SIRT1 Activation by Resveratrol Alleviates Cardiac Dysfunction via Mitochondrial Regulation in Diabetic Cardiomyopathy Mice

Figures

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  Figure 1. Resveratrol alleviated cardiac dysfunction in DCM mouse heart. (a) Increased heart weight/tibia length in STZ-induced DCM mice (∗p < 0.05). (b) Increased ventricle/body weight in DCM mice as compared with Con mice (∗p < 0.05). (c) Elevated expression of myocardial hypertrophy markers of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) in DCM mouse heart (∗p < 0.05). (d) Impaired cardiac function in diabetic hearts (∗p < 0.05), while resveratrol treatment significantly reversed these changes in DCM mice (#p < 0.05). (e) DCM mouse hearts displayed structural changes and fibrogenesis, and resveratrol markedly alleviated these changes. (f) SIRT1 was markedly downregulated in DCM mouse myocardium (∗p < 0.05), while resveratrol enhanced SIRT1 expression in DCM + RES mice (#p < 0.05). ∗p < 0.05 versus Con; #p < 0.05 versus DCM.

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  Figure 2. Cardiac-specific SIRT1 knockout (SIRT1KO) mice displayed symptoms of DCM. (a) Mouse tail tissue PCR showed different genotypes during the process of crossbreeding. (b) There was almost no SIRT1 mRNA in SIRT1KO mouse cardiac tissue (∗p < 0.05 in Heter group, ∗∗p < 0.01 in SIRT1KO group). (c) SIRT1 protein was also barely expressed in SIRT1KO mice myocardium (∗∗p < 0.01 in SIRT1KO group). (d) Increased heart weight/tibia length ratio in SIRT1KO mouse myocardium and ventricle/heart weight in SIRT1KO mouse myocardium (∗p < 0.05). (e) The mRNA expressions of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) were significantly increased in SIRT1KO mice (∗p < 0.05). (f) SIRT1KO mouse hearts displayed structural changes and fibrogenesis in the myocardium. (g) Cardiac function was impaired in SIRT1KO mice as compared with WT and Heter mice (∗p < 0.05) (Figure 1(g)). ∗p < 0.05 versus WT. WT: wild type; Heter: heterozygous.

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  Figure 3. Myocardial metabolic and mitochondrial alterations in DCM and SIRT1KO mice. (a) TEM images revealed morphological mitochondrial impairment in both DCM and SIRT1KO mice, and SIRT1 activation by resveratrol could alleviate these mitochondrial changes in DCM + RES mice. (b) Mitochondrial DNA (mtDNA) amount was significantly decreased in the DCM, SIRT1KO, and SIRT1KO + DCM groups as compared with the Con group (∗p < 0.05). Resveratrol elevated mtDNA amount in the DCM + R group (#p < 0.05), while resveratrol's protective effects were diminished in the SIRT1KO + DCM + R group as compared with the DCM + RES group (&p < 0.05). (c) Complex IV activity was significantly reduced in the DCM, SIRT1KO, and SIRT1KO + DCM groups (∗p < 0.05) than in the Con group. Resveratrol markedly alleviated complex IV activity reduction in DCM mice (#p < 0.05). While in contrast, the beneficial effect of resveratrol was completely abolished in the SIRT1KO + DCM + R group (&p < 0.05, SIRT1KO + DCM + R versus DCM + RES). (d) Both DCM and SIRT1KO led to defective 18F-FDG uptake in myocardium, and resveratrol ameliorated this in DCM + R mice but not in SIRT1KO + DCM + R. (e) Insulin receptor substrate 2 (IRS2)…

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  Figure 4. High glucose and SIRT1 deletion impaired mitochondrial biogenesis and function. (a and b) Both HG and SIRT1 deletion by shRNA lentiviral vector increased apoptotic cell ratio in H9c2 cells (∗p < 0.05), but resveratrol did not suppress HG-induced apoptosis in SIRT1KD + HG + R cells as in HG + RES cells (#p < 0.05, &p < 0.05). (c) Control cells showed strong TMRM fluorescence, while HG and SIRT1KD resulted in decreased TMRM fluorescence due to depolarization of the mitochondria. Additionally, resveratrol relatively increased TMRM fluorescence in HG + RES cells but not in SIRT1KD + HG + R cells. (d) mtDNA amount was reduced in HG and SIRT1KD-treated cells (∗p < 0.05), and resveratrol markedly increased mtDNA amount in the HG + RES group but not in the SIRT1KD + HG + R group (#p < 0.05, &p < 0.05). (e) Mitochondrial enzyme activity was reduced in HG and SIRT1KD-grouped cells (∗p < 0.05), and resveratrol markedly elevated it in the HG + RES group but not in the SIRT1KD + HG + R group (#p < 0.05, &p < 0.05). (f) ATP generation demonstrated the similar tendency with mitochondrial enzyme activity. ∗p < 0.05 versus Con; #p < 0.05 versus DCM; &p < 0.05 versus DCM + RES.

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  Figure 5. SIRT1 regulated mitochondrial-related proteins expression through PGC-1α deacetylation. (a) Nonfunctional PGC-1α acetylation (about 105 kDa, indicated by the red box) was upregulated in the HG and SIRT1KD groups; meanwhile, SIRT1 activation in HG + RES cells promoted functional PGC-1α deacetylation. (b) HG and SIRT1KD decreased the mRNA expression of mitochondrion-related genes such as NRF-1, NRF-2, ERR-α, and TFAM (∗p < 0.05). SIRT1 activation by resveratrol upregulated the expression of NRF-1, NRF-2, ERR-α, and TFAM mRNA expression in HG + RES cells but not in SIRT1KD + HG + R cells (#p < 0.05, &p < 0.05). (c) Protein expression level changes of NRF-1, NRF-2, ERR-α, and TFAM showed the similar tendency with mRNA. ∗p < 0.05 versus Con; #p < 0.05 versus DCM; &p < 0.05 versus DCM + RES. NRF-1: nuclear respiratory factor 1; NRF-2: nuclear respiratory factor 2; ERR-α: estrogen-related receptor-α; TFAM: mitochondrial transcription factor A.