HDAC Inhibition Regulates Cardiac Function by Increasing Myofilament Calcium Sensitivity and Decreasing Diastolic Tension
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SUBMITTER: Eaton D
PROVIDER: S-EPMC9323146 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:RationalePreviously, we demonstrated that a deoxycorticosterone acetate (DOCA)-salt hypertensive mouse model produces cardiac oxidative stress and diastolic dysfunction with preserved systolic function. Oxidative stress has been shown to increase late inward sodium current (I(Na)), reducing the net cytosolic Ca(2+) efflux.ObjectiveOxidative stress in the DOCA-salt model may increase late I(Na), resulting in diastolic dysfunction amenable to treatment with ranolazine.Methods and resultsEchocardiography detected evidence of diastolic dysfunction in hypertensive mice that improved after treatment with ranolazine (E/E':sham, 31.9 ± 2.8, sham+ranolazine, 30.2 ± 1.9, DOCA-salt, 41.8 ± 2.6, and DOCA-salt+ranolazine, 31.9 ± 2.6; P=0.018). The end-diastolic pressure-volume relationship slope was elevated in DOCA-salt mice, improving to sham levels with treatment (sham, 0.16 ± 0.01 versus sham+ranolazine, 0.18 ± 0.01 versus DOCA-salt, 0.23 ± 0.2 versus DOCA-salt+ranolazine, 0.17 ± 0.0 1 mm Hg/L; P<0.005). DOCA-salt myocytes demonstrated impaired relaxation, τ, improving with ranolazine (DOCA-salt, 0.18 ± 0.02, DOCA-salt+ranolazine, 0.13 ± 0.01, sham, 0.11 ± 0.01, sham+ranolazine, 0.09 ± 0.02 seconds; P=0.0004). Neither late I(Na) nor the Ca(2+) transients were different from sham myocytes. Detergent extracted fiber bundles from DOCA-salt hearts demonstrated increased myofilament response to Ca(2+) with glutathionylation of myosin binding protein C. Treatment with ranolazine ameliorated the Ca(2+) response and cross-bridge kinetics.ConclusionsDiastolic dysfunction could be reversed by ranolazine, probably resulting from a direct effect on myofilaments, indicating that cardiac oxidative stress may mediate diastolic dysfunction through altering the contractile apparatus.
| S-EPMC3314887 | biostudies-literature
Project description:Aberrant myofilament Ca(2+) sensitivity is commonly observed with multiple cardiac diseases, especially familial cardiomyopathies. Although the etiology of the cardiomyopathies remains unclear, improving cardiac muscle Ca(2+) sensitivity through either pharmacological or genetic approaches shows promise of alleviating the disease-related symptoms. Due to its central role as the Ca(2+) sensor for cardiac muscle contraction, troponin C (TnC) stands out as an obvious and versatile target to reset disease-associated myofilament Ca(2+) sensitivity back to normal. To test the hypothesis that aberrant myofilament Ca(2+) sensitivity and its related function can be corrected through rationally engineered TnC constructs, three thin filament protein modifications representing different proteins (troponin I or troponin T), modifications (missense mutation, deletion, or truncation), and disease subtypes (familial or acquired) were studied. A fluorescent TnC was utilized to measure Ca(2+) binding to TnC in the physiologically relevant biochemical model system of reconstituted thin filaments. Consistent with the pathophysiology, the restrictive cardiomyopathy mutation, troponin I R192H, and ischemia-induced truncation of troponin I (residues 1-192) increased the Ca(2+) sensitivity of TnC on the thin filament, whereas the dilated cardiomyopathy mutation, troponin T ΔK210, decreased the Ca(2+) sensitivity of TnC on the thin filament. Rationally engineered TnC constructs corrected the abnormal Ca(2+) sensitivities of the thin filament, reconstituted actomyosin ATPase activity, and force generation in skinned trabeculae. Thus, the present study provides a novel and versatile therapeutic strategy to restore diseased cardiac muscle Ca(2+) sensitivity.
| S-EPMC3370186 | biostudies-literature
Project description:Tropomyosin (TM) plays a central role in calcium mediated striated muscle contraction. There are three muscle TM isoforms: alpha-TM, beta-TM, and gamma-TM. alpha-TM is the predominant cardiac and skeletal muscle isoform. beta-TM is expressed in skeletal and embryonic cardiac muscle. gamma-TM is expressed in slow-twitch musculature, but is not found in the heart. Our previous work established that muscle TM isoforms confer different physiological properties to the cardiac sarcomere. To determine whether one of these isoforms is dominant in dictating its functional properties, we generated single and double transgenic mice expressing beta-TM and/or gamma-TM in the heart, in addition to the endogenously expressed alpha-TM. Results show significant TM protein expression in the betagamma-DTG hearts: alpha-TM: 36%, beta-TM: 32%, and gamma-TM: 32%. These betagamma-DTG mice do not develop pathological abnormalities; however, they exhibit a hyper contractile phenotype with decreased myofilament calcium sensitivity, similar to gamma-TM transgenic hearts. Biophysical studies indicate that gamma-TM is more rigid than either alpha-TM or beta-TM. This is the first report showing that with approximately equivalent levels of expression within the same tissue, there is a functional dominance of gamma-TM over alpha-TM or beta-TM in regulating physiological performance of the striated muscle sarcomere. In addition to the effect expression of gamma-TM has on Ca(2+) activation of the cardiac myofilaments, our data demonstrates an effect on cooperative activation of the thin filament by strongly bound rigor cross-bridges. This is significant in relation to current ideas on the control mechanism of the steep relation between Ca(2+) and tension.
| S-EPMC3805252 | biostudies-literature
Project description:ObjectiveCardiac troponin I (cTnI) is an essential physiological and pathological regulator of cardiac relaxation. Significant to this regulation, the post-translational modification of cTnI through phosphorylation functions as a key mechanism to accelerate myofibril relaxation. Similar to phosphorylation, post-translational modification by acetylation alters amino acid charge and protein function. Recent studies have demonstrated that the acetylation of cardiac myofibril proteins accelerates relaxation and that cTnI is acetylated in the heart. These findings highlight the potential significance of myofilament acetylation; however, it is not known if site-specific acetylation of cTnI can lead to changes in myofilament, myofibril, and/or cellular mechanics. The objective of this study was to determine the effects of mimicking acetylation at a single site of cTnI (lysine-132; K132) on myofilament, myofibril, and cellular mechanics and elucidate its influence on molecular function.MethodsTo determine if pseudo-acetylation of cTnI at 132 modulates thin filament regulation of the acto-myosin interaction, we reconstituted thin filaments containing WT or K132Q (to mimic acetylation) cTnI and assessed in vitro motility. To test if mimicking acetylation at K132 alters cellular relaxation, adult rat ventricular cardiomyocytes were infected with adenoviral constructs expressing either cTnI K132Q or K132 replaced with arginine (K132R; to prevent acetylation) and cell shortening and isolated myofibril mechanics were measured. Finally, to confirm that changes in cell shortening and myofibril mechanics were directly due to pseudo-acetylation of cTnI at K132, we exchanged troponin containing WT or K132Q cTnI into isolated myofibrils and measured myofibril mechanical properties.ResultsReconstituted thin filaments containing K132Q cTnI exhibited decreased calcium sensitivity compared to thin filaments reconstituted with WT cTnI. Cardiomyocytes expressing K132Q cTnI had faster relengthening and myofibrils isolated from these cells had faster relaxation along with decreased calcium sensitivity compared to cardiomyocytes expressing WT or K132R cTnI. Myofibrils exchanged with K132Q cTnI ex vivo demonstrated faster relaxation and decreased calcium sensitivity.ConclusionsOur results indicate for the first time that mimicking acetylation of a specific cTnI lysine accelerates myofilament, myofibril, and myocyte relaxation. This work underscores the importance of understanding how acetylation of specific sarcomeric proteins affects cardiac homeostasis and disease and suggests that modulation of myofilament lysine acetylation may represent a novel therapeutic target to alter cardiac relaxation.
| S-EPMC7363438 | biostudies-literature
Project description:We investigated the mechanisms associated with E22K mutation in myosin regulatory light chain (RLC), found to cause hypertrophic cardiomyopathy (HCM) in humans and mice. Specifically, we characterized the mechanical profiles of papillary muscle fibers from transgenic mice expressing human ventricular RLC wild-type (Tg-WT) or E22K mutation (Tg-E22K). Because the two mouse models expressed different amounts of transgene, the B6SJL mouse line (NTg) was used as an additional control. Mechanical experiments were carried out on Ca2+ - and ATP-activated fibers and in rigor. Sinusoidal analysis was performed to elucidate the effect of E22K on tension and stiffness during activation/rigor, tension-pCa, and myosin cross-bridge (CB) kinetics. We found significant reductions in active tension (by 54%) and stiffness (active by 40% and rigor by 54%). A decrease in the Ca2+ sensitivity of tension (by ∆pCa ~ 0.1) was observed in Tg-E22K compared with Tg-WT fibers. The apparent (=measured) rate constant of exponential process B (2πb: force generation step) was not affected by E22K, but the apparent rate constant of exponential process C (2πc: CB detachment step) was faster in Tg-E22K compared with Tg-WT fibers. Both 2πb and 2πc were smaller in NTg than in Tg-WT fibers, suggesting a kinetic difference between the human and mouse RLC. Our results of E22K-induced reduction in myofilament stiffness and tension suggest that the main effect of this mutation was to disturb the interaction of RLC with the myosin heavy chain and impose structural abnormalities in the lever arm of myosin CB. When placed in vivo, the E22K mutation is expected to result in reduced contractility and decreased cardiac output whereby leading to HCM.Sub-disciplineBioenergetics.DatabaseThe data that support the findings of this study are available from the corresponding authors upon reasonable request.Animal protocolBK20150353 (Soochow University).Research governanceSchool of Nursing: Hua-Gang Hu: seuboyh@163.com; Soochow University: Chen Ge chge@suda.edu.cn.
| S-EPMC8668163 | biostudies-literature
Project description:This study determined whether deficiency of ovarian estrogen starting very early in life promoted age-associated Ca(2+) dysregulation and contractile dysfunction in isolated ventricular myocytes. Myocytes were isolated from anesthetized C57BL/6 female mice. Animals received an ovariectomy or sham-operation at one month and were aged to ~24 months. Excitation-contraction coupling parameters were compared in fura-2 loaded myocytes (37°C). While Ca(2+) transients were larger and faster in field-stimulated myocytes from ovariectomized mice, ovariectomy had no effect on peak fractional shortening. Similarly, ovariectomy had no effect on fractional shortening measured in vivo by echocardiography (values were 60.5 ± 2.9 vs. 60.3 ± 2.5% in sham and ovariectomized, respectively; n=5 mice/group). Ovariectomy did decrease myofilament Ca(2+) sensitivity, as evidenced by a 26% increase in the Ca(2+) required to activate actomyosin MgATPase in ovariectomized hearts. Larger Ca(2+) transients were attributable to a 48% increase in peak Ca(2+) current, along with an increase in the amplitude, width and frequency of Ca(2+) sparks measured in fluo-4 loaded myocytes. These changes in Ca(2+) handling were not due to increased expression of Ca(2+) channels (Cav1.2), sarcoplasmic reticulum Ca(2+) ATPase (SERCA2) or Na(+)-Ca(2+) exchanger in ovariectomized hearts. However, ovariectomy increased sarcoplasmic reticulum Ca(2+) stores by ~90% and promoted spontaneous Ca(2+) release from the sarcoplasmic reticulum when compared to sham controls. These observations demonstrate that long-term ovariectomy promotes intracellular Ca(2+) dysregulation, reduces myofilament Ca(2+) sensitivity and increases spontaneous Ca(2+) release in the aging female heart.
| S-EPMC3776741 | biostudies-literature
Project description:Heart failure with preserved ejection fraction (HFpEF) is a major health problem without effective therapies. This study assessed the effects of histone deacetylase (HDAC) inhibition on cardiopulmonary structure, function, and metabolism in a large mammalian model of pressure overload recapitulating features of diastolic dysfunction common to human HFpEF. Male domestic short-hair felines (n = 31, aged 2 months) underwent a sham procedure (n = 10) or loose aortic banding (n = 21), resulting in slow-progressive pressure overload. Two months after banding, animals were treated daily with suberoylanilide hydroxamic acid (b + SAHA, 10 mg/kg, n = 8), a Food and Drug Administration-approved pan-HDAC inhibitor, or vehicle (b + veh, n = 8) for 2 months. Echocardiography at 4 months after banding revealed that b + SAHA animals had significantly reduced left ventricular hypertrophy (LVH) (P < 0.0001) and left atrium size (P < 0.0001) versus b + veh animals. Left ventricular (LV) end-diastolic pressure and mean pulmonary arterial pressure were significantly reduced in b + SAHA (P < 0.01) versus b + veh. SAHA increased myofibril relaxation ex vivo, which correlated with in vivo improvements of LV relaxation. Furthermore, SAHA treatment preserved lung structure, compliance, blood oxygenation, and reduced perivascular fluid cuffs around extra-alveolar vessels, suggesting attenuated alveolar capillary stress failure. Acetylation proteomics revealed that SAHA altered lysine acetylation of mitochondrial metabolic enzymes. These results suggest that acetylation defects in hypertrophic stress can be reversed by HDAC inhibitors, with implications for improving cardiac structure and function in patients.
| S-EPMC7065257 | biostudies-literature
Project description:BackgroundDiastolic dysfunction (DD) is associated with the development of heart failure and contributes to the pathogenesis of other cardiac maladies, including atrial fibrillation. Inhibition of histone deacetylases (HDACs) has been shown to prevent DD by enhancing myofibril relaxation. We addressed the therapeutic potential of HDAC inhibition in a model of established DD with preserved ejection fraction.MethodsFour weeks after uninephrectomy and implantation with deoxycorticosterone acetate pellets, when DD was clearly evident, 1 cohort of mice was administered the clinical-stage HDAC inhibitor ITF2357/Givinostat. Echocardiography, blood pressure measurements, and end point invasive hemodynamic analyses were performed. Myofibril mechanics and intact cardiomyocyte relaxation were assessed ex vivo. Cardiac fibrosis was evaluated by picrosirius red staining and second harmonic generation microscopy of left ventricle (LV) sections, RNA sequencing of LV mRNA, mass spectrometry-based evaluation of decellularized LV biopsies, and atomic force microscopy determination of LV stiffness. Mechanistic studies were performed with primary rat and human cardiac fibroblasts.ResultsHDAC inhibition normalized DD without lowering blood pressure in this model of systemic hypertension. In contrast to previous models, myofibril relaxation was unimpaired in uninephrectomy/deoxycorticosterone acetate mice. Furthermore, cardiac fibrosis was not evident in any mouse cohort on the basis of picrosirius red staining or second harmonic generation microscopy. However, mass spectrometry revealed induction in the expression of >100 extracellular matrix proteins in LVs of uninephrectomy/deoxycorticosterone acetate mice, which correlated with profound tissue stiffening based on atomic force microscopy. ITF2357/Givinostat treatment blocked extracellular matrix expansion and LV stiffening. The HDAC inhibitor was subsequently shown to suppress cardiac fibroblast activation, at least in part, by blunting recruitment of the profibrotic chromatin reader protein BRD4 (bromodomain-containing protein 4) to key gene regulatory elements.ConclusionsThese findings demonstrate the potential of HDAC inhibition as a therapeutic intervention to reverse existing DD and establish blockade of extracellular matrix remodeling as a second mechanism by which HDAC inhibitors improve ventricular filling. Our data reveal the existence of pathophysiologically relevant covert or hidden cardiac fibrosis that is below the limit of detection of histochemical stains such as picrosirius red, highlighting the need to evaluate fibrosis of the heart using diverse methodologies.
| S-EPMC8884170 | biostudies-literature
Project description:Troponin I (TnI) variant Pro82Ser (cTnIP82S) was initially considered a disease-causing mutation; however, later studies suggested the contrary. We tested the hypothesis of whether a causal link exists between cTnIP82S and cardiac structural and functional remodeling, such as during aging or chronic pressure overload. A cardiac-specific transgenic (Tg) mouse model of cTnIP82S was created to test this hypothesis. During aging, Tg cTnIP82S displayed diastolic dysfunction, characterized by longer isovolumetric relaxation time, and impaired ejection and relaxation time. In young, Tg mice in vivo pressure-volume loops and intact trabecular preparations revealed normal cardiac contractility at baseline. However, upon β-adrenergic stimulation, a blunted contractile reserve and no hastening in left ventricle relaxation were evident in vivo, whereas, in isolated muscles, Ca(2+) transient amplitude isoproterenol dose-response was blunted. In addition, when exposed to chronic pressure overload, Tg mice show exacerbated hypertrophy and decreased contractility compared with age-matched non-Tg littermates. At the molecular level, this mutation significantly impairs myofilament cooperative activation. Importantly, this occurs in the absence of alterations in TnI or myosin-binding protein C phosphorylation. The cTnIP82S variant occurs near a region of interactions with troponin T; therefore, structural changes in this region could explain its meaningful effects on myofilament cooperativity. Our data indicate that cTnIP82S mutation modifies age-dependent diastolic dysfunction and impairs overall contractility after β-adrenergic stimulation or chronic pressure overload. Thus cTnIP82S variant should be regarded as a disease-modifying factor for dysfunction and adverse remodeling with aging and chronic pressure overload.
| S-EPMC4297775 | biostudies-literature
Project description:AMP-activated protein kinase (AMPK) is an energy-sensing enzyme central to the regulation of metabolic homeostasis. In the heart AMPK is activated during cardiac stress-induced ATP depletion and functions to stimulate metabolic pathways that restore the AMP/ATP balance. Recently it was demonstrated that AMPK phosphorylates cardiac troponin I (cTnI) at Ser-150 in vitro. We sought to determine if the metabolic regulatory kinase AMPK phosphorylates cTnI at Ser-150 in vivo to alter cardiac contractile function directly at the level of the myofilament. Rabbit cardiac myofibrils separated by two-dimensional isoelectric focusing subjected to a Western blot with a cTnI phosphorylation-specific antibody demonstrates that cTnI is endogenously phosphorylated at Ser-150 in the heart. Treatment of myofibrils with the AMPK holoenzyme increased cTnI Ser-150 phosphorylation within the constraints of the muscle lattice. Compared with controls, cardiac fiber bundles exchanged with troponin containing cTnI pseudo-phosphorylated at Ser-150 demonstrate increased sensitivity of calcium-dependent force development, blunting of both PKA-dependent calcium desensitization, and PKA-dependent increases in length dependent activation. Thus, in addition to the defined role of AMPK as a cardiac metabolic energy gauge, these data demonstrate AMPK Ser-150 phosphorylation of cTnI directly links the regulation of cardiac metabolic demand to myofilament contractile energetics. Furthermore, the blunting effect of cTnI Ser-150 phosphorylation cross-talk can uncouple the effects of myofilament PKA-dependent phosphorylation from β-adrenergic signaling as a novel thin filament contractile regulatory signaling mechanism.
| S-EPMC3365946 | biostudies-literature