Sirt3 blocks the cardiac hypertrophic response by augmenting Foxo3a-dependent antioxidant defense mechanisms in mice
Nagalingam R. Sundaresan, … , Ayman Isbatan, Mahesh P. Gupta
Nagalingam R. Sundaresan, … , Ayman Isbatan, Mahesh P. Gupta
Published August 3, 2009
Citation Information: J Clin Invest. 2009;119(9):2758-2771. https://doi.org/10.1172/JCI39162.
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Research Article Aging

Sirt3 blocks the cardiac hypertrophic response by augmenting Foxo3a-dependent antioxidant defense mechanisms in mice

Abstract

Sirtuin 3 (SIRT3) is a member of the sirtuin family of proteins that promote longevity in many organisms. Increased expression of SIRT3 has been linked to an extended life span in humans. Here, we have shown that Sirt3 protects the mouse heart by blocking the cardiac hypertrophic response. Although Sirt3-deficient mice appeared to have normal activity, they showed signs of cardiac hypertrophy and interstitial fibrosis at 8 weeks of age. Application of hypertrophic stimuli to these mice produced a severe cardiac hypertrophic response, whereas Sirt3-expressing Tg mice were protected from similar stimuli. In primary cultures of cardiomyocytes, Sirt3 blocked cardiac hypertrophy by activating the forkhead box O3a–dependent (Foxo3a-dependent), antioxidant–encoding genes manganese superoxide dismutase (MnSOD) and catalase (Cat), thereby decreasing cellular levels of ROS. Reduced ROS levels suppressed Ras activation and downstream signaling through the MAPK/ERK and PI3K/Akt pathways. This resulted in repressed activity of transcription factors, specifically GATA4 and NFAT, and translation factors, specifically eukaryotic initiation factor 4E (elf4E) and S6 ribosomal protein (S6P), which are involved in the development of cardiac hypertrophy. These results demonstrate that SIRT3 is an endogenous negative regulator of cardiac hypertrophy, which protects hearts by suppressing cellular levels of ROS.

Authors

Nagalingam R. Sundaresan, Madhu Gupta, Gene Kim, Senthilkumar B. Rajamohan, Ayman Isbatan, Mahesh P. Gupta

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Figure 7

Sirt3 blocks Ras activation and mitochondrial ROS accumulation during hypertrophy.

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Sirt3 blocks Ras activation and mitochondrial ROS accumulation during hy...
(A) Rat cardiomyocytes were infected with the indicated adenoviruses and treated with PE (20 μM) for indicated time intervals. From the cell extract, Raf was immunoprecipitated with use of Raf-RBD beads, and the complex was analyzed for coprecipitation of active Ras by Western blotting (WB). Sirt3 overexpression suppressed the Ras-Raf binding both at 5 and 10 minutes after PE treatment, an indication of Ras deactivation. (B and C) Heart extracts of different groups of mice subjected to ISO-mediated hypertrophy were analyzed for coprecipitation of active Ras with Raf-RBD. Ras was highly activated in Sirt3-KO hearts, whereas it was suppressed in Sirt3-Tg hearts compared with controls. (D) Real-time measurement of mitochondrial ROS production in rat cardiomyocytes infected with Ad.Sirt3 or Ad.Smut virus. Cells were induced with 20 μM of PE in a thermoregulatory chamber of microscope, and the production of ROS (red fluorescence) from cells was determined at regular intervals with use of MitoSox dye by time-lapse confocal microscopy. (E) The increase of ROS was quantified by measurement of the intensity of fluorescence at different time intervals (mean ± SEM, n = 100 cells). *P < 0.01 compared with Ad.Sirt3-infected cells. (F) Cardiomyocytes cultured from neonatal hearts of Sirt3-KO and WT mice were stimulated with PE (20 μM) and ROS production measured as in D. (G) Quantification of ROS levels in 2 groups of cardiomyocytes shown in F (mean ± SEM, n = 50 cells). *P < 0.01 compared with WT cells. Original magnification, ×630 (D and F). GST, glutathione-S-transferase.