Fig 2: Characterization of Ucn2 overexpression in vivo in mice skeletal muscle phenotype. The expression of Ucn2 mRNA (A) and Ucn2 protein (B) of normal mice transfected in vivo with the plasmid expressing pcDNA 3.1+ empty vector + GFP or mUcn2 + GFP for 7 and 14 days. The effect of Ucn2 overexpression for 7 and 14 days in the protein content of p-Ser133 CREB and PGC1-α (C, D), in CREB target genes (E) and in the protein content of Epac (F, G). The temporal effect of Ucn2 overexpression in vivo for 7 and 14 days in TA muscle mass of normal mice (H, I). P: phospho; AF: Alexa Fluor; GFP: Green Fluorescent Protein. Microscopy images with 40x of magnification. Negative control was obtained by omission of primary antibody. The nuclei were stained with Dapi. Results are expressed as mean ± SEM. (∗P ≤ 0.05 vs control) (n = 4–5).

Fig 3: CEA enhances activated PGC-1α to regulate fatty acid metabolism. a-c Volcano plot (a), GO enrichment analysis (b) of the differentially expressed genes in Veh and CEA treated A549 cells, Heatmap of differentially expressed genes that involved in fatty acid metabolism (c), n = 3. d A549 cells were treated with CEA (50 ng/mL) for 24 h, PPARA, PPARG, PPARGC1A and UCP2 expression were assessed; n = 3. e A549 cells were treated with CEA for 24 h, PGC-1α expression was determined by western blot analysis. Data are expressed as means ± SD. *P < 0.05, **P < 0.01, ***P < 0.0001, by two-sided unpaired student’s t test

Fig 4: Specific amino-acid mixtures prevent mitochondrial dysfunction in HL-1 cardiomyocytes acutely exposed to DOX. (A) Mitochondrial biogenesis marker expression: peroxisome proliferator-activated receptor-γ coactivator 1α (PGC1-α), nuclear respiratory factor-1 (NRF1), transcription factor A (Tfam), cytochrome c (cyt c), and cytochrome c oxidase subunit IV (COX IV) mRNA levels were analysed by quantitative RT-PCR. Relative expression values for the untreated (CTRL) cells were taken as 1.0 (n = 5 experiments). (B) COX IV and cyt c protein levels were detected by immunoblot analysis. The relative values were determined by densitometric analysis relative to GAPDH levels; values for untreated (CTRL) cells were taken as 1.0 (n = 5 experiments). (C) Citrate synthase activity. The values were normalized to protein content (n = 3 experiments). * p < 0.05, ** p < 0.01, and ***p < 0.001 vs. untreated cells; † p < 0.01 vs. DOX-treated cells; § p < 0.01 vs. BCAAem-treated cells. All data are presented as the mean ± SD.

Fig 5: Effect of PSP on the expression of related proteins (Bcl-2, Bax and caspase-3, SIRT1 and PGC-1α) in the ischemic hippocampus of rats in each group. (A) The protein analyzed by western blotting. (B) The ratio of Bax to Bcl protein level. The quantitative graph of (C) caspase-3, (D) SIRT1 and (E) PGC-1 α protein levels. PSP1 is the low dose group (150 mg/kg), PSP2 is the middle dose group (200 mg/kg) and PSP3 is the high dose group (250 mg/kg). The data are expressed as the mean ± standard deviation. **P<0.01 vs. respective sham; #P<0.05, ##P<0.01 vs. respective IR. PGC-1α, peroxisome proliferator-activated receptor γ coactivator-1α; Bcl-2, B-cell lymphoma 2; Bax, Bcl-2-like protein 4; SIRT1, silent information regulator protein 1; PGC-1α, peroxisome proliferator-activated receptor γ coactivator-1α; PSP, polysaccharide peptide; IR, ischemia reperfusion.