Sulfur Amino Acid Restriction Mitigates High-Fat Diet-Induced Molecular Alterations in Cardiac Remodeling Primarily via FGF21-Independent Mechanisms

Figures

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  Figure 1. SAAR reduced body weight and visceral adiposity independently of FGF21. Body weight (n = 10–14 animals per group) was measured prior to sacrifice. The weights of eWAT and rpWAT were measured post-sacrifice (n = 7–11 animals per group). Statistical analysis was performed using one-way ANOVA followed by Tukey–Kramer post-hoc multiple comparison tests. Significance levels to the NFD group are denoted by the * symbol: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Significance levels to the HFD group are denoted by the # symbol: ### p < 0.001, and #### p < 0.0001. Abbreviations: eWAT, epididymal white adipose tissue; rpWAT, retroperitoneal white adipose tissue.

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  Figure 2. SAAR modulated the expression levels of the cardioprotective hormones FGF21 and Adiponectin. Serum FGF21 levels were quantified using an ELISA kit (n = 4–6 animals per group). mRNA expression levels were quantified via RT-PCR (n = 7–11 animals per group). Statistical analysis was performed using one-way ANOVA followed by Tukey–Kramer post-hoc multiple comparison tests. Significance levels to the NFD group are denoted by the * symbol: * p < 0.05, ** p < 0.01, **** p < 0.0001. Significance levels to the HFD group are denoted by the # symbol: # p < 0.05 and ## p < 0.01.

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  Figure 3. Serum FGF21 levels are associated with Liver Fgf21 mRNA expression but not with Cardiac Fgf21 mRNA expression. Serum FGF21 levels were quantified using an ELISA kit. mRNA expression levels were quantified via RT-PCR. Relationship between serum FGF21 concentrations and liver Fgf21 (n = 20) and cardiac Fgf21 (n = 19) mRNA concentrations was determined with linear regression analysis. The errors of the linear regression lines are denoted in yellow.

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  Figure 4. SAAR enhanced the expression of cardiac lipid utilization genes and Slc2a1 in WT mice but not in Fgf21−/− mice. mRNA expression levels were quantified via RT-PCR (n = 6–11 animals per group). Statistical analysis was performed using one-way ANOVA followed by Tukey–Kramer post-hoc multiple comparison tests. Significance levels to the NFD group are denoted by the * symbol: * p < 0.05. Significance levels to the HFD group are denoted by the # symbol: # p < 0.05, ## p < 0.01 and ### p < 0.001.

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  Figure 5. SAAR modulated the expression of genes related to mitochondrial biogenesis across both genotypes in the heart. mRNA expression levels were quantified via RT-PCR (n = 7–11 animals per group). Protein quantification was conducted using Western blot (n = 6–11 animals per group). Statistical analysis was performed using one-way ANOVA followed by Tukey–Kramer post-hoc multiple comparison tests. Significance levels to the NFD group are denoted by the * symbol: * p < 0.05 and ** p < 0.01. Significance levels to the HFD group are denoted by the # symbol: # p < 0.05, ## p < 0.01 and ### p < 0.001.

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  Figure 6. SAAR abrogated the HFD-induced expression of genes associated with abnormal cellular stress responses across both genotypes in the heart. mRNA expression levels were quantified via RT-PCR (n = 6–11 animals per group). Statistical analysis was performed using one-way ANOVA followed by Tukey–Kramer post-hoc multiple comparison tests. Significance levels to the NFD group are denoted by the * symbol: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Significance levels to the HFD group are denoted by the # symbol: # p < 0.05, ## p < 0.01, ### p < 0.001, and #### p < 0.0001.

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  Figure 7. SAAR modulated the expression of genes associated with cardiac hypertrophy and fibrosis pathways in both genotypes. mRNA expression levels were quantified via RT-PCR (n = 7–11 animals per group). Statistical analysis was performed using one-way ANOVA followed by Tukey–Kramer post-hoc multiple comparison tests. Significance levels to the NFD group are denoted by the * symbol: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Significance levels to the HFD group are denoted by the # symbol: # p < 0.05, ## p < 0.01 and ### p < 0.001.

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  Figure 8. SAAR-induced molecular alterations in the heart are more indicative of physiological than pathological cardiac remodeling. Created with BioRender.com.