Late-life restoration of mitochondrial function reverses cardiac dysfunction in old mice
- Department of Pathology, University of Washington, United States
- Aging and Metabolism Program, Oklahoma Medical Research Foundation, United States
- Department of Genome Science, University of Washington, United States
- Buck Institute for Research on Aging, United States
- Department of Radiology, University of Washington, United States
- Biological Sciences Division, Pacific Northwest National Laboratory, United States
- Department of Environmental and Occupational Health Sciences, University of Washington, United States
- Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, United States
- Department of Cellular and Molecular Medicine, University of Arizona, United States
- Social Profit Network, United States
Figures
Figure 1 with 3 supplements
SS-31 treatment reverses cardiac aging phenotypes and improves exercise performance in old mice.
Doppler echocardiography showed that 8-week SS-31 treatment (a) improved diastolic function (increased Ea/Aa) and (b) enhanced myocardial performance (reduced myocardial performance index, MPI) of …
Figure 1—figure supplement 1
Individual trajectories of SS-31 responses in cardiac function of male mice.
8-week SS-31 treatment improved diastolic function and myocardial performance over time. Linear regression analyses treating each mouse in a group as a separate point revealed increased Ea/Aa (a) …
Figure 1—figure supplement 2
Individual trajectories of SS-31 responses in exercise performance of male mice.
Treadmill running time trended to decline in majority of old saline control mice (p=0.11) but not in old SS-31 treated mice (p=0.48); n = 10 for old control and n = 9 for old SS-31, analyzed by …
Figure 1—figure supplement 3
Individual trajectories of SS-31 responses in cardiac function of female mice.
8-week SS-31 treatment improved diastolic function in old female mice (p<0.001 between 0 week and 8 week by paired T-test); this improvement persisted 2 weeks after cessation of treatment (p=0.006 …
Figure 2 with 1 supplement
SS-31 treatment reduces ROS production and improves respiration in cardiomyocytes.
(a) SS-31 treated cardiomyocytes showed reduced mitochondrial superoxide, indicated by reduced MitoSox signal (normalized to mitochondrial content by the ratio to MitoTracker Green), compared to old …
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Figure 2—source data 1
All image files for MitoSOX analysis in Figure 2a.
- https://cdn.elifesciences.org/articles/55513/elife-55513-fig2-data1-v2.zip
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Figure 2—source data 2
All image files for MitoPY1 analysis in Figure 2b.
- https://cdn.elifesciences.org/articles/55513/elife-55513-fig2-data2-v2.zip
Figure 2—figure supplement 1
SS-31 treatment increases mitochondrial membrane potential in aged cardiomyocytes.
*p<0.05 vs old saline. n = 19 (Control) and 18 (SS-31) cells were analyzed by unpaired T-test.
Figure 3
SS-31 treatment does not alter expression of subunits of oxidative phosphorylation complexes.
Immunoblotting using anti-OXPHOS antibody detected no differences in expression levels of OXPHOS subunits (NDUFB8, SDHB, UQCRC2, MTCO1, and ATP5A) in hearts of old male mice treated with SS-31 for 8 …
Figure 4
SS-31 treatment reduces protein oxidation and senescence in old hearts.
(a) A histogram of the distribution of changes in glutathionylation levels in peptides from old control and old SS-31 treated hearts; n = 3 female mice per group, analyzed as described in the …
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Figure 4—source data 1
All image files for p16 analysis in Figure 4c.
- https://cdn.elifesciences.org/articles/55513/elife-55513-fig4-data1-v2.zip
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Figure 4—source data 2
All image files for p19 analysis in Figure 4d.
- https://cdn.elifesciences.org/articles/55513/elife-55513-fig4-data2-v2.zip
Figure 5 with 2 supplements
SS-31 treatment partially restores age-related proteomic remodeling.
A heatmap of z-scores the 88 proteins that were significantly altered by both aging (q < 0.05 for old control vs. young control) and SS-31 treatment (q < 0.05 for old SS-31 vs. old control); n = 9, …
Figure 5—figure supplement 1
SS-31 treatment induces modest changes in metabolome that partially attenuates the age-related changes.
A heat map of the relative levels of the 18 metabolites that were significantly (FDR < 0.05) different among treatment groups; n = 10 (for YCL, YSS, OCL) and n = 8 (for OSS) were male mice were …
Figure 5—figure supplement 2
A network of metabolite set enrichment for the 11 metabolites that were significantly (p<0.05, by Tukey’s HSD) altered by aging.
Figure 6
SS-31 rescues the age-related hypo-phosphorylation of MyBP-C.
(a) Old murine hearts displayed reduced levels of MyBP-C phosphorylation at Ser282, which is normalized by SS-31 treatment. (b–c) Aging and SS-31 treatment did not alter phosphorylation of cTnI at …
Figure 7
The cardiac benefit of SS-31 treatment is not additive to that of mCAT expression but the two interventions differentially regulate myofilament protein phosphorylation.
(a) Diastolic function (Ea/Aa) improved at both 8 and 12 weeks after AAV9-mCAT administration. n = 3 (saline) and n = 5 (AAV-mCAT) female mice were analyzed by repeated measure ANOVA with Tukey’s …
Figure 8
Schematic outline of results and interpretation.
While mCAT and SS-31 both inhibit electron transport chain produced ROS, they do so by different mechanisms. Both inhibit a ROS-mediated vicious cycle (ROS induced mtDNA and protein damage leads to …
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Strain, strain background (M. musculus; male and female) | C57BL/6J | National Institute of Aging Charles River colony | RRID:IMSR_JAX:000664 | |
| Genetic reagent (M. musculus) | mCAT | Rabinovitch Lab; PMID:15879174 | RRID:IMSR_JAX:016197; Tg(CAG-OTC/CAT)4033Prab | Now available at The Jackson Lab |
| Transfected construct (M. musculus) | AAV9-mCAT | Duan Lab, PMID:19690612 | AV.RSV.MCAT | Adeno-associated virus construct to transfect and express mCAT transgene |
| Antibody | Anti-OXPHOS (Rabbit polyclonal) | Abcam | ab110413 RRID:AB_2629281 | (1:500) |
| Antibody | Anti-Troponin I (Rabbit polyclonal) | Cell Signaling Technology | #4002 RRID:AB_2206278 | (1:1000) |
| Antibody | Anti-pSer23/24-Troponin I (Rabbit polyclonal) | Cell Signaling Technology | #4004 RRID:AB_2206275 | (1:1000) |
| Antibody | Anti-pSer150-Troponin I (Rabbit polyclonal) | ThermoFisher | PA5-35410 RRID:AB_2552720 | (1:1000) |
| Antibody | Anti- cMyBP-C (Mouse monoclonal) | Santa Cruz SC-137237 | SC-137237 RRID:AB_2148327 | (1:1000) |
| Antibody | Anti-pSer282-cMyBP-C (Rabbit polyclonal) | Enzo | ALX-215–057 R050 RRID:AB_2050502 | (1:2000) |
| Antibody | anti-p19 (Rabbit polyclonal) | LSBio | LS-C49180 RRID:AB_1192824 | (1:300) |
| Antibody | anti-p16 (Rabbit polyclonal) | Abcam | ab211542 | (1:300) |
| Antibody | Donkey anti-Rabbit Secondary Antibody, HRP | ThermoFisher | A16035 RRID:AB_2534709 | (1:10000) |
| Antibody | Goat anti-Mouse Secondary Antibody, HRP | ThermoFisher | A16072 RRID:AB_2534745 | (1:10000) |
| Peptide, recombinant protein | SS-31 peptide (Elamipretide) | Stealth BioTherapeutics | 3 µg/g body weight/day | |
| Commercial assay or kit | OxiSelect protein carbonyl ELISA kit | Cell Biolabs | STA-310 | |
| Commercial assay or kit | ImmPRESS-VR Anti-Rabbit IgG HRP Polymer Detection Kit | Vector Laboratories | MP-6401–15 | |
| Commercial assay or kit | Seahorse XF Cell Mito Stress Test Kit | Aligent/Seahorse Bioscience | 103015–100 | |
| Commercial assay or kit | MitoSOX Red | ThermoFisher | M36008 | |
| Commercial assay or kit | MitoPY1 | Fisher Scientific/Tocris Bioscience | 44-281-0 | |
| Commercial assay or kit | MitoTracker Green | ThermoFisher | M7514 | |
| Commercial assay or kit | MitoTracker Deep Red | ThermoFisher | M22426 | |
| Commercial assay or kit | JC-1 Dye | ThermoFisher | T3168 | |
| Commercial assay or kit | BCA protein assay | Thermo Scientific | 23225 | |
| Commercial assay or kit | Pierce Reversible Protein Stain Kit for PVDF Membranes | Thermo Scientific | 24585 | |
| Commercial assay or kit | SuperSignal West Pico PLUS Chemiluminescent Substrate | Thermo Scientific | 34580 | |
| Software, algorithm | Graphpad Prism | Graphpad | RRID:SCR_002798 | |
| Software, algorithm | AlphaView Software | ProteinSimple | ||
| Software, algorithm | Metaboanalyst 4.0 | www.metaboanalyst.ca; PMID:29762782 | RRID:SCR_015539 | |
| Software, algorithm | Topograph | MacCoss Lab software; PMID:22865922 | ||
| Software, algorithm | ComplexHeatmap | https://github.com/jokergoo/ComplexHeatmap PMID:27207943 | RRID:SCR_017270 |
Additional files
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Supplementary file 1
A dataset of relative abundances of all metabolites measured.
- https://cdn.elifesciences.org/articles/55513/elife-55513-supp1-v2.csv
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Supplementary file 2
A table of all metabolites that showed significant differences in one or more comparisons.
- https://cdn.elifesciences.org/articles/55513/elife-55513-supp2-v2.xlsx
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Supplementary file 3
A dataset of identities, fold changes and statistics of all proteins identified in the proteomic analysis.
- https://cdn.elifesciences.org/articles/55513/elife-55513-supp3-v2.csv
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Transparent reporting form
- https://cdn.elifesciences.org/articles/55513/elife-55513-transrepform-v2.pdf
