A combination of aerobic and resistance training induces white adipose tissue browning

Figures

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  Fig. 1. Improvement in exercise training performance after 8 weeks of CT exercise. Vpeak_K and maximum load lifted were determined during the 1st, 4th, and 8th weeks. (A) Combined training protocol and experimental design, (B) Aerobic performance was assessed by peak velocity during the incremental test (Vpeak_K); (C) Resistance performance was assessed by the maximum load lifted (g). Data are presented as mean ± SD and were analyzed using two-way repeated-measures ANOVA, followed by Šídák’s multiple-comparisons test. **p < 0.01 and ****p < 0.0001 versus the corresponding SED group at the same time point. Abbreviations: SED, sedentary; CT, combined training; AER, aerobic training; RES, resistance training. SED, n = 10 animals; CT, n = 10 animals

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  Fig. 2. Food intake and body mass before and after 8 weeks of CT. (A) Food intake was assessed in SED (n = 10) and CT (n = 10) animals. (B) Body mass was assessed in SED (n = 30) and CT (n = 30) animals. Data are presented as mean ± SD and were analyzed using two-way repeated-measures ANOVA, followed by Šídák’s multiple-comparisons test. *p < 0.05 and ****p < 0.0001 versus the respective pre-training value. Abbreviations: SED, sedentary; CT, combined training; Pre, pre-training; Post, post-training

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  Fig. 3. CT increased VEGF expression in WAT depots after 8 weeks. (A) VEGF was detected by immunohistochemistry; (B) VEGF positive-staining area per field (µm²) after 8 weeks of CT. Data are presented as mean ± SD. Comparisons between SED and CT were performed using an unpaired two-tailed Student’s t-test. *p < 0.05 versus the corresponding SED group. The reported sample size corresponds to the final number of animals with successfully processed and analyzed tissue sections. Black arrows indicate VEGF-positive structures. Scale bars, 50 μm. Abbreviations: SED, sedentary (n = 5 animals); CT, combined training (n = 5 animals); WAT, white adipose tissue; iWAT, inguinal white adipose tissue; rpWAT, retroperitoneal white adipose tissue

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  Fig. 4. CT enhances CD31 expression in WAT after 8 weeks. (A) Representative immunofluorescence images of CD31 staining in iWAT and rpWAT; (B) CD31 positive-staining area per field (µm²) in each adipose tissue depot. Data are presented as mean ± SD. Comparisons between SED and CT were performed using an unpaired two-tailed Student’s t-test. *p < 0.05 versus the corresponding SED group. The reported sample size corresponds to the final number of animals with successfully processed and analyzed tissue sections. White arrows indicate CD31-positive structures. Scale bars, 50 μm. Abbreviations: SED, sedentary (n = 5 animals); CT, combined training (n = 5 animals); WAT, white adipose tissue; iWAT, inguinal white adipose tissue; rpWAT, retroperitoneal white adipose tissue; DAPI, 4′,6-diamidino-2-phenylindole; CD31, cluster of differentiation 31

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  Fig. 5. CT increased UCP1 expression in iWAT and rpWAT after 8 weeks. (A) Representative immunohistochemistry images of UCP1 staining in iWAT and rpWAT; (B) UCP1 positive-staining area per field (µm²) in each adipose tissue depot; and (C) Multilocular adipocyte area normalized to the total WAT area analyzed. Data are presented as mean ± SD. Comparisons between SED and CT were performed using an unpaired two-tailed Student’s t-test. *p < 0.05 versus the corresponding SED group. The reported sample size corresponds to the final number of animals with successfully processed and analyzed tissue sections. Scale bars, 50 μm. Abbreviations: SED, sedentary (n = 5 animals); CT, combined training (n = 5 animals); WAT, white adipose tissue; iWAT, inguinal white adipose tissue; rpWAT, retroperitoneal white adipose tissue; UCP1, uncoupling protein 1

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  Fig. 6. CT increased the expression of selected genes associated with brown and beige adipocyte phenotypes. Gene expression was assessed by qPCR in iWAT and rpWAT. Each data point represents one pooled biological sample generated from two mice. (A) Ucp1: SED, n = 9; CT, n = 11. iWAT, q = 0.000165; rpWAT, q = 0.000005. (B) Pgc1α: SED, n = 9; CT, n = 10. iWAT, q = 0.0011; rpWAT, q < 0.000001. (C) Cidea: SED, n = 9; CT, n = 7. iWAT, q = 0.000549; rpWAT, q = 0.001232. (D) Prdm16: iWAT: SED, n = 7; CT, n = 7; q = 0.008779. rpWAT: SED, n = 8; CT, n = 8; q = 0.009927. (E) Eva1: SED, n = 7; CT, n = 7. iWAT, q = 0.020144; rpWAT, q = 0.000011. (F) Tmem26: iWAT: SED, n = 9; CT, n = 11; q = 0.0011. rpWAT: SED, n = 9; CT, n = 6; q = 0.00047. (G) Cd137: SED, n = 9; CT, n = 9. iWAT, q = 0.008779; rpWAT, q = 0.000106. (H) Tbx1: iWAT: SED, n = 8; CT, n = 7; q = 0.041811. rpWAT: SED, n = 7; CT, n = 8; q < 0.000001. Data are presented as mean ± SD. Between-group comparisons were performed using Welch’s unpaired t-tests for each gene. Multiple testing across the eight genes was controlled separately within each adipose tissue depot using the two-stage step-up FDR procedure of Benjamini, Krieger, and…

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  Fig. 7. Representative illustration of white adipose tissue (WAT) from sedentary and combined training (CT) mice after 8 weeks of intervention. CT promoted depot-specific changes in WAT mass and reduced adipocyte area, increased angiogenesis-associated markers (VEGF/CD31), presence of multilocular adipocytes expressing UCP1, and upregulation of brown-like and beige adipogenic genes