1, E) gene expression during energy deficit (ED) relative to weight maintenance (WM) and associations between AR, total RNA, Fn14, IL-6R, and MuRF1 changes relative to WM (F–I) in subjects receiving 200 mg testosterone enanthate/wk (TEST) or placebo (PLA). Total AR was normalized to heat shock protein 90 (HSP90), and muscle total RNA concentrations (µg RNA/mg muscle) were calculated based on muscle sample total RNA yield relative to muscle weight. Gene data were normalized to the geometric mean of GUSB and TUBB, and fold changes were calculated using the ΔΔC_T method (45). Differences between TEST and PLA were examined at each time point using unpaired t tests, and associations were examined using Pearson’s correlation. Values are means ± SD [TEST, n = 9 and PLA, n = 10 (n = 9 for Fn14)]. *TEST different from PLA, P < 0.05.

Fig. 4.

Within [Resting (black bars), Post (gray bars), and Recovery (white bars)]- and between [weight maintenance (WM) and energy deficit (ED)]-study phase responses to exercise and a high-protein mixed meal for fibroblast growth factor-inducible 14 (Fn14, A), TNF-like weak inducer of apoptosis (TWEAK, B), TNF-α receptor (TNFα-R, C), TNF-α (D), IL-6 receptor (IL-6R, E), IL-6 (F), myogenic differentiation-1 (MyoD, G), myogenin (H), paired box 7 (Pax7, I), myogenic factor 5 (Myf5, J), myogenic factor 6 (Myf6, K), muscle ring-finger protein-1 (MuRF1, L), and muscle atrophy F-box (MAFbx, M) in subjects receiving 200 mg testosterone enanthate/wk (TEST) or placebo (PLA). Data were normalized to the geometric mean of GUSB and TUBB, and fold changes were calculated using the ΔΔC_T method (45). Changes in gene expression were evaluated using mixed-model repeated-measure ANOVA [n = 10 except Post for PLA during WM and ED (n = 9) and Resting for TEST during ED (n = 9); Fn14 and IL-6 Post for PLA during WM (n = 8), Recovery for PLA during WM (n = 9) and Resting for PLA during ED (n = 9). *Different from Resting, time main effect, P < 0.05. ⁺Different from Post, time main effect, P < 0.05. **Different from Resting, time-by-treatment interaction, P < 0.05. ⁺⁺Different from Post, time-by-treatment interaction, P < 0.05. #Different from WM, phase main effect, P < 0.05. ††Different from PLA, phase-by-treatment interaction, P < 0.05. ##Different from PLA, time-by-treatment interaction, P < 0.05. ‡Different from Resting, time-by-phase interaction, P < 0.05. ◊Different from Post, time-by-phase interaction, P < 0.05.

Fig. 5.

Within [Resting (black bars), Post (gray bars), and Recovery (white bars)]- and between [weight maintenance (WM) and energy deficit (ED)]-study phase responses to exercise and a high-protein mixed meal for phosphorylated (p) mechanistic target of rapamycin (mTOR)^Ser2448 (A), p-p70 ribosomal protein S6 kinase (p70S6K)^{Ser424/Thr421} (B), and p-ribosomal protein S6 (rpS6)^Ser240/244 (C) in subjects receiving 200 mg testosterone enanthate/wk (TEST) or placebo (PLA). For A–C, a representative band for the target phosphorylation site is on top and a representative band for total protein is on bottom. Protein phosphorylation status was expressed relative to totals of each protein, and changes were examined using mixed-model repeated-measure ANOVA. [n = 10 except WM Recovery (n = 9) and ED Resting (n = 9) for TEST and WM Resting (n = 9), WM Post (n = 9), and ED Post (n = 8) for PLA]. *Different from Resting, time main effect, P < 0.05.