Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Nov 23;144(21):1714-1731.
doi: 10.1161/CIRCULATIONAHA.121.053575. Epub 2021 Oct 21.

Altered Cardiac Energetics and Mitochondrial Dysfunction in Hypertrophic Cardiomyopathy

Sara Ranjbarvaziri  1   2 , Kristina B Kooiker  3 , Mathew Ellenberger  4 , Giovanni Fajardo  1   2 , Mingming Zhao  1   2 , Alison Schroer Vander Roest  1   2 , Rahel A Woldeyes  5 , Tiffany T Koyano  6 , Robyn Fong  6 , Ning Ma  2   7 , Lei Tian  2   7 , Gavin M Traber  4 , Frandics Chan  8 , John Perrino  9 , Sushma Reddy  1   2 , Wah Chiu  5   10 , Joseph C Wu  2   7 , Joseph Y Woo  6 , Kathleen M Ruppel  1   11 , James A Spudich, Michael P Snyder  4 , Kévin Contrepois  4 , Daniel Bernstein  1   2
Affiliations

Altered Cardiac Energetics and Mitochondrial Dysfunction in Hypertrophic Cardiomyopathy

Sara Ranjbarvaziri et al. Circulation. .

Abstract

Background: Hypertrophic cardiomyopathy (HCM) is a complex disease partly explained by the effects of individual gene variants on sarcomeric protein biomechanics. At the cellular level, HCM mutations most commonly enhance force production, leading to higher energy demands. Despite significant advances in elucidating sarcomeric structure-function relationships, there is still much to be learned about the mechanisms that link altered cardiac energetics to HCM phenotypes. In this work, we test the hypothesis that changes in cardiac energetics represent a common pathophysiologic pathway in HCM.

Methods: We performed a comprehensive multiomics profile of the molecular (transcripts, metabolites, and complex lipids), ultrastructural, and functional components of HCM energetics using myocardial samples from 27 HCM patients and 13 normal controls (donor hearts).

Results: Integrated omics analysis revealed alterations in a wide array of biochemical pathways with major dysregulation in fatty acid metabolism, reduction of acylcarnitines, and accumulation of free fatty acids. HCM hearts showed evidence of global energetic decompensation manifested by a decrease in high energy phosphate metabolites (ATP, ADP, and phosphocreatine) and a reduction in mitochondrial genes involved in creatine kinase and ATP synthesis. Accompanying these metabolic derangements, electron microscopy showed an increased fraction of severely damaged mitochondria with reduced cristae density, coinciding with reduced citrate synthase activity and mitochondrial oxidative respiration. These mitochondrial abnormalities were associated with elevated reactive oxygen species and reduced antioxidant defenses. However, despite significant mitochondrial injury, HCM hearts failed to upregulate mitophagic clearance.

Conclusions: Overall, our findings suggest that perturbed metabolic signaling and mitochondrial dysfunction are common pathogenic mechanisms in patients with HCM. These results highlight potential new drug targets for attenuation of the clinical disease through improving metabolic function and reducing mitochondrial injury.

Keywords: cardiomyopathy, hypertrophic; metabolism; mitochondria; mitophagy; reactive oxygen species.

PubMed Disclaimer

Conflict of interest statement

DISCLOSURES

The authors declare no conflict of interest.

Figures

Figure 1.
Figure 1.. Study design and molecular profiles in HCM.
(A) Summary of the experimental design workflow. (B) Metabolomic, lipidomic, and transcriptomic profiling was performed on left ventricle tissue from HCM patients and donor controls. Principal component analysis (PCA) plots of all 6,189 metabolite features(left), 728 lipids (middle), and 48,167 transcripts (right) clearly separate the profiles of HCM from controls. (C) Top metabolic pathways enriched in HCM identified by IMPaLA, integrating metabolites, lipids, and transcripts (FDR<0.05). Metabolic pathways were categorized as follows: proteins, blue; lipids, orange; carbohydrates, black; nucleotides, gray and energy metabolism (purple). (D) Integrated network analysis combining all omic datasets using Shiny Genes and Metabolites (GAM) platform. Resulting networks were annotated and plotted in Cytoscape. Each node (circle) represents a metabolite or lipid and each edge connecting nodes represents an enzyme-encoding transcript based on biochemical relationships. The arrowhead determines the principal direction of the biochemical reaction. The size of each node and of connecting line reflects their p value; the color represents the relative change (blue, decrease; red, increase; black, not significant; and gray, not detected). n=6 control and n=13 HCM samples for metabolites/lipids; n=7 control and n=13 HCM for transcripts. FFA, free fatty acids; M/S-chain, medium/short-chain; L-chain, long chain; AC, acylcarnitine; G3P, glyceraldehyde 3-phosphate; PPP, pentose phosphate pathway; Ru5P, ribulose-5-phosphate; R5P, ribose 5-phosphate; CER, ceramide; CE, cholesterol ester; PC, phosphatidylcholine; PE, phosphatidylethanolamine; LPC, lysophosphatidylcholine; LPE, lysophosphatidylethanolamine; DAG, diacylglycerol; and TAG, triacylglycerol.
Figure 2.
Figure 2.. Dysregulated metabolism in HCM.
(A) Total concentration (log2 scale) of free fatty acids (FFA) in HCM as measured by the targeted lipidomics platform. (B) Heatmap shows overall changes in genes regulating myocardial energetics. Shown are expression of genes encoding major fatty acid transporters, β-oxidation enzymes, a glucose transporter, and enzymes involved in regulation of glycolysis, TCA cycle, and mitochondrial energy metabolism. (C-F) Levels of acylcarnitines (total concentration, log2 scale), carnitine, sphingolipids (total concentration), and TCA cycle metabolites measured by untargeted metabolomics. (G) Citrate synthase (CS) activity in HCM vs control (spectrophotometry analysis). (H) High energy phosphate metabolites measured by untargeted metabolomics. Error bars represent mean ± SEM. n=6 control and n=13 HCM in A, C-F, and H; n=7 control and n=13 HCM in B; n=5 control and n=5 HCM in G. Between-group comparisons performed using Whitney U test in A, C, E, and G. Two-sided Welch’s t-test or Wald test with Benjamini-Hochberg’s FDR correction method used in B, D, F and H. *p<0.05, **p<0.01, ***p<0.005, or ****p<0.001.
Figure 3.
Figure 3.. Mitochondrial damage in HCM.
(A) Representative electron micrographs of myocardial tissue from donor and HCM. Each image represents one individual subject (scale bar=2µm). (B-D) Quantitative measurements of interfibrillar mitochondrial cristae density using ImageJ (~20 randomly selected images from each sample). (E) A slice through reconstructed 3D volume and corresponding segmented 3D volumes showing heterogeneous mitochondrial morphology within a single HCM sample (scale bar=100nm). (e-e”) Mitochondrial membranes are color coded in pink and cristae are shown in blue. (F) Chord plot representation of 51 differentially expressed genes (HCM vs control, FDR<0.05) from 7 enriched pathways generated by GOplot. The color map represents fold change of genes (log2 scale) and p value of go terms (−log10 scale). Error bars represent mean ± SEM. n=3 control and n=14 HCM in B-D; n=7 control and n=13 HCM in F. Between-group comparisons performed using Whitney U test. *p<0.05 or n.s., not significant (p>0.05).
Figure 4.
Figure 4.. Mitochondrial respiratory function and capacity in HCM.
(A) Average oxygen consumption rate (OCR) showing respiratory activity measured by Oroboros oximeter. Arrows indicate the sequential additions of mitochondrial substrates to assess respiratory states in freshly isolated myocardial tissue. (B) Analysis of ATP production capacity through oxidative phosphorylation (OXPHOS). (C) Mitochondrial complex activities in HCM vs control (spectrophotometry analysis). (D) The transcript level of UCP2 (log2 scale) measured by RNA-Seq. (E and F) Representative western blot and quantification of phosphorylated- and total AMPK. GAPDH was used as a loading control. (G) ELISA measurements of pAMPK to total AMPK. Error bars represent mean ± SEM. n=3 control and n=9 HCM in A and B; n=5 control and n=5 HCM in C; n=7 control and n=13 HCM in D; n=3 control and n=5 HCM in E and F; n=6 control and n=11 HCM in G. Between-group comparisons performed using Whitney U test in B, C, F, and G. Wald test with Benjamini-Hochberg’s FDR correction method used in D. *p<0.05, ***p<0.005, or n.s, not significant (p>0.05). Mal, malate; Glut, glutamate; Oligo, oligomycin A; FCCP, carbonyl cyanide-4-(trifluoromethoxy) phenylhydrazone; Anti-A, antimycin A; CI-CV, mitochondrial complex I-V; UCP, uncoupling protein.
Figure 5.
Figure 5.. Increased oxidative damage in HCM.
(A) 4-hydroxynonenal (4HNE) immunoreactivity indicative of oxidative stress measured by western blot. (B) Major cardiolipin species were measured by mass spec. (C) Quantification of mitochondrial DNA (mtDNA) to nuclear DNA (nDNA) using qPCR. (D) Cystine level measured by mass spec (log2 scale). (E) The ratio of oxidized glutathione (GSSG) to glutathione (GSH) (calculated from mass spec data). (F) The enzymatic activity of glutathione peroxidase (spectrophotometry analysis). (G) Violin plot showing mitophagy associated genes. (H and I) Representative Western blots and quantitative measurements of LC3A and B. GAPDH was used as a loading control. Violin plot was reserved to show RNA-Seq data. Error bars represent mean ± SEM. n=3 control and n=8 HCM in A; n=6 control and n=13 HCM in B, D, and E; n=5 control and n=10 HCM in C; n=5 control and n=5 HCM in F; n=7 control and n=13 HCM in G. n=6 control and n=10 HCM in H and I. Between-group comparisons performed using Whitney U test in A, C, E, F, and I. Two-sided Welch’s t-test and Benjamini-Hochberg’s FDR correction method used in B and D. *p<0.05, **p<0.01, ***p<0.005, ****p<0.001 or n.s, not significant (p>0.05). L4, tetralinoleoyl (18:2)4; L3O, trilinoleoyl-oleoyl (18:2)3(18:1)1; L2O2, dilinoleoyl-dioleoyl (18:2)2(18:1)2; LO3, linoleoyl-trioleoyl (18:2)1(18:1)3; O.2, superoxide radical; H2O2, hydrogen peroxide; GPx, glutathione peroxidase; LC3, Microtubule-associated proteins 1A/1B-light chain 3.
Figure 6.
Figure 6.. Mitochondrial alterations in HCM.
HCM hearts exhibit impaired fatty acid oxidation and reduced glucose metabolism. Along with these metabolic changes, increased energy demands in hypercontractile HCM hearts enhance ROS production, which together with insufficient antioxidant contents causes damage to diverse mitochondrial sites including mtDNA, cardiolipins, cristae and mitochondrial respiratory complexes. These mitochondrial abnormalities lead to reduced mitochondrial respiration and high energy phosphate molecules ultimately causing myocardial energy deprivation. Italics represents changes at the transcript level while metabolites and enzymatic activities are shown in regular font. Color indicates relative changes in the respective biomolecules: blue, decrease; red, increase. FABP, fatty acid binding protein; CD36, cluster of differentiation 36; CPT1, carnitine palmitoyl transferase type 1; PEP, phosphoenolpyruvate; CoA, Coenzyme A; CI-V, Complex I-V; mtDNA, mitochondrial DNA; TCA, tricarboxylic acid; ACs, acylcarnitines; ROS, reactive oxygen species; GSH, reduced glutathione; GSSG, oxidized GSH; SCLC2A1, SLC25A4 and SLC22A5 genes encode glucose transporter1 (GLUT1), the mitochondrial ADP/ATP translocator (ANT1), and carnitine transporter respectively.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Maron BJ and Maron MS. Hypertrophic cardiomyopathy. Lancet. 2013;381:242–55. - PubMed
    1. Semsarian C, Ingles J, Maron MS and Maron BJ. New perspectives on the prevalence of hypertrophic cardiomyopathy. J Am Coll Cardiol. 2015;65:1249–1254. - PubMed
    1. Spudich JA. Three perspectives on the molecular basis of hypercontractility caused by hypertrophic cardiomyopathy mutations. Pflugers Arch. 2019;471:701–717. - PMC - PubMed
    1. Ritterhoff J and Tian R. Metabolism in cardiomyopathy: every substrate matters. Cardiovascular Research. 2017;113:411–421. - PMC - PubMed
    1. Marian AJ and Braunwald E. Hypertrophic Cardiomyopathy: Genetics, Pathogenesis, Clinical Manifestations, Diagnosis, and Therapy. Circ Res. 2017;121:749–770. - PMC - PubMed

Publication types

MeSH terms

Substances