Hypertrophic Cardiomyopathy: A Vicious Cycle Triggered by Sarcomere Mutations and Secondary Disease Hits

Abstract

Significance: Hypertrophic cardiomyopathy (HCM) is a cardiac genetic disease characterized by left ventricular hypertrophy, diastolic dysfunction, and myocardial disarray. Disease onset occurs between 20 and 50 years of age, thus affecting patients in the prime of their life. HCM is caused by mutations in sarcomere proteins, the contractile building blocks of the heart. Despite increased knowledge of causal mutations, the exact path from genetic defect leading to cardiomyopathy is complex and involves additional disease hits. Recent Advances: Laboratory-based studies indicate that HCM development not only depends on the primary sarcomere impairment caused by the mutation but also on secondary disease-related alterations in the heart. Here we propose a vicious mutation-induced disease cycle, in which a mutation-induced energy depletion alters cellular metabolism with increased mitochondrial work, which triggers secondary disease modifiers that will worsen disease and ultimately lead to end-stage HCM. Critical Issues: Evidence shows excessive cellular reactive oxygen species (ROS) in HCM patients and HCM animal models. Oxidative stress markers are increased in the heart (oxidized proteins, DNA, and lipids) and serum of HCM patients. In addition, increased mitochondrial ROS production and changes in endogenous antioxidants are reported in HCM. Mutant sarcomeric protein may drive excessive levels of cardiac ROS via changes in cardiac efficiency and metabolism, mitochondrial activation and/or dysfunction, impaired protein quality control, and microvascular dysfunction. Future Directions: Interventions restoring metabolism, mitochondrial function, and improved ROS balance may be promising therapeutic approaches. We discuss the effects of current HCM pharmacological therapies and potential future therapies to prevent and reverse HCM. Antioxid. Redox Signal. 31, 318-358.

Keywords: hypertrophic cardiomyopathy; mitochondrion; pathophysiological mechanism; reactive oxygen species; redox state; sarcomeric gene mutation.

FIG. 1.

Cardiac muscle cell activation and role of sarcomeres. Contraction is initiated on Ca²⁺ entry in the muscle cell, which activates Ca²⁺-release from the SR. Ca²⁺ binds to the myofilaments, which causes contraction. To relax Ca²⁺ detaches and is pumped back into the SR via the SR-Ca²⁺-ATPase pump SERCA. During increased cardiac stress (exercise), Ca²⁺ detachment from the myofilaments is increased via reduced myofilament Ca²⁺-sensitivity by activation of β₁-AR and via cAMP of PKA. Activation of β₃-AR increases the activity of PKG via cGMP, which also reduces myofilament Ca²⁺-sensitivity. The inset shows the composition of the myofilaments, with the most affected HCM sarcomeric proteins cardiac myosin-binding protein C, myosin heavy chain, and troponin T. β₁-AR, β₁-adrenergic receptors; cAMP, cyclic AMP; cGMP, cyclic GMP; HCM, hypertrophic cardiomyopathy; PKA, protein kinase A; PKG, protein kinase G; SR, sarcoplasmic reticulum. Color images are available online.

FIG. 2.

Decline in the protein quality control system. Due to an age-related decline in the PQC system, mutant protein levels may increase above a toxic threshold, while levels of normal proteins decrease. This imbalance in protein composition may alter cardiac function, which will trigger a secondary disease-related decline in the PQC system further aggravating cardiac dysfunction and remodeling. Left: Cardiac MRI image of a mutation carrier without a phenotype (G+/Ph−). Right: Cardiac MRI image of a symptomatic HCM patient. MRI, magnetic resonance imaging; PQC, protein-quality-control. Color images are available online.

FIG. 3.

Proposed mutation-induced cycle in hypertrophic cardiomyopathy. Mutant protein increases energetic costs for sarcomere contraction and thereby impairs efficiency of cardiac performance. High energetic cost of sarcomere contraction may also be caused by increased sensitivity of the sarcomeres to Ca²⁺ (i.e., increased myofilament Ca²⁺-sensitivity), which is a hallmark of HCM. The high energetic costs for sarcomere contraction will cause energy depletion and thereby affect other energy-consuming processes in the cell involved in the maintenance of intracellular Ca²⁺-handling, metabolic substrate levels, and mitochondrial function. Perturbations in intracellular [Ca²⁺], metabolites, and mitochondrial function will impair diastolic function, which will cause hypoperfusion of the heart. Reduced cardiac perfusion, together with mitochondrial dysfunction, will reduce energy supply to the heart. Reduced energy supply will inhibit the PQC system resulting in an accumulation of mutant relative to normal protein. Disease-related derailments in the PQC system will exacerbate disease by the imbalance in sarcomere protein composition. In addition, hypoperfusion of the heart will trigger cell death and initiate secondary remodeling and dysfunction of endothelial and muscle cells, and will trigger maladaptive processes such as an increased adrenergic drive, post-translation protein modifications, cellular hypertrophy, reduced myofibril density, and fibrosis. Color images are available online.

FIG. 4.

Impaired efficiency of cardiac contraction at early stage of the disease. Imaging studies in mutation carriers and patients with obstructive HCM revealed reduced efficiency of cardiac performance at an early stage of the disease before onset of hypertrophy. The energy deficiency was larger in carriers of MYH7 mutation compared with MYBPC3 mutation carriers. By use of cardiovascular MRI, left ventricular volumes and mass were defined to calculate myocardial external efficiency, that is, the ratio between external work and myocardial oxygen consumption. Left: Cardiac MRI image of a mutation carrier without a phenotype. Right: Cardiac MRI image of a symptomatic HCM patient. Figure has been adapted from Witjas-Paalberends et al. (280) and Güçlü et al. (86) with permission (Oxford University Press and Wolters Kluwer). MYBPC3, gene encoding cardiac myosin-binding protein-C; MYH7, gene encoding β-myosin heavy chain. Color images are available online.

FIG. 5.

Defects at the myofilament level in hypertrophic cardiomyopathy. At the myofilament level, sarcomere mutations were shown to (A) increase tension cost (i.e., reduce efficiency of sarcomere contraction) and (B) increase myofilament Ca²⁺-sensitivity (indicated by the arrow). (C) Maximal force-generating capacity (black arrow) and myofilament Ca²⁺-sensitivity (white arrow) increase on stretching from short (1.8 μm) to long (2.2 μm) sarcomere lengths for the donor. Sarcomere mutations (MYH7_mis and TNNI3_mis) impair this length-dependent myofilament activation. (A) Adapted from Witjas-Paalberends et al. (280) with permission (Oxford University Press). (B) Adapted from Sequeira et al. (225) with permission. (C) Donor has been adapted from Sequeira et al. (225) with permission. (C) (MYH7_mis and TNNI3_mis) original data. Color images are available online.

FIG. 6.

Excitation/contraction energetics coupling in healthy cardiomyocytes. Left, Ca²⁺-induced Ca²⁺-release from the SR increases cytosolic [Ca^2±], leading to activation of myofilaments. Ca²⁺-release from myofilaments allows for myofilament relaxation. Right, Zoomed view of the creatine (Cr) and phosphocreatine (PCr) export pathway in healthy cardiomyocytes. Mitochondrial ATP synthase regenerates ATP from ADP, which via mitochondrial CK the phosphoryl group in ATP is used to generate PCr inside the mitochondria. Muscle CK uses the shuttled PCr to rapidly regenerate ATP from ADP at the myofilaments. CK, creatine kinase; NCX, Na⁺/Ca²⁺ exchanger. Color images are available online.

FIG. 7.

Excitation/contraction energetics coupling in hypertrophic cardiomyopathy cardiomyocytes. Left, HCM-causing mutations lead to higher myofilament ATPase-activity in cardiomyocytes, thereby enhancing cellular [ADP]. Defective CK and mitochondria function are unable to rapidly regenerate ATP. Elevated [ADP] leads to high myofilament Ca²⁺-sensitivity and Ca²⁺-buffering (sticky myofilaments, designed as a fishnet trapping Ca²⁺ at the myofilaments). Right, Zoomed view of the creatine (Cr)/phosphocreatine (PCr) export pathway in HCM cardiomyocytes. Impaired mitochondrial function in concert with reduced CK function and low PCr amount limits the capacity of the Cr-PCr pathway to buffer ATP, leading to ADP accumulation at the myofilaments. Color images are available online.

FIG. 8.

Effects of pathological ADP levels on cardiomyocyte function. Myofilament Ca²⁺-tension in HCM and nonfailing donor membrane-permeabilized cardiomyocytes. Ca²⁺-tension of HCM cardiomyocytes was measured in the presence of pathological ADP levels (100 μM ADP), while ADP was absent in donor cardiomyocytes, mimicking in vivo intracellular milieu. Gray panel depicts the free [Ca²⁺] range in in vivo cardiomyocytes (i.e., ∼0.15 diastolic to 1.6 μM systolic [Ca²⁺]). Cardiomyocyte Ca²⁺-tension is corrected for myofibril density. HCM cardiomyopathy tissue is hypercontractile versus donors at the free [Ca²⁺] in vivo range. Figure has been adapted from Sequeira et al. (225) and Sequeira et al. (224) with permission. Color images are available online.

FIG. 9.

Interaction between myofilament Ca²⁺-sensitivity, mutant protein, and secondary disease effects. (A) Activation of β-adrenergic receptors desensitizes the myofilaments for Ca²⁺ via PKA-mediated phosphorylation of sarcomeric proteins. (B) A hallmark of HCM is increased myofilament Ca²⁺-sensitivity, directly caused by mutant protein, reduced PKA-mediated protein phosphorylation, oxidative protein modifications, and increased [ADP]. High Ca²⁺-sensitivity will increase energetic costs for contraction and limit relaxation of the heart. Schematic figure based on original data from Sequeira et al. (225) and Sequeira et al. (224) and Wilder et al. (277). Color images are available online.

FIG. 10.

Cellular reactive oxygen species production and damaging effects. (A) Four (one-electron) steps for oxygen reduction. Electron transfer to oxygen (O₂) generates superoxide radical anion (O₂^•−), which progressively forms hydrogen peroxide (H₂O₂) and the hydroxyl radical (HO^•). (B) ROS-mediated cardiomyocyte injury. O₂^•− and HO^• initiate myofilament and cytosolic protein damage with amino acid residues oxidized and degraded/deactivated. Lipid peroxidation of sarcolemma, SR, mitochondrial and nuclear membranes, disrupts phospholipids, and increases cellular permeability to ions and water. Mitochondrial membrane disruption additionally causes more ROS production. Nuclear and mtDNA can be oxidized, leading to strand breaks and transcription impairment. The O₂^•− anion is extremely reactive, nevertheless due to its poor lipid solubility cannot diffuse far within cell compartments. This contrasts with H₂O₂, which due to its high lipid solubility can diffuse through membranes and generate HO^• at specific sites. The HO^• is considered the most potent form of ROS. (C) DNA oxidative damage. Conversion of guanine to 8-oxodG by the HO^• radical is the most frequently observed damage to nucleotides. Addition of HO^• to the guanine backbone interferes with polymerases and/or allows nucleotide mispairing. Cellular levels of 8-oxodG are used as a marker to estimate the amount of oxidative damage to DNA in cells. (D) Lipid peroxidation. Lipid peroxidation initiates when hydrogen atoms, closed to the double bond of a (poly)unsaturated fatty acid from phospholipids, are extracted by HO^• (i.e., desaturation) forming a very unstable lipid radical. Rearrangements of the single electron change the molecular structure of the lipid radical. In the presence of O₂, the lipid radical chain reaction is propagated, forming a lipid peroxyl radical. Removal of hydrogen atoms from other (poly)unsaturated fatty acids forms lipid hydroperoxide. Eventually, lipid degradation occurs with formation of MDA and 4-hydroxyalkenal products. MDA appears in the urine and blood and its levels are used to assess lipid damage. ROS-induced damage to lipid membranes, including the sarcolemma, nucleus, sarcoplasmic reticulum, and the mitochondria, forms lipid radicals and lipid peroxides; lipid peroxidation of cellular membranes alters their structure, disrupts lipid bilayer integrity, and increases susceptibility to permeability of ions and water. 8-oxodG, 8-oxo-7-hydrodeoxyguanosine; MDA, malondialdehyde; mtDNA, mitochondrial DNA; ROS, reactive oxygen species. Color images are available online.

FIG. 11.

Mitochondrial electron transport chain. (A) Components of the electron transport chain. NADH dehydrogenase (complex I), cytochrome b–c1 complex (complex III), cytochrome c oxidase (complex IV), and ATP synthase (complex V) span the inner mitochondrial membrane. Reduced forms of NADH (complex I) and FAD(2H) donate electrons (e⁻) to the transport chain via complex I or complex II, respectively, which are sequentially transferred to electron carriers, including the lipid soluble CoQ, complex III, CytC, and complex IV. Complex IV accepts electrons from the electron transport chain and reduces O₂ into H₂O. As electrons pass through the electron transfer chain, protons (H⁺) are pumped across the mitochondrial matrix to the inner mitochondrial space (at complexes I, III, and IV; complex II lacks a proton pumping mechanism), responsible for establishing an electrochemical proton gradient at the inner mitochondrial membrane. The creation of the electrochemical proton gradient forces protons back inside the matrix at the complex V, which uses the proton gradient energy to regenerate ATP from ADP (and Pi). The electron transport chain is coupled to the rate of ATP regeneration by the electrochemical proton gradient-coupled oxidative phosphorylation. Under physiological conditions, approximately up to 5% of O₂ in cells is converted to ROS, with complexes I and III the main sites for ROS production. (B) Supercomplex formation in the presence of cardiolipin. Cardiolipin is a unique type of phospholipid only found on mitochondrial membranes with higher enrichment at the inner mitochondrial membrane (∼22%) with minute levels of enrichment at the outer mitochondrial membrane (∼3%). Cardiolipin promotes membrane curvature of the inner mitochondrial membrane and importantly restructures the electron transport chain complexes into supercomplexes to improve electron transfer efficiency. (C) Cardiolipin peroxidation. Due to the high (poly)unsaturated fatty acid content, cardiolipin is particularly vulnerable to lipid peroxidation, which reduces formation of supercomplexes. In addition, affinity of CytC, and potentially CoQ, to the inner mitochondrial membrane is lost. CytC release from the mitochondrial intermembrane compartments into the cytosol activates programmed cell death. (D) SS-31 cardioprotective effects of cardiolipin. SS-31 specifically targets and stabilizes cardiolipin's location at the inner mitochondrial membrane, likely facilitating the formation of mitochondrial supercomplexes, in combination with reductions of electron leakage (and ROS formation) with improved ATP regeneration. CoQ, coenzyme Q; CytC, cytochrome c. Color images are available online.

FIG. 12.

Sarcomeric mutations contribute to excessive levels of ROS and the HCM phenotype. Mutations in sarcomeric genes may affect the cardiac redox balance via multiple pathways. Sarcomeric mutations alter sarcomere function, which may affect mechanotransduction and increase energetic cost and mitochondrial activity. In addition, expression of mutant proteins may contribute to UPS dysfunction, which might result in ER stress. These processes affect the cardiac redox balance. Other mechanisms that are likely to contribute to cardiac oxidative stress in HCM are microvascular dysfunction causing local ischemia, mitochondrial dysfunction, a strong adrenergic drive, activation of the angiotensin II (Ang II)-dependent pathway, changes in antioxidant capacity, and possibly an increased xanthine oxidase activity. Excessive levels of cardiac ROS in HCM lead to oxidative modifications of sarcomeric proteins, which may contribute to diastolic dysfunction. It may also activate the MAPK family, which has been linked to cardiac fibrosis and hypertrophy. In addition, oxidation of Ca²⁺/calmodulin kinase II (CaMKII) may contribute to its sustained activation, which slows down L-type Ca²⁺ current inactivation and increases the late Na⁺ current amplitude. This contributes to action potential duration prolongation and related arrhythmias. Lipid peroxidation and DNA oxidation may cause cell damage and/or effect cell signaling. This shows that a disturbed cardiac redox state has a central role in HCM pathology. ER, endoplasmic reticulum; ERK, extracellular signal-regulated protein kinases; GSH, reduced glutathione; JNK, c-Jun N-terminal protein kinases; MAPK, mitogen-activated protein kinases; MyHC, myosin heavy chain; NOS, nitric oxide synthase; UA, uric acid; UPS, ubiquitin/proteasome system. Color images are available online.

FIG. 13.

Breaking the mutation-induced cycle in hypertrophic cardiomyopathy. The vicious cycle may be broken by interventions to target metabolism (metabolic therapy), to lower myofilament Ca²⁺-sensitivity (e.g., stimulation of β₃AR, β₃AR agonist) and thereby reduce sarcomeric energy consumption, by interventions targeted at the mitochondria to reduce oxidative stress and improve energy supply, and by components that boost the PQC system and restore protein balance in the cell. HSPs, heat shock proteins. Color images are available online.