Peroxisomal dysfunction in cardiac adipose tissue is involved in obesity-associated cardiac hypertrophy

Fig. 1. Cardiac adipose tissue accumulation and browning inability induce cardiac hypertrophy in HFD-fed rats. (A) Common pathway categories identified by GSEA with the gene set database from the GSE159612 dataset are presented. (B) The body weight of SD rats was monitored weekly, and rats on a HFD were compared with those on a standard diet. (C) Relative tissue weights normalized to body weight were measured. (D) The morphology of the EAT and PVAT was observed using H&E staining. (E) UCP1 (green), TOM20 (red), and nuclei (blue) were detected by IF staining in the EAT and PVAT (original magnification × 400, scale bar = 75 μm). (F) Representative cross-sectional view of the mid-ventricular region of the whole rat heart by H&E staining (scale bar = 2 mm). Colored overlays indicate segmented anatomical structures: total heart (black), RV (green), LV (red), interventricular septum (orange), RV wall thickness (yellow), LV wall thickness (dark blue), and septal wall thickness (light blue). (G) Overview of serial short-axis sections (5 μm thickness) collected at 30 μm intervals through the mid-ventricular region (scale bar = 2 mm). (H) Cavity volume of the LV and RV was measured based on serial…

Fig. 2. Cardiac hypertrophy is induced by lipid metabolism plasticity in the EAT and PVAT of HFD-fed rats. (A) A schematic diagram of the H9C2 cardiomyocytes treated with lipid metabolite extracted from the EAT and PVAT. (B, C) H9C2 cells treated with lipid metabolite were stained for collagen using Masson’s Trichrome staining (magnification × 200, scale bar = 150 μm), while α-SMA (green) and nuclei (blue) were detected by IF staining in cardiomyofibers (magnification × 200, scale bar = 150 μm). (D, E) The gene expressions of Anp, Col3a1, Tnfa, Il6, Myh7, and Abcd1 were measured by RT-qPCR in the EAT and PVAT lipid metabolite-treated H9C2 cells. Results are expressed relative to Gapdh. (F) PCA is shown. (G) Heatmap visualization of fatty acid profiles in the EAT and PVAT from ND and HFD rats based on MSEA. (H, I) Volcano plots illustrate differential fatty acid composition between the HFD and ND groups in the EAT and PVAT. The data are presented as the mean ± SEM from three or more independent experiments. Statistical differences were evaluated using an unpaired t-test and a subsequent post hoc one-tailed Mann-Whitney U test. *p < 0.05 vs. ND.

Fig. 3. Peroxisomal lipid metabolism is altered in the EAT and PVAT of the HFD-fed rats. (A) Dot plot of significantly enriched pathways identified using the Small Molecule Pathway Database (SMPDB). (B) Pathway impact analysis based on the Kyoto Encyclopedia of Genes and Genomes (KEGG). (C) Expressions of genes associated with LCFA synthesis/degradation (Fasn, Acc, Elovl6, Scd1, and Hsd17b4) and SCFA synthesis/degradation (Acads, Acss2, and Ffar2) were accessed by RT-qPCR. The results are expressed relative to Gapdh. (D) The PPARα, PMP70, PEX13, PEX14, PEX16, and ACOX1 protein expressions in the PVAT and EAT were measured with a western blot assay. Results are expressed relative to β-actin. (E) PMP70 (green) and nuclei (blue) were detected by IF staining in the PVAT and EAT (original magnification × 400, scale bar = 30 μm). The data are presented as the mean ± SEM from three or more independent experiments. Statistical differences were evaluated using an unpaired t-test and a subsequent post hoc one-tailed Mann-Whitney U test. *p < 0.05 vs. ND.

Fig. 4. Peroxisomal dysfunction in cardiac adipocytes induces cardiac muscle dysfunction in H9C2 cardiomyocytes. (A) ROS and (B) catalase activity were measured in the SVF cells treated with 10, 12-TA, Mdivi-1, and WY-14643. (C) The gene expressions of Pex5, Pex11a, Pex13, Pex14, and Pex16 were determined by RT-qPCR. The results are expressed relative to Gapdh. (D) Lipid accumulation was shown by Oil Red O staining in SVF cells treated with 10, 12-TA, Mdivi-1, and WY-14643. Lipid droplet distributions were evaluated and normalized to DM (−). (E) A schematic diagram of the H9C2 cardiomyocytes treated with white ACM is presented. PMP70 (green) and nuclei (blue) were detected by IF staining in differentiated rat SVF cells (original magnification × 200, scale bar = 150 μm). (F) α-SMA (green) and nuclei (blue) were detected by IF staining in H9C2 cardiomyocytes (original magnification × 200, scale bar = 150 μm). (G) The H9C2 cardiomyocytes were stained with Masson’s Trichrome staining (original magnification × 200, scale bar = 150 μm). The cell surface area was measured using image J. (H) The gene expressions of Anp, Col3a1, Tnfa, and Myh7 were determined by RT-qPCR. The results are expressed…