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Wu T, Huang T, Ren H, Shen C, Qian J, Fu X, Liu S, Xie C, Lin X, Wan J, Xiong S, Ji Y, Liu M, Zheng H, Liang T, Liu W, Zou Y, Lai K, Yang M, Song Z, Lan P, Li X, Wu Y, Yang M, Li H, Huang X, Chen H, Tan J, Cai W. Metabolic Coordination Structures Contribute to Diabetic Myocardial Dysfunction. Circ Res 2025; 136:946-967. [PMID: 40190276 DOI: 10.1161/circresaha.124.326044] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/17/2024] [Revised: 03/11/2025] [Accepted: 03/14/2025] [Indexed: 04/26/2025]
Abstract
BACKGROUND Individuals with diabetes are susceptible to cardiac dysfunction and heart failure, potentially resulting in mortality. Metabolic disorders frequently occur in patients with diabetes, and diabetes usually leads to remodeling of heart structure and cardiac dysfunction. However, the contribution and underlying mechanisms of metabolic and structural coupling in diabetic cardiac dysfunction remain elusive. METHODS Two mouse models of type 2 diabetes (T2DM) were used to assess alterations in glucose/lipid metabolism and cardiac structure. The potential metabolic-structural coupling molecule ACBP (acyl-coenzyme A-binding protein) was screened from 4 published datasets of T2DM-associated heart disease. In vivo loss-of-function and gain-of-function approaches were used to investigate the role of ACBP in diabetic cardiac dysfunction. The underlying mechanisms of metabolic and structural coupling were investigated by stable-isotope tracing metabolomics, coimmunoprecipitation coupled with mass spectrometry, and chromatin immunoprecipitation sequencing. RESULTS Diabetic mouse hearts exhibit enhanced lipid metabolism and impaired ultrastructure with marked cardiac systolic and diastolic dysfunction. Analysis of 4 T2DM public datasets revealed that Acbp was a significant lipid metabolism gene whose expression was upregulated. Consistently, ACBP expression levels were markedly elevated in the hearts of patients with diabetes and diabetic mice. Moreover, we constructed cardiomyocyte-specific Acbp knockout mice that exhibited attenuation of T2DM-induced cardiac remodeling and cardiac dysfunction, including attenuation of cardiac hypertrophy, fibrosis, ultrastructural damage, and enhanced cardiomyocyte contractility and cardiac function. Conversely, cardiac-specific Acbp overexpression via adeno-associated virus type 9, which encodes Acbp under the cTnT (cardiac troponin T) promoter, recapitulated cardiac dysfunction. Mechanistically, cardiac-specific Acbp knockout enhances glucose utilization in diabetic cardiomyocytes, suggesting a potential compensatory mechanism for insufficient ATP levels, highlighting its metabolic role. In addition, combined with mass spectrometry analysis revealed that ACBP binds MyBPC3 (myosin-binding protein C3) in T2DM individuals, which potentially prevents MyBPC3 from assisting the formation of cross-bridge structures between myosin and actin, thereby impairing myocardial contraction. Importantly, chromatin immunoprecipitation sequencing revealed that peroxisome proliferator-activated receptor γ regulates the transcriptional activity of Acbp. CONCLUSIONS Our findings demonstrated that ACBP mediates the bidirectional regulation of cardiomyocyte metabolic and structural associations and identified a promising therapeutic target for ameliorating cardiac dysfunction in patients with T2DM.
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Affiliation(s)
- Teng Wu
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Reproductive Medicine Center, Sun Yat-sen Memorial Hospital (T.W., T.L., W.L., P.L., H.C.), Sun Yat-sen University, Guangzhou, China
| | - Tongsheng Huang
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Honglin Ren
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Conghui Shen
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Jiang Qian
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Xinlu Fu
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Shangyuan Liu
- GMU-GIBH Joint School of Life Sciences, The Guangdong-Hong Kong-Macau Joint Laboratory for Cell Fate Regulation and Diseases, Guangzhou Medical University, China (S.L., C.X.)
| | - Chengshu Xie
- GMU-GIBH Joint School of Life Sciences, The Guangdong-Hong Kong-Macau Joint Laboratory for Cell Fate Regulation and Diseases, Guangzhou Medical University, China (S.L., C.X.)
| | - Xi Lin
- ZEISS Microscopy Customer Center China, Shanghai (X. Lin)
| | - Junhong Wan
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Shijie Xiong
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Yuanjun Ji
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Mengying Liu
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Huiting Zheng
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Ting Liang
- Reproductive Medicine Center, Sun Yat-sen Memorial Hospital (T.W., T.L., W.L., P.L., H.C.), Sun Yat-sen University, Guangzhou, China
| | - Wenyi Liu
- Reproductive Medicine Center, Sun Yat-sen Memorial Hospital (T.W., T.L., W.L., P.L., H.C.), Sun Yat-sen University, Guangzhou, China
| | - Yan Zou
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Kingwai Lai
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Maoquan Yang
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Zeyi Song
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Peixuan Lan
- Reproductive Medicine Center, Sun Yat-sen Memorial Hospital (T.W., T.L., W.L., P.L., H.C.), Sun Yat-sen University, Guangzhou, China
| | - Xinghui Li
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Yandi Wu
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Ming Yang
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Hui Li
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Xuezhe Huang
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Hui Chen
- Reproductive Medicine Center, Sun Yat-sen Memorial Hospital (T.W., T.L., W.L., P.L., H.C.), Sun Yat-sen University, Guangzhou, China
| | - Jing Tan
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
| | - Weibin Cai
- Guangdong Engineering & Technology Research Center for Disease-Model Animals, Laboratory Animal Center (T.W., T.H., H.R., C.S., J.Q., X.F., J.W., S.X., Y.J., M.L., H.Z., Y.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, H.L., X.H., J.T., W.C.), Sun Yat-sen University, Guangzhou, China
- Department of Biochemistry, Zhongshan School of Medicine (T.W., T.H., H.R., C.S., J.W., S.X., Y.J., M.L., H.Z., K.L., Maoquan Yang, Z.S., X. Li, Y.W., Ming Yang, J.T., W.C.), Sun Yat-sen University, Guangzhou, China
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Ekhator ES, Fazzari M, Newman RH. Redox Regulation of cAMP-Dependent Protein Kinase and Its Role in Health and Disease. Life (Basel) 2025; 15:655. [PMID: 40283209 PMCID: PMC12029036 DOI: 10.3390/life15040655] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/06/2025] [Revised: 04/02/2025] [Accepted: 04/03/2025] [Indexed: 04/29/2025] Open
Abstract
Protein kinase A (PKA) is a key regulator of cellular signaling that regulates key physiological processes such as metabolism, cell proliferation, and neuronal function. While its activation by the second messenger 3',5'-cyclic adenosine triphosphate (cAMP) is well characterized, recent research highlights additional regulatory mechanisms, particularly oxidative post-translational modifications, that influence PKA's structure, activity, and substrate specificity. Both the regulatory and catalytic subunits of PKA are susceptible to redox modifications, which have been shown to play important roles in the regulation of key cellular functions, including cardiac contractility, lipid metabolism, and the immune response. Likewise, redox-dependent modulation of PKA signaling has been implicated in numerous diseases, including cardiovascular disorders, diabetes, and neurodegenerative conditions, making it a potential therapeutic target. However, the mechanisms of crosstalk between redox- and PKA-dependent signaling remain poorly understood. This review examines the structural and functional regulation of PKA, with a focus on redox-dependent modifications and their impact on PKA-dependent signaling. A deeper understanding of these mechanisms may provide new strategies for targeting oxidative stress in disease and restoring balanced PKA signaling in cells.
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Affiliation(s)
- Ese S. Ekhator
- Department of Biology, North Carolina A&T State University, Greensboro, NC 27411, USA;
- Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA 15261, USA;
| | - Marco Fazzari
- Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA 15261, USA;
| | - Robert H. Newman
- Department of Biology, North Carolina A&T State University, Greensboro, NC 27411, USA;
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Pietsch N, Chen CY, Kupsch S, Bacmeister L, Geertz B, Herrera-Rivero M, Siebels B, Voß H, Krämer E, Braren I, Westermann D, Schlüter H, Mearini G, Schlossarek S, van der Velden J, Caporizzo MA, Lindner D, Prosser BL, Carrier L. Chronic Activation of Tubulin Tyrosination Improves Heart Function. Circ Res 2024; 135:910-932. [PMID: 39279670 PMCID: PMC11465905 DOI: 10.1161/circresaha.124.324387] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/05/2024] [Revised: 08/28/2024] [Accepted: 09/03/2024] [Indexed: 09/18/2024]
Abstract
BACKGROUND Hypertrophic cardiomyopathy (HCM) is the most common cardiac genetic disorder caused by sarcomeric gene variants and associated with left ventricular hypertrophy and diastolic dysfunction. The role of the microtubule network has recently gained interest with the findings that microtubule detyrosination (dTyr-MT) is markedly elevated in heart failure. Acute reduction of dTyr-MT by inhibition of the detyrosinase (VASH [vasohibin]/SVBP [small VASH-binding protein] complex) or activation of the tyrosinase (TTL [tubulin tyrosine ligase]) markedly improved contractility and reduced stiffness in human failing cardiomyocytes and thus posed a new perspective for HCM treatment. In this study, we tested the impact of chronic tubulin tyrosination in an HCM mouse model (Mybpc3 knock-in), in human HCM cardiomyocytes, and in SVBP-deficient human engineered heart tissues (EHTs). METHODS Adeno-associated virus serotype 9-mediated TTL transfer was applied in neonatal wild-type rodents, in 3-week-old knock-in mice, and in HCM human induced pluripotent stem cell-derived cardiomyocytes. RESULTS We show (1) TTL for 6 weeks dose dependently reduced dTyr-MT and improved contractility without affecting cytosolic calcium transients in wild-type cardiomyocytes; (2) TTL for 12 weeks reduced the abundance of dTyr-MT in the myocardium, improved diastolic filling, compliance, cardiac output, and stroke volume in knock-in mice; (3) TTL for 10 days normalized cell area in HCM human induced pluripotent stem cell-derived cardiomyocytes; (4) TTL overexpression activated transcription of tubulins and other cytoskeleton components but did not significantly impact the proteome in knock-in mice; (5) SVBP-deficient EHTs exhibited reduced dTyr-MT levels, higher force, and faster relaxation than TTL-deficient and wild-type EHTs. RNA sequencing and mass spectrometry analysis revealed distinct enrichment of cardiomyocyte components and pathways in SVBP-deficient versus TTL-deficient EHTs. CONCLUSIONS This study provides the first proof of concept that chronic activation of tubulin tyrosination in HCM mice and in human EHTs improves heart function and holds promise for targeting the nonsarcomeric cytoskeleton in heart disease.
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Affiliation(s)
- Niels Pietsch
- Department of Experimental Pharmacology and Toxicology (N.P., B.G., E.K., G.M., S.S., L.C.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- German Centre for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Germany (N.P., D.W., G.M., S.S., D.L., L.C.)
| | - Christina Y. Chen
- Department of Physiology, Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA (C.Y.C., M.A.C., B.L.P.)
- Now with Gene Therapy Program, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA (C.Y.C.)
| | - Svenja Kupsch
- Department of Cardiology, University Heart and Vascular Center (S.K., L.B., D.W., D.L.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- Now with Institute of Experimental Cardiovascular Research, University Medical Center Hamburg-Eppendorf, Germany (S.K.)
| | - Lucas Bacmeister
- Department of Cardiology, University Heart and Vascular Center (S.K., L.B., D.W., D.L.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- Now with Faculty of Medicine, Department of Cardiology and Angiology, University Heart Center Freiburg-Bad Krozingen, University of Freiburg, Germany (L.B., D.W., D.L.)
| | - Birgit Geertz
- Department of Experimental Pharmacology and Toxicology (N.P., B.G., E.K., G.M., S.S., L.C.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Marisol Herrera-Rivero
- Department of Genetic Epidemiology, Institute of Human Genetics, University of Münster, Germany (M.H.-R.)
- Joint Institute for Individualisation in a Changing Environment, University of Münster and Bielefeld University, Münster, Germany (M.H.-R.)
| | - Bente Siebels
- Section Mass Spectrometric Proteomics (B.S., H.V., H.S.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Hannah Voß
- Section Mass Spectrometric Proteomics (B.S., H.V., H.S.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Elisabeth Krämer
- Department of Experimental Pharmacology and Toxicology (N.P., B.G., E.K., G.M., S.S., L.C.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Ingke Braren
- Vector Facility, Department of Experimental Pharmacology and Toxicology (I.B.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Dirk Westermann
- Department of Cardiology, University Heart and Vascular Center (S.K., L.B., D.W., D.L.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- Now with Faculty of Medicine, Department of Cardiology and Angiology, University Heart Center Freiburg-Bad Krozingen, University of Freiburg, Germany (L.B., D.W., D.L.)
- German Centre for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Germany (N.P., D.W., G.M., S.S., D.L., L.C.)
| | - Hartmut Schlüter
- Section Mass Spectrometric Proteomics (B.S., H.V., H.S.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Giulia Mearini
- Department of Experimental Pharmacology and Toxicology (N.P., B.G., E.K., G.M., S.S., L.C.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- German Centre for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Germany (N.P., D.W., G.M., S.S., D.L., L.C.)
| | - Saskia Schlossarek
- Department of Experimental Pharmacology and Toxicology (N.P., B.G., E.K., G.M., S.S., L.C.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- German Centre for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Germany (N.P., D.W., G.M., S.S., D.L., L.C.)
| | - Jolanda van der Velden
- Amsterdam University Medical Center, Vrije Universiteit Amsterdam, Department of Physiology, Amsterdam Cardiovascular Sciences, Amsterdam, the Netherlands (J.v.d.V.)
| | - Matthew A. Caporizzo
- Department of Physiology, Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA (C.Y.C., M.A.C., B.L.P.)
- Now with Department of Molecular Physiology and Biophysics, University of Vermont Larner College of Medicine, Burlington, VT (M.A.C.)
| | - Diana Lindner
- Department of Cardiology, University Heart and Vascular Center (S.K., L.B., D.W., D.L.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- Now with Faculty of Medicine, Department of Cardiology and Angiology, University Heart Center Freiburg-Bad Krozingen, University of Freiburg, Germany (L.B., D.W., D.L.)
- German Centre for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Germany (N.P., D.W., G.M., S.S., D.L., L.C.)
| | - Benjamin L. Prosser
- Department of Physiology, Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA (C.Y.C., M.A.C., B.L.P.)
| | - Lucie Carrier
- Department of Experimental Pharmacology and Toxicology (N.P., B.G., E.K., G.M., S.S., L.C.), University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- German Centre for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Germany (N.P., D.W., G.M., S.S., D.L., L.C.)
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4
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Ishizaka M, Katagiri K, Ogawa M, Hsu HH, Miyagawa Y, Takemura N. A pilot study of the proarrhythmic effects of pimobendan injection in clinically healthy cats. Vet Res Commun 2024; 48:3177-3186. [PMID: 39141288 DOI: 10.1007/s11259-024-10478-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2023] [Accepted: 07/21/2024] [Indexed: 08/15/2024]
Abstract
Pimobendan is not currently approved for use in cats, although its usefulness in feline hypertrophic cardiomyopathy has been suggested. Reports indicate an increase in arrhythmic events following oral administration to healthy cats. Given the greater potency of intravenous administration compared to oral intake, it is conceivable that the incidence of arrhythmias may be increased following pimobendan injection. Therefore, this study aimed to investigate the proarrhythmic effects of pimobendan injection in cats. Five clinically healthy cats underwent physical examination, echocardiography, blood pressure measurements, and 24-hour Holter electrocardiography immediately before and after receiving pimobendan as an intravenous bolus dose of 0.15 mg/kg twice daily for 3 days. Additionally, a 24-hour Holter electrocardiography recording was conducted on the third day of pimobendan or placebo IV administration to assess heart rate, arrhythmias, and heart rate variability. Following pimobendan administration, there was a significant increase in total 24-hour heart rate. Echocardiography revealed a significant increase in mitral valve annulus systolic velocity (S') on the ventricular septal wall side, indicative of enhanced contractility. Only one cat exhibited paroxysmal ventricular tachycardia and an increase in the frequency of arrhythmic events. Conversely, in the remaining cats, a decreasing trend in the number of arrhythmias was observed. These findings indicate that intravenous administration of pimobendan may not be implicated in the onset of arrhythmias. Nevertheless, further research is warranted to explore the effects of intravenous pimobendan administration in cats with myocardial disease.
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Affiliation(s)
- Mio Ishizaka
- Laboratory of Veterinary Internal Medicine II, School of Veterinary Medicine, Nippon Veterinary and Life Science University, 1-7-1 Kyonan-cho, Musashino-shi, 180-8602, Tokyo, Japan
| | - Keisuke Katagiri
- Honda Animal Hospital, 1-789-3 Hondacho, Midori-ku, Chiba-shi, 266-0005, Chiba, Japan
| | - Mizuki Ogawa
- Veterinary Medical Center, Graduate School of Agricultural and Life Science, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, 113-0032, Tokyo, Japan
| | - Huai-Hsun Hsu
- Laboratory of Veterinary Internal Medicine II, School of Veterinary Medicine, Nippon Veterinary and Life Science University, 1-7-1 Kyonan-cho, Musashino-shi, 180-8602, Tokyo, Japan
| | - Yuichi Miyagawa
- Laboratory of Veterinary Internal Medicine II, School of Veterinary Medicine, Nippon Veterinary and Life Science University, 1-7-1 Kyonan-cho, Musashino-shi, 180-8602, Tokyo, Japan.
| | - Naoyuki Takemura
- Laboratory of Veterinary Internal Medicine II, School of Veterinary Medicine, Nippon Veterinary and Life Science University, 1-7-1 Kyonan-cho, Musashino-shi, 180-8602, Tokyo, Japan
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5
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Arts T, Lyon A, Delhaas T, Kuster DWD, van der Velden J, Lumens J. Translating myosin-binding protein C and titin abnormalities to whole-heart function using a novel calcium-contraction coupling model. J Mol Cell Cardiol 2024; 190:13-23. [PMID: 38462126 DOI: 10.1016/j.yjmcc.2024.03.001] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/01/2023] [Revised: 01/15/2024] [Accepted: 03/01/2024] [Indexed: 03/12/2024]
Abstract
Mutations in cardiac myosin-binding protein C (cMyBP-C) or titin may respectively lead to hypertrophic (HCM) or dilated (DCM) cardiomyopathies. The mechanisms leading to these phenotypes remain unclear because of the challenge of translating cellular abnormalities to whole-heart and system function. We developed and validated a novel computer model of calcium-contraction coupling incorporating the role of cMyBP-C and titin based on the key assumptions: 1) tension in the thick filament promotes cross-bridge attachment mechanochemically, 2) with increasing titin tension, more myosin heads are unlocked for attachment, and 3) cMyBP-C suppresses cross-bridge attachment. Simulated stationary calcium-tension curves, isotonic and isometric contractions, and quick release agreed with experimental data. The model predicted that a loss of cMyBP-C function decreases the steepness of the calcium-tension curve, and that more compliant titin decreases the level of passive and active tension and its dependency on sarcomere length. Integrating this cellular model in the CircAdapt model of the human heart and circulation showed that a loss of cMyBP-C function resulted in HCM-like hemodynamics with higher left ventricular end-diastolic pressures and smaller volumes. More compliant titin led to higher diastolic pressures and ventricular dilation, suggesting DCM-like hemodynamics. The novel model of calcium-contraction coupling incorporates the role of cMyBP-C and titin. Its coupling to whole-heart mechanics translates changes in cellular calcium-contraction coupling to changes in cardiac pump and circulatory function and identifies potential mechanisms by which cMyBP-C and titin abnormalities may develop into HCM and DCM phenotypes. This modeling platform may help identify distinct mechanisms underlying clinical phenotypes in cardiac diseases.
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Affiliation(s)
- Theo Arts
- Department of Biomedical Engineering, Cardiovascular Research Center Maastricht (CARIM), Maastricht University, 6200MD Maastricht, the Netherlands.
| | - Aurore Lyon
- Department of Biomedical Engineering, Cardiovascular Research Center Maastricht (CARIM), Maastricht University, 6200MD Maastricht, the Netherlands
| | - Tammo Delhaas
- Department of Biomedical Engineering, Cardiovascular Research Center Maastricht (CARIM), Maastricht University, 6200MD Maastricht, the Netherlands
| | - Diederik W D Kuster
- Department of Physiology, Amsterdam University Medical Center, 1081HZ Amsterdam, the Netherlands
| | - Jolanda van der Velden
- Department of Physiology, Amsterdam University Medical Center, 1081HZ Amsterdam, the Netherlands
| | - Joost Lumens
- Department of Biomedical Engineering, Cardiovascular Research Center Maastricht (CARIM), Maastricht University, 6200MD Maastricht, the Netherlands
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6
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Kochurova AM, Beldiia EA, Nefedova VV, Ryabkova NS, Yampolskaya DS, Matyushenko AM, Bershitsky SY, Kopylova GV, Shchepkin DV. N-Terminal Fragment of Cardiac Myosin Binding Protein C Modulates Cooperative Mechanisms of Thin Filament Activation in Atria and Ventricles. BIOCHEMISTRY. BIOKHIMIIA 2024; 89:116-129. [PMID: 38467549 DOI: 10.1134/s0006297924010073] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/16/2023] [Revised: 12/09/2023] [Accepted: 12/11/2023] [Indexed: 03/13/2024]
Abstract
Cardiac myosin binding protein C (cMyBP-C) is one of the essential control components of the myosin cross-bridge cycle. The C-terminal part of cMyBP-C is located on the surface of the thick filament, and its N-terminal part interacts with actin, myosin, and tropomyosin, affecting both kinetics of the ATP hydrolysis cycle and lifetime of the cross-bridge, as well as calcium regulation of the actin-myosin interaction, thereby modulating contractile function of myocardium. The role of cMyBP-C in atrial contraction has not been practically studied. We examined effect of the N-terminal C0-C1-m-C2 (C0-C2) fragment of cMyBP-C on actin-myosin interaction using ventricular and atrial myosin in an in vitro motility assay. The C0-C2 fragment of cMyBP-C significantly reduced the maximum sliding velocity of thin filaments on both myosin isoforms and increased the calcium sensitivity of the actin-myosin interaction. The C0-C2 fragment had different effects on the kinetics of ATP and ADP exchange, increasing the affinity of ventricular myosin for ADP and decreasing the affinity of atrial myosin. The effect of the C0-C2 fragment on the activation of the thin filament depended on the myosin isoforms. Atrial myosin activates the thin filament less than ventricular myosin, and the C0-C2 fragment makes these differences in the myosin isoforms more pronounced.
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Affiliation(s)
- Anastasia M Kochurova
- Institute of Immunology and Physiology, Russian Academy of Sciences, Yekaterinburg, 620049, Russia
| | - Evgenia A Beldiia
- Institute of Immunology and Physiology, Russian Academy of Sciences, Yekaterinburg, 620049, Russia
- Ural Federal University, Ekaterinburg, 620002, Russia
| | - Victoria V Nefedova
- Bach Institute of Biochemistry, Research Center of Biotechnology, Russian Academy of Sciences, Moscow, 119071, Russia
| | - Natalia S Ryabkova
- Department of Biochemistry, Faculty of Biology, Lomonosov Moscow State University, Moscow, 119234, Russia
- HyTest Ltd., Turku, 20520, Finland
| | - Daria S Yampolskaya
- Bach Institute of Biochemistry, Research Center of Biotechnology, Russian Academy of Sciences, Moscow, 119071, Russia
| | - Alexander M Matyushenko
- Bach Institute of Biochemistry, Research Center of Biotechnology, Russian Academy of Sciences, Moscow, 119071, Russia
| | - Sergey Y Bershitsky
- Institute of Immunology and Physiology, Russian Academy of Sciences, Yekaterinburg, 620049, Russia
| | - Galina V Kopylova
- Institute of Immunology and Physiology, Russian Academy of Sciences, Yekaterinburg, 620049, Russia
| | - Daniil V Shchepkin
- Institute of Immunology and Physiology, Russian Academy of Sciences, Yekaterinburg, 620049, Russia.
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Escribá R, Larrañaga-Moreira JM, Richaud-Patin Y, Pourchet L, Lazis I, Jiménez-Delgado S, Morillas-García A, Ortiz-Genga M, Ochoa JP, Carreras D, Pérez GJ, de la Pompa JL, Brugada R, Monserrat L, Barriales-Villa R, Raya A. iPSC-Based Modeling of Variable Clinical Presentation in Hypertrophic Cardiomyopathy. Circ Res 2023; 133:108-119. [PMID: 37317833 DOI: 10.1161/circresaha.122.321951] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/06/2022] [Accepted: 06/01/2023] [Indexed: 06/16/2023]
Abstract
BACKGROUND Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiac disease and a frequent cause of heart failure and sudden cardiac death. Our understanding of the genetic bases and pathogenic mechanisms underlying HCM has improved significantly in the recent past, but the combined effect of various pathogenic gene variants and the influence of genetic modifiers in disease manifestation are very poorly understood. Here, we set out to investigate genotype-phenotype relationships in 2 siblings with an extensive family history of HCM, both carrying a pathogenic truncating variant in the MYBPC3 gene (p.Lys600Asnfs*2), but who exhibited highly divergent clinical manifestations. METHODS We used a combination of induced pluripotent stem cell (iPSC)-based disease modeling and CRISPR (clustered regularly interspersed short palindromic repeats)/Cas9 (CRISPR-associated protein 9)-mediated genome editing to generate patient-specific cardiomyocytes (iPSC-CMs) and isogenic controls lacking the pathogenic MYBPC3 variant. RESULTS Mutant iPSC-CMs developed impaired mitochondrial bioenergetics, which was dependent on the presence of the mutation. Moreover, we could detect altered excitation-contraction coupling in iPSC-CMs from the severely affected individual. The pathogenic MYBPC3 variant was found to be necessary, but not sufficient, to induce iPSC-CM hyperexcitability, suggesting the presence of additional genetic modifiers. Whole-exome sequencing of the mutant carriers identified a variant of unknown significance in the MYH7 gene (p.Ile1927Phe) uniquely present in the individual with severe HCM. We finally assessed the pathogenicity of this variant of unknown significance by functionally evaluating iPSC-CMs after editing the variant. CONCLUSIONS Our results indicate that the p.Ile1927Phe variant of unknown significance in MYH7 can be considered as a modifier of HCM expressivity when found in combination with truncating variants in MYBPC3. Overall, our studies show that iPSC-based modeling of clinically discordant subjects provides a unique platform to functionally assess the effect of genetic modifiers.
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Affiliation(s)
- Rubén Escribá
- Regenerative Medicine Program, Institut d'Investigació Biomèdica de Bellvitge - IDIBELL, L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Program for Clinical Translation of Regenerative Medicine in Catalonia - P-[CMRC], L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Center for Networked Biomedical Research on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Barcelona, Spain (R.E., Y.R.-P., L.P., A.R.)
| | - José M Larrañaga-Moreira
- Unidad de Cardiopatías Familiares, Servicio de Cardiología, Complexo Hospitalario Universitario de A Coruña, Servizo Galego de Saúde (SERGAS) (J.M.L.-M., R.B.-V.)
- Instituto de Investigación Biomédica de A Coruña (INIBIC), Universidade da Coruña, A Coruña, Spain (J.M.L.-M., M.O.-G., J.P.O., R.B.-V.)
| | - Yvonne Richaud-Patin
- Regenerative Medicine Program, Institut d'Investigació Biomèdica de Bellvitge - IDIBELL, L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Program for Clinical Translation of Regenerative Medicine in Catalonia - P-[CMRC], L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Center for Networked Biomedical Research on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Barcelona, Spain (R.E., Y.R.-P., L.P., A.R.)
| | - Léa Pourchet
- Regenerative Medicine Program, Institut d'Investigació Biomèdica de Bellvitge - IDIBELL, L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Program for Clinical Translation of Regenerative Medicine in Catalonia - P-[CMRC], L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Center for Networked Biomedical Research on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Barcelona, Spain (R.E., Y.R.-P., L.P., A.R.)
| | - Ioannis Lazis
- Regenerative Medicine Program, Institut d'Investigació Biomèdica de Bellvitge - IDIBELL, L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Program for Clinical Translation of Regenerative Medicine in Catalonia - P-[CMRC], L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
| | - Senda Jiménez-Delgado
- Regenerative Medicine Program, Institut d'Investigació Biomèdica de Bellvitge - IDIBELL, L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Program for Clinical Translation of Regenerative Medicine in Catalonia - P-[CMRC], L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
| | - Alba Morillas-García
- Regenerative Medicine Program, Institut d'Investigació Biomèdica de Bellvitge - IDIBELL, L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Program for Clinical Translation of Regenerative Medicine in Catalonia - P-[CMRC], L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
| | - Martín Ortiz-Genga
- Instituto de Investigación Biomédica de A Coruña (INIBIC), Universidade da Coruña, A Coruña, Spain (J.M.L.-M., M.O.-G., J.P.O., R.B.-V.)
| | - Juan Pablo Ochoa
- Instituto de Investigación Biomédica de A Coruña (INIBIC), Universidade da Coruña, A Coruña, Spain (J.M.L.-M., M.O.-G., J.P.O., R.B.-V.)
- Health in Code S.L., Scientific Department, A Coruña, Spain (J.P.O., L.M.)
| | - David Carreras
- Cardiovascular Genetics Center, Biomedical Research Institute of Girona, Spain (D.C., G.J.P., R.B.)
- Department of Medical Sciences, Universitat de Girona, Spain (D.C., G.J.P., R.B.)
| | - Guillermo Javier Pérez
- Cardiovascular Genetics Center, Biomedical Research Institute of Girona, Spain (D.C., G.J.P., R.B.)
- Department of Medical Sciences, Universitat de Girona, Spain (D.C., G.J.P., R.B.)
- Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares (CIBERCV), Spain (G.J.P., J.L.d.l.P., R.B., R.B.-V.)
| | - José Luis de la Pompa
- Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares (CIBERCV), Spain (G.J.P., J.L.d.l.P., R.B., R.B.-V.)
- Intercellular Signalling in Cardiovascular Development & Disease Laboratory, Centro Nacional de Investigaciones Cardiovasculares (CNIC), Madrid, Spain (J.L.d.l.P.)
| | - Ramón Brugada
- Cardiovascular Genetics Center, Biomedical Research Institute of Girona, Spain (D.C., G.J.P., R.B.)
- Department of Medical Sciences, Universitat de Girona, Spain (D.C., G.J.P., R.B.)
- Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares (CIBERCV), Spain (G.J.P., J.L.d.l.P., R.B., R.B.-V.)
- Hospital Josep Trueta, Girona, Spain (R.B.)
| | - Lorenzo Monserrat
- Health in Code S.L., Scientific Department, A Coruña, Spain (J.P.O., L.M.)
| | - Roberto Barriales-Villa
- Unidad de Cardiopatías Familiares, Servicio de Cardiología, Complexo Hospitalario Universitario de A Coruña, Servizo Galego de Saúde (SERGAS) (J.M.L.-M., R.B.-V.)
- Instituto de Investigación Biomédica de A Coruña (INIBIC), Universidade da Coruña, A Coruña, Spain (J.M.L.-M., M.O.-G., J.P.O., R.B.-V.)
- Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares (CIBERCV), Spain (G.J.P., J.L.d.l.P., R.B., R.B.-V.)
| | - Angel Raya
- Regenerative Medicine Program, Institut d'Investigació Biomèdica de Bellvitge - IDIBELL, L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Program for Clinical Translation of Regenerative Medicine in Catalonia - P-[CMRC], L'Hospitalet de Llobregat, Spain (R.E., Y.R.-P., L.P., I.L., S.J.-D., A.M.-G., A.R.)
- Center for Networked Biomedical Research on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Barcelona, Spain (R.E., Y.R.-P., L.P., A.R.)
- Institució Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain (A.R.)
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Lymperopoulos A. Clinical pharmacology of cardiac cyclic AMP in human heart failure: too much or too little? Expert Rev Clin Pharmacol 2023; 16:623-630. [PMID: 37403791 PMCID: PMC10529896 DOI: 10.1080/17512433.2023.2233891] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2023] [Accepted: 07/04/2023] [Indexed: 07/06/2023]
Abstract
INTRODUCTION Cyclic 3', 5'-adenosine monophosphate (cAMP) is a major signaling hub in cardiac physiology. Although cAMP signaling has been extensively studied in cardiac cells and animal models of heart failure (HF), not much is known about its actual amount present inside human failing or non-failing cardiomyocytes. Since many drugs used in HF work via cAMP, it is crucial to determine the status of its intracellular levels in failing vs. normal human hearts. AREAS COVERED Only studies performed on explanted/excised cardiac tissues from patients were examined. Studies that contained no data from human hearts or no data on cAMP levels per se were excluded from this perspective's analysis. EXPERT OPINION Currently, there is no consensus on the status of cAMP levels in human failing vs. non-failing hearts. Several studies on animal models may suggest maladaptive (e.g. pro-apoptotic) effects of cAMP on HF, advocating for cAMP lowering for therapy, but human studies almost universally indicate that myocardial cAMP levels are deficient in human failing hearts. It is the expert opinion of this perspective that intracellular cAMP levels are too low in human failing hearts, contributing to the disease. Strategies to increase (restore), not decrease, these levels should be pursued in human HF.
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Affiliation(s)
- Anastasios Lymperopoulos
- Laboratory for the Study of Neurohormonal Control of the Circulation, Department of Pharmaceutical Sciences, Nova Southeastern University Barry and Judy Silverman College of Pharmacy, Fort Lauderdale, FL, USA
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Turner KL, Morris HS, Awinda PO, Fitzsimons DP, Tanner BCW. RLC phosphorylation amplifies Ca2+ sensitivity of force in myocardium from cMyBP-C knockout mice. J Gen Physiol 2023; 155:213841. [PMID: 36715675 PMCID: PMC9930131 DOI: 10.1085/jgp.202213250] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2022] [Revised: 11/11/2022] [Accepted: 01/18/2023] [Indexed: 01/31/2023] Open
Abstract
Hypertrophic cardiomyopathy (HCM) is the leading genetic cause of heart disease. The heart comprises several proteins that work together to properly facilitate force production and pump blood throughout the body. Cardiac myosin binding protein-C (cMyBP-C) is a thick-filament protein, and mutations in cMyBP-C are frequently linked with clinical cases of HCM. Within the sarcomere, the N-terminus of cMyBP-C likely interacts with the myosin regulatory light chain (RLC); RLC is a subunit of myosin located within the myosin neck region that modulates contractile dynamics via its phosphorylation state. Phosphorylation of RLC is thought to influence myosin head position along the thick-filament backbone, making it more favorable to bind the thin filament of actin and facilitate force production. However, little is known about how these two proteins interact. We tested the effects of RLC phosphorylation on Ca2+-regulated contractility using biomechanical assays on skinned papillary muscle strips isolated from cMyBP-C KO mice and WT mice. RLC phosphorylation increased Ca2+ sensitivity of contraction (i.e., pCa50) from 5.80 ± 0.02 to 5.95 ± 0.03 in WT strips, whereas RLC phosphorylation increased Ca2+ sensitivity of contraction from 5.86 ± 0.02 to 6.15 ± 0.03 in cMyBP-C KO strips. These data suggest that the effects of RLC phosphorylation on Ca2+ sensitivity of contraction are amplified when cMyBP-C is absent from the sarcomere. This implies that cMyBP-C and RLC act in concert to regulate contractility in healthy hearts, and mutations to these proteins that lead to HCM (or a loss of phosphorylation with disease progression) may disrupt important interactions between these thick-filament regulatory proteins.
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Affiliation(s)
- Kyrah L Turner
- School of Molecular Biosciences & Neuroscience, Washington State University , Pullman, WA, USA
| | - Haley S Morris
- School of Molecular Biosciences & Neuroscience, Washington State University , Pullman, WA, USA
| | - Peter O Awinda
- Department of Integrative Physiology & Neuroscience, Washington State University , Pullman, WA, USA
| | - Daniel P Fitzsimons
- Department of Animal, Veterinary and Food Sciences, University of Idaho , Moscow, ID, USA
| | - Bertrand C W Tanner
- Department of Integrative Physiology & Neuroscience, Washington State University , Pullman, WA, USA
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10
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Borges JI, Suster MS, Lymperopoulos A. Cardiac RGS Proteins in Human Heart Failure and Atrial Fibrillation: Focus on RGS4. Int J Mol Sci 2023; 24:6136. [PMID: 37047106 PMCID: PMC10147095 DOI: 10.3390/ijms24076136] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2023] [Revised: 03/22/2023] [Accepted: 03/23/2023] [Indexed: 04/14/2023] Open
Abstract
The regulator of G protein signaling (RGS) proteins are crucial for the termination of G protein signals elicited by G protein-coupled receptors (GPCRs). This superfamily of cell membrane receptors, by far the largest and most versatile in mammals, including humans, play pivotal roles in the regulation of cardiac function and homeostasis. Perturbations in both the activation and termination of their G protein-mediated signaling underlie numerous heart pathologies, including heart failure (HF) and atrial fibrillation (AFib). Therefore, RGS proteins play important roles in the pathophysiology of these two devasting cardiac diseases, and several of them could be targeted therapeutically. Although close to 40 human RGS proteins have been identified, each RGS protein seems to interact only with a specific set of G protein subunits and GPCR types/subtypes in any given tissue or cell type. Numerous in vitro and in vivo studies in animal models, and also in diseased human heart tissue obtained from transplantations or tissue banks, have provided substantial evidence of the roles various cardiomyocyte RGS proteins play in cardiac normal homeostasis as well as pathophysiology. One RGS protein in particular, RGS4, has been reported in what are now decades-old studies to be selectively upregulated in human HF. It has also been implicated in protection against AFib via knockout mice studies. This review summarizes the current understanding of the functional roles of cardiac RGS proteins and their implications for the treatment of HF and AFib, with a specific focus on RGS4 for the aforementioned reasons but also because it can be targeted successfully with small organic molecule inhibitors.
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Affiliation(s)
| | | | - Anastasios Lymperopoulos
- Laboratory for the Study of Neurohormonal Control of the Circulation, Department of Pharmaceutical Sciences, Barry and Judy Silverrman College of Pharmacy, Nova Southeastern University, Fort Lauderdale, FL 33328-2018, USA
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11
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Lewis CTA, Ochala J. Myosin Heavy Chain as a Novel Key Modulator of Striated Muscle Resting State. Physiology (Bethesda) 2023; 38:0. [PMID: 36067133 DOI: 10.1152/physiol.00018.2022] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
After years of intense research using structural, biological, and biochemical experimental procedures, it is clear that myosin molecules are essential for striated muscle contraction. However, this is just the tip of the iceberg of their function. Interestingly, it has been shown recently that these molecules (especially myosin heavy chains) are also crucial for cardiac and skeletal muscle resting state. In the present review, we first overview myosin heavy chain biochemical states and how they influence the consumption of ATP. We then detail how neighboring partner proteins including myosin light chains and myosin binding protein C intervene in such processes, modulating the ATP demand in health and disease. Finally, we present current experimental drugs targeting myosin ATP consumption and how they can treat muscle diseases.
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Affiliation(s)
| | - Julien Ochala
- Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark
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12
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Del Calvo G, Baggio Lopez T, Lymperopoulos A. The therapeutic potential of targeting cardiac RGS4. Ther Adv Cardiovasc Dis 2023; 17:17539447231199350. [PMID: 37724539 PMCID: PMC10510358 DOI: 10.1177/17539447231199350] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/01/2023] [Accepted: 08/16/2023] [Indexed: 09/21/2023] Open
Abstract
G protein-coupled receptors (GPCRs) play pivotal roles in regulation of cardiac function and homeostasis. To function properly, every cell needs these receptors to be stimulated only when a specific extracellular stimulus is present, and to be silenced the moment that stimulus is removed. The regulator of G protein signaling (RGS) proteins are crucial for the latter to occur at the cell membrane, where the GPCR normally resides. Perturbations in both activation and termination of G protein signaling underlie numerous heart pathologies. Although more than 30 mammalian RGS proteins have been identified, each RGS protein seems to interact only with a specific set of G protein subunits and GPCR types/subtypes in any given tissue or cell type, and this applies to the myocardium as well. A large number of studies have provided substantial evidence for the roles various RGS proteins expressed in cardiomyocytes play in cardiac physiology and heart disease pathophysiology. This review summarizes the current understanding of the functional roles of cardiac RGS proteins and their implications for the treatment of specific heart diseases, such as heart failure and atrial fibrillation. We focus on cardiac RGS4 in particular, since this isoform appears to be selectively (among the RGS protein family) upregulated in human heart failure and is also the target of ongoing drug discovery efforts for the treatment of a variety of diseases.
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Affiliation(s)
- Giselle Del Calvo
- Laboratory for the Study of Neurohormonal Control of the Circulation, Department of Pharmaceutical Sciences, Barry and Judy Silverman College of Pharmacy, Nova Southeastern University, Fort Lauderdale, FL, USA
| | - Teresa Baggio Lopez
- Laboratory for the Study of Neurohormonal Control of the Circulation, Department of Pharmaceutical Sciences, Barry and Judy Silverman College of Pharmacy, Nova Southeastern University, Fort Lauderdale, FL, USA
| | - Anastasios Lymperopoulos
- Laboratory for the Study of Neurohormonal Control of the Circulation, Department of Pharmaceutical Sciences, Barry and Judy Silverman College of Pharmacy, Nova Southeastern University, 3200 South University Drive, HPD (Terry) Building/Room 1350, Fort Lauderdale, FL 33328-2018, USA
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13
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Da’as SI, Hasan W, Salem R, Younes N, Abdelrahman D, Mohamed IA, Aldaalis A, Temanni R, Mathew LS, Lorenz S, Yacoub M, Nomikos M, Nasrallah GK, Fakhro KA. Transcriptome Profile Identifies Actin as an Essential Regulator of Cardiac Myosin Binding Protein C3 Hypertrophic Cardiomyopathy in a Zebrafish Model. Int J Mol Sci 2022; 23:ijms23168840. [PMID: 36012114 PMCID: PMC9408294 DOI: 10.3390/ijms23168840] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2022] [Revised: 07/24/2022] [Accepted: 07/26/2022] [Indexed: 01/15/2023] Open
Abstract
Variants in cardiac myosin-binding protein C (cMyBP-C) are the leading cause of inherited hypertrophic cardiomyopathy (HCM), demonstrating the key role that cMyBP-C plays in the heart’s contractile machinery. To investigate the c-MYBPC3 HCM-related cardiac impairment, we generated a zebrafish mypbc3-knockout model. These knockout zebrafish displayed significant morphological heart alterations related to a significant decrease in ventricular and atrial diameters at systolic and diastolic states at the larval stages. Immunofluorescence staining revealed significant hyperplasia in the mutant’s total cardiac and ventricular cardiomyocytes. Although cardiac contractility was similar to the wild-type control, the ejection fraction was significantly increased in the mypbc3 mutants. At later stages of larval development, the mutants demonstrated an early cardiac phenotype of myocardium remodeling, concurrent cardiomyocyte hyperplasia, and increased ejection fraction as critical processes in HCM initiation to counteract the increased ventricular myocardial wall stress. The examination of zebrafish adults showed a thickened ventricular cardiac wall with reduced heart rate, swimming speed, and endurance ability in both the mypbc3 heterozygous and homozygous groups. Furthermore, heart transcriptome profiling showed a significant downregulation of the actin-filament-based process, indicating an impaired actin cytoskeleton organization as the main dysregulating factor associated with the early ventricular cardiac hypertrophy in the zebrafish mypbc3 HCM model.
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Affiliation(s)
- Sahar Isa Da’as
- Department of Human Genetics, Sidra Medicine, Doha P.O. Box 26999, Qatar
- Australian Regenerative Medicine Institute, College of Health and Life Sciences, Hamad Bin Khalifa University, Doha P.O. Box 34110, Qatar
- Correspondence:
| | - Waseem Hasan
- Department of Human Genetics, Sidra Medicine, Doha P.O. Box 26999, Qatar
| | - Rola Salem
- Health Center, Qatar University, Doha P.O. Box 2713, Qatar
| | - Nadine Younes
- Department of Biomedical Sciences, College of Health Science, Member of QU Health, Qatar University, Doha P.O. Box 2713, Qatar
- Biomedical Research Center, Qatar University, Doha P.O. Box 2713, Qatar
| | - Doua Abdelrahman
- Department of Human Genetics, Sidra Medicine, Doha P.O. Box 26999, Qatar
| | - Iman A. Mohamed
- Australian Regenerative Medicine Institute, Monash University, Melbourne 3168, Australia
| | - Arwa Aldaalis
- Australian Regenerative Medicine Institute, College of Health and Life Sciences, Hamad Bin Khalifa University, Doha P.O. Box 34110, Qatar
| | - Ramzi Temanni
- Integrated Genomics Services, Sidra Medicine, Doha P.O. Box 26999, Qatar
| | - Lisa Sara Mathew
- Integrated Genomics Services, Sidra Medicine, Doha P.O. Box 26999, Qatar
| | - Stephan Lorenz
- Integrated Genomics Services, Sidra Medicine, Doha P.O. Box 26999, Qatar
| | | | - Michail Nomikos
- College of Medicine, Member of QU Health, Qatar University, Doha P.O. Box 2713, Qatar
| | - Gheyath K. Nasrallah
- Department of Biomedical Sciences, College of Health Science, Member of QU Health, Qatar University, Doha P.O. Box 2713, Qatar
- Biomedical Research Center, Qatar University, Doha P.O. Box 2713, Qatar
| | - Khalid A. Fakhro
- Department of Human Genetics, Sidra Medicine, Doha P.O. Box 26999, Qatar
- Australian Regenerative Medicine Institute, College of Health and Life Sciences, Hamad Bin Khalifa University, Doha P.O. Box 34110, Qatar
- Weill Cornell Medical College, Doha P.O. Box 24811, Qatar
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14
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Stathopoulou K, Schnittger J, Raabe J, Fleischer F, Mangels N, Piasecki A, Findlay J, Hartmann K, Krasemann S, Schlossarek S, Uebeler J, Wixler V, Blake DJ, Baillie GS, Carrier L, Ehler E, Cuello F. CMYA5 is a novel interaction partner of FHL2 in cardiac myocytes. FEBS J 2022; 289:4622-4645. [PMID: 35176204 DOI: 10.1111/febs.16402] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2021] [Revised: 01/13/2022] [Accepted: 02/15/2022] [Indexed: 11/27/2022]
Abstract
Four-and-a-half LIM domains protein 2 (FHL2) is an anti-hypertrophic adaptor protein that regulates cardiac myocyte signalling and function. Herein, we identified cardiomyopathy-associated 5 (CMYA5) as a novel FHL2 interaction partner in cardiac myocytes. In vitro pull-down assays demonstrated interaction between FHL2 and the N- and C-terminal regions of CMYA5. The interaction was verified in adult cardiac myocytes by proximity ligation assays. Immunofluorescence and confocal microscopy demonstrated co-localisation in the same subcellular compartment. The binding interface between FHL2 and CMYA5 was mapped by peptide arrays. Exposure of neonatal rat ventricular myocytes to a CMYA5 peptide covering one of the FHL2 interaction sites led to an increase in cell area at baseline, but a blunted response to chronic phenylephrine treatment. In contrast to wild-type hearts, loss or reduced FHL2 expression in Fhl2-targeted knockout mouse hearts or in a humanised mouse model of hypertrophic cardiomyopathy led to redistribution of CMYA5 into the perinuclear and intercalated disc region. Taken together, our results indicate a direct interaction of the two adaptor proteins FHL2 and CMYA5 in cardiac myocytes, which might impact subcellular compartmentation of CMYA5.
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Affiliation(s)
- Konstantina Stathopoulou
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Germany
| | - Josef Schnittger
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Germany
| | - Janice Raabe
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Germany
| | - Frederic Fleischer
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Germany
| | - Nils Mangels
- Department of Biochemistry and Molecular Cell Biology, University Medical Center Hamburg-Eppendorf, Germany
| | - Angelika Piasecki
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Germany
| | - Jane Findlay
- Institute of Cardiovascular and Medical Sciences, University of Glasgow, UK
| | - Kristin Hartmann
- Institute of Neuropathology, University Medical Center Hamburg-Eppendorf, Germany
| | - Susanne Krasemann
- Institute of Neuropathology, University Medical Center Hamburg-Eppendorf, Germany
| | - Saskia Schlossarek
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Germany
| | - June Uebeler
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Germany
| | - Viktor Wixler
- Institute of Molecular Virology, Centre for Molecular Biology of Inflammation, Westfaelische Wilhelms-University, Germany
| | - Derek J Blake
- Division of Psychological Medicine and Clinical Neurosciences, School of Medicine, Cardiff University, UK
| | - George S Baillie
- Institute of Cardiovascular and Medical Sciences, University of Glasgow, UK
| | - Lucie Carrier
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Germany
| | - Elisabeth Ehler
- School of Cardiovascular Medicine and Sciences, BHF Research Excellence Centre, King's College London, UK.,Randall Centre for Cell and Molecular Biophysics (School of Basic and Medical Biosciences), King's College London, UK
| | - Friederike Cuello
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Germany
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15
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Suay-Corredera C, Alegre-Cebollada J. The mechanics of the heart: zooming in on hypertrophic cardiomyopathy and cMyBP-C. FEBS Lett 2022; 596:703-746. [PMID: 35224729 DOI: 10.1002/1873-3468.14301] [Citation(s) in RCA: 16] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2021] [Revised: 01/10/2022] [Accepted: 01/14/2022] [Indexed: 11/10/2022]
Abstract
Hypertrophic cardiomyopathy (HCM), a disease characterized by cardiac muscle hypertrophy and hypercontractility, is the most frequently inherited disorder of the heart. HCM is mainly caused by variants in genes encoding proteins of the sarcomere, the basic contractile unit of cardiomyocytes. The most frequently mutated among them is MYBPC3, which encodes cardiac myosin-binding protein C (cMyBP-C), a key regulator of sarcomere contraction. In this review, we summarize clinical and genetic aspects of HCM and provide updated information on the function of the healthy and HCM sarcomere, as well as on emerging therapeutic options targeting sarcomere mechanical activity. Building on what is known about cMyBP-C activity, we examine different pathogenicity drivers by which MYBPC3 variants can cause disease, focussing on protein haploinsufficiency as a common pathomechanism also in nontruncating variants. Finally, we discuss recent evidence correlating altered cMyBP-C mechanical properties with HCM development.
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16
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Methawasin M, Farman GP, Granzier-Nakajima S, Strom J, Kiss B, Smith JE, Granzier H. Shortening the thick filament by partial deletion of titin's C-zone alters cardiac function by reducing the operating sarcomere length range. J Mol Cell Cardiol 2022; 165:103-114. [PMID: 35031281 PMCID: PMC8940690 DOI: 10.1016/j.yjmcc.2022.01.002] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/24/2021] [Revised: 01/06/2022] [Accepted: 01/07/2022] [Indexed: 10/19/2022]
Abstract
Titin's C-zone is an inextensible segment in titin, comprised of 11 super-repeats and located in the cMyBP-C-containing region of the thick filament. Previously we showed that deletion of titin's super-repeats C1 and C2 (TtnΔC1-2 model) results in shorter thick filaments and contractile dysfunction of the left ventricular (LV) chamber but that unexpectedly LV diastolic stiffness is normal. Here we studied the contraction-relaxation kinetics from the time-varying elastance of the LV and intact cardiomyocyte, cellular work loops of intact cardiomyocytes, Ca2+ transients, cross-bridge kinetics, and myofilament Ca2+ sensitivity. Intact cardiomyocytes of TtnΔC1-2 mice exhibit systolic dysfunction and impaired relaxation. The time-varying elastance at both LV and single-cell levels showed that activation kinetics are normal in TtnΔC1-2 mice, but that relaxation is slower. The slowed relaxation is, in part, attributable to an increased myofilament Ca2+ sensitivity and slower early Ca2+ reuptake. Cross-bridge dynamics showed that cross-bridge kinetics are normal but that the number of force-generating cross-bridges is reduced. In vivo sarcomere length (SL) measurements revealed that in TtnΔC1-2 mice the operating SL range of the LV is shifted towards shorter lengths. This normalizes the apparent cell and LV diastolic stiffness but further reduces systolic force as systole occurs further down on the ascending limb of the force-SL relation. We propose that the reduced working SLs reflect titin's role in regulating diastolic stiffness by altering the number of sarcomeres in series. Overall, our study reveals that thick filament length regulation by titin's C-zone is critical for normal cardiac function.
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Affiliation(s)
- Mei Methawasin
- Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85721, United States of America.
| | - Gerrie P Farman
- Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85721, United States of America
| | - Shawtaroh Granzier-Nakajima
- Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85721, United States of America
| | - Joshua Strom
- Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85721, United States of America
| | - Balazs Kiss
- Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85721, United States of America
| | - John E Smith
- Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85721, United States of America
| | - Henk Granzier
- Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85721, United States of America.
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17
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Cheng Z, Fang T, Huang J, Guo Y, Alam M, Qian H. Hypertrophic Cardiomyopathy: From Phenotype and Pathogenesis to Treatment. Front Cardiovasc Med 2021; 8:722340. [PMID: 34760939 PMCID: PMC8572854 DOI: 10.3389/fcvm.2021.722340] [Citation(s) in RCA: 26] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2021] [Accepted: 09/17/2021] [Indexed: 02/05/2023] Open
Abstract
Hypertrophic cardiomyopathy (HCM) is a very common inherited cardiovascular disease (CAD) and the incidence is about 1/500 of the common population. It is caused by more than 1,400 mutations in 11 or more genes encoding the proteins of the cardiac sarcomere. HCM presents a heterogeneous clinical profile and complex pathophysiology and HCM is the most important cause of sudden cardiac death (SCD) in young people. HCM also contributes to functional disability from heart failure and stroke (caused by atrial fibrillation). Current treatments for HCM (medication, myectomy, and alcohol septal ablation) are geared toward slowing down the disease progression and symptom relief and implanted cardiac defibrillator (ICD) to prevent SCD. HCM is, however, entering a period of tight translational research that holds promise for the major advances in disease-specific therapy. Main insights into the genetic landscape of HCM have improved our understanding of molecular pathogenesis and pointed the potential targets for the development of therapeutic agents. We reviewed the critical discoveries about the treatments, mechanism of HCM, and their implications for future research.
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Affiliation(s)
- Zeyi Cheng
- Department of Cardiovascular Surgery, West China Hospital, Sichuan University, Chengdu, China
| | - Tingting Fang
- Department of Cardiology, West China Hospital, Sichuan University, Chengdu, China
| | - Jinglei Huang
- School of Medicine, Lanzhou University, Lanzhou, China
| | - Yingqiang Guo
- Department of Cardiovascular Surgery, West China Hospital, Sichuan University, Chengdu, China
| | - Mahboob Alam
- Division of Cardiovascular Medicine, Department of Medicine, Baylor College of Medicine, Houston, TX, United States
| | - Hong Qian
- Department of Cardiovascular Surgery, West China Hospital, Sichuan University, Chengdu, China
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18
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Düsener S, Flenner F, Maack C, Kohlhaas M, Bay J, Carrier L, Friedrich FW. Ouabain worsens diastolic sarcomere length in myocytes from a cardiomyopathy mouse model. Eur J Pharmacol 2021; 904:174170. [PMID: 33984298 DOI: 10.1016/j.ejphar.2021.174170] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2021] [Revised: 04/28/2021] [Accepted: 05/06/2021] [Indexed: 12/01/2022]
Abstract
Diastolic dysfunction is a major feature of hypertrophic cardiomyopathy (HCM). Data from patient tissue and animal models associate increased Ca2+ sensitivity of myofilaments with altered Na+ and Ca2+ ion homeostasis in cardiomyocytes with diastolic dysfunction. In this study, we tested the acute effects of ouabain on ventricular myocytes of an HCM mouse model. The effects of ouabain on contractility and Ca2+ transients were tested in intact adult mouse ventricular myocytes (AMVMs) of Mybpc3-targeted knock-in (KI) and wild-type (WT) mice. Concentration-response assessment of contractile function revealed low sensitivity of AMVMs to ouabain (10 μM) compared to literature data on human cardiomyocytes (100 nM). Three hundred μM ouabain increased contraction amplitude (WT ~1.8-fold; KI ~1.5-fold) and diastolic intracellular Ca2+ in both WT and KI (+12-18%), but further decreased diastolic sarcomere length in KI cardiomyocytes (-5%). Western Blot analysis of whole heart protein extracts revealed 50% lower amounts of Na+/K+ ATPase (NKA) in KI than in WT. Ouabain worsened the diastolic phenotype of KI cardiomyocytes at concentrations which did not impair WT diastolic function. Ouabain led to an elevation of intracellular Ca2+, which was poorly tolerated in KI showing already high cytosolic Ca2+ at baseline due to increased myofilament Ca2+ sensitivity. Lower amounts of NKA in KI could amplify the need to exchange excessive intracellular Na+ for Ca2+ and thereby explain the general tendency to higher diastolic Ca2+ in KI.
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Affiliation(s)
- Silke Düsener
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Frederik Flenner
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Christoph Maack
- Department of Translational Research, Comprehensive Heart Failure Center (CHFC), University Clinic Würzburg, Würzburg, Germany
| | - Michael Kohlhaas
- Department of Translational Research, Comprehensive Heart Failure Center (CHFC), University Clinic Würzburg, Würzburg, Germany
| | - Johannes Bay
- Department of Pediatrics and Neonatology, Saarland University Hospital, Homburg, Saar, Germany
| | - Lucie Carrier
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany.
| | - Felix W Friedrich
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany.
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19
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Bunch TA, Guhathakurta P, Lepak VC, Thompson AR, Kanassatega RS, Wilson A, Thomas DD, Colson BA. Cardiac myosin-binding protein C interaction with actin is inhibited by compounds identified in a high-throughput fluorescence lifetime screen. J Biol Chem 2021; 297:100840. [PMID: 34052227 PMCID: PMC8233204 DOI: 10.1016/j.jbc.2021.100840] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/04/2021] [Revised: 05/19/2021] [Accepted: 05/26/2021] [Indexed: 11/26/2022] Open
Abstract
Cardiac myosin-binding protein C (cMyBP-C) interacts with actin and myosin to modulate cardiac muscle contractility. These interactions are disfavored by cMyBP-C phosphorylation. Heart failure patients often display decreased cMyBP-C phosphorylation, and phosphorylation in model systems has been shown to be cardioprotective against heart failure. Therefore, cMyBP-C is a potential target for heart failure drugs that mimic phosphorylation or perturb its interactions with actin/myosin. Here we have used a novel fluorescence lifetime-based assay to identify small-molecule inhibitors of actin-cMyBP-C binding. Actin was labeled with a fluorescent dye (Alexa Fluor 568, AF568) near its cMyBP-C binding sites; when combined with the cMyBP-C N-terminal fragment, C0-C2, the fluorescence lifetime of AF568-actin decreases. Using this reduction in lifetime as a readout of actin binding, a high-throughput screen of a 1280-compound library identified three reproducible hit compounds (suramin, NF023, and aurintricarboxylic acid) that reduced C0-C2 binding to actin in the micromolar range. Binding of phosphorylated C0-C2 was also blocked by these compounds. That they specifically block binding was confirmed by an actin-C0-C2 time-resolved FRET (TR-FRET) binding assay. Isothermal titration calorimetry (ITC) and transient phosphorescence anisotropy (TPA) confirmed that these compounds bind to cMyBP-C, but not to actin. TPA results were also consistent with these compounds inhibiting C0-C2 binding to actin. We conclude that the actin-cMyBP-C fluorescence lifetime assay permits detection of pharmacologically active compounds that affect cMyBP-C-actin binding. We now have, for the first time, a validated high-throughput screen focused on cMyBP-C, a regulator of cardiac muscle contractility and known key factor in heart failure.
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Affiliation(s)
- Thomas A Bunch
- Department of Cellular & Molecular Medicine, University of Arizona, Tucson Arizona, USA
| | - Piyali Guhathakurta
- Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, Minnesota, USA
| | - Victoria C Lepak
- Department of Cellular & Molecular Medicine, University of Arizona, Tucson Arizona, USA
| | - Andrew R Thompson
- Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, Minnesota, USA
| | | | - Anna Wilson
- Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, Minnesota, USA
| | - David D Thomas
- Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, Minnesota, USA
| | - Brett A Colson
- Department of Cellular & Molecular Medicine, University of Arizona, Tucson Arizona, USA.
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20
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Flenner F, Jungen C, Küpker N, Ibel A, Kruse M, Koivumäki JT, Rinas A, Zech ATL, Rhoden A, Wijnker PJM, Lemoine MD, Steenpass A, Girdauskas E, Eschenhagen T, Meyer C, van der Velden J, Patten-Hamel M, Christ T, Carrier L. Translational investigation of electrophysiology in hypertrophic cardiomyopathy. J Mol Cell Cardiol 2021; 157:77-89. [PMID: 33957110 PMCID: PMC8320769 DOI: 10.1016/j.yjmcc.2021.04.009] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/10/2021] [Revised: 04/14/2021] [Accepted: 04/29/2021] [Indexed: 12/25/2022]
Abstract
Hypertrophic cardiomyopathy (HCM) patients are at increased risk of ventricular arrhythmias and sudden cardiac death, which can occur even in the absence of structural changes of the heart. HCM mouse models suggest mutations in myofilament components to affect Ca2+ homeostasis and thereby favor arrhythmia development. Additionally, some of them show indications of pro-arrhythmic changes in cardiac electrophysiology. In this study, we explored arrhythmia mechanisms in mice carrying a HCM mutation in Mybpc3 (Mybpc3-KI) and tested the translatability of our findings in human engineered heart tissues (EHTs) derived from CRISPR/Cas9-generated homozygous MYBPC3 mutant (MYBPC3hom) in induced pluripotent stem cells (iPSC) and to left ventricular septum samples obtained from HCM patients. We observed higher arrhythmia susceptibility in contractility measurements of field-stimulated intact cardiomyocytes and ventricular muscle strips as well as in electromyogram recordings of Langendorff-perfused hearts from adult Mybpc3-KI mice than in wild-type (WT) controls. The latter only occurred in homozygous (Hom-KI) but not in heterozygous (Het-KI) mouse hearts. Both Het- and Hom-KI are known to display pro-arrhythmic increased Ca2+ myofilament sensitivity as a direct consequence of the mutation. In the electrophysiological characterization of the model, we observed smaller repolarizing K+ currents in single cell patch clamp, longer ventricular action potentials in sharp microelectrode recordings and longer ventricular refractory periods in Langendorff-perfused hearts in Hom-KI, but not Het-KI. Interestingly, reduced K+ channel subunit transcript levels and prolonged action potentials were already detectable in newborn, pre-hypertrophic Hom-KI mice. Human iPSC-derived MYBPC3hom EHTs, which genetically mimicked the Hom-KI mice, did exhibit lower mutant mRNA and protein levels, lower force, beating frequency and relaxation time, but no significant alteration of the force-Ca2+ relation in skinned EHTs. Furthermore, MYBPC3hom EHTs did show higher spontaneous arrhythmic behavior, whereas action potentials measured by sharp microelectrode did not differ to isogenic controls. Action potentials measured in septal myectomy samples did not differ between patients with HCM and patients with aortic stenosis, except for the only sample with a MYBPC3 mutation. The data demonstrate that increased myofilament Ca2+ sensitivity is not sufficient to induce arrhythmias in the Mybpc3-KI mouse model and suggest that reduced K+ currents can be a pro-arrhythmic trigger in Hom-KI mice, probably already in early disease stages. However, neither data from EHTs nor from left ventricular samples indicate relevant reduction of K+ currents in human HCM. Therefore, our study highlights the species difference between mouse and human and emphasizes the importance of research in human samples and human-like models.
Sudden cardiac death is threatening hypertrophic cardiomyopathy (HCM) patients. Arrhythmia mechanisms are not well understood. Mouse HCM models showed relevant reduction in K+ currents. Human iPSC-EHT model and HCM patient septal myectomies did not display this mechanism.
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Affiliation(s)
- Frederik Flenner
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Christiane Jungen
- DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany; Department of Cardiology-Electrophysiology, cardiac Neuro- and Electrophysiology Research Group (cNEP), University Heart and Vascular Center, University Hospital Hamburg-Eppendorf, Hamburg, Germany; Department of Cardiology, Willem Einthoven Center for Cardiac Arrhythmia Research and Management, Leiden University Medical Center, Leiden, the Netherlands
| | - Nadine Küpker
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Antonia Ibel
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Martin Kruse
- Department of Biology and Program in Neuroscience, Bates College, Lewiston, ME, USA
| | - Jussi T Koivumäki
- BioMediTech, Faculty of Medicine and Health Technology, Tampere University, Tampere, Finland
| | - Anna Rinas
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Antonia T L Zech
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Alexandra Rhoden
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Paul J M Wijnker
- Amsterdam UMC, Vrije Universiteit Amsterdam, Department of Physiology, Amsterdam Cardiovascular Sciences, Amsterdam, the Netherlands
| | - Marc D Lemoine
- DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany; Department of Cardiology-Electrophysiology, cardiac Neuro- and Electrophysiology Research Group (cNEP), University Heart and Vascular Center, University Hospital Hamburg-Eppendorf, Hamburg, Germany
| | - Anna Steenpass
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Evaldas Girdauskas
- Department of Cardiovascular Surgery, University Heart and Vascular Center, University Hospital Hamburg-Eppendorf, Hamburg, Germany
| | - Thomas Eschenhagen
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Christian Meyer
- Department of Cardiology-Electrophysiology, cardiac Neuro- and Electrophysiology Research Group (cNEP), University Heart and Vascular Center, University Hospital Hamburg-Eppendorf, Hamburg, Germany; Division of Cardiology/Angiology/Intensiv Care, cardiac Neuro- and Electrophysiology Research Consortium (cNEP), EVK Düsseldorf, Teaching Hospital University of Düsseldorf, Düsseldorf, Germany; Institute of Neural and Sensory Physiology, cardiac Neuro- and Electrophysiology Research Consortium (cNEP), University of Düsseldorf, Düsseldorf, Germany
| | - Jolanda van der Velden
- Amsterdam UMC, Vrije Universiteit Amsterdam, Department of Physiology, Amsterdam Cardiovascular Sciences, Amsterdam, the Netherlands
| | - Monica Patten-Hamel
- Department of General and Interventional Cardiology, University Heart Center, University Hospital Hamburg-Eppendorf, Hamburg, Germany
| | - Torsten Christ
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Lucie Carrier
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany.
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21
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Feyen DAM, McKeithan WL, Bruyneel AAN, Spiering S, Hörmann L, Ulmer B, Zhang H, Briganti F, Schweizer M, Hegyi B, Liao Z, Pölönen RP, Ginsburg KS, Lam CK, Serrano R, Wahlquist C, Kreymerman A, Vu M, Amatya PL, Behrens CS, Ranjbarvaziri S, Maas RGC, Greenhaw M, Bernstein D, Wu JC, Bers DM, Eschenhagen T, Metallo CM, Mercola M. Metabolic Maturation Media Improve Physiological Function of Human iPSC-Derived Cardiomyocytes. Cell Rep 2021; 32:107925. [PMID: 32697997 PMCID: PMC7437654 DOI: 10.1016/j.celrep.2020.107925] [Citation(s) in RCA: 230] [Impact Index Per Article: 57.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2020] [Revised: 05/15/2020] [Accepted: 06/26/2020] [Indexed: 12/15/2022] Open
Abstract
Induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) have enormous potential for the study of human cardiac disorders. However, their physiological immaturity severely limits their utility as a model system and their adoption for drug discovery. Here, we describe maturation media designed to provide oxidative substrates adapted to the metabolic needs of human iPSC (hiPSC)-CMs. Compared with conventionally cultured hiPSC-CMs, metabolically matured hiPSC-CMs contract with greater force and show an increased reliance on cardiac sodium (Na+) channels and sarcoplasmic reticulum calcium (Ca2+) cycling. The media enhance the function, long-term survival, and sarcomere structures in engineered heart tissues. Use of the maturation media made it possible to reliably model two genetic cardiac diseases: long QT syndrome type 3 due to a mutation in the cardiac Na+ channel SCN5A and dilated cardiomyopathy due to a mutation in the RNA splicing factor RBM20. The maturation media should increase the fidelity of hiPSC-CMs as disease models. Physiological immaturity of iPSC-derived cardiomyocytes limits their fidelity as disease models. Feyen et al. developed a low glucose, high oxidative substrate media that increase maturation of ventricular-like hiPSC-CMs in 2D and 3D cultures relative to standard protocols. Improved characteristics include a low resting Vm, rapid depolarization, and increased Ca2+ dependence and force generation.
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Affiliation(s)
- Dries A M Feyen
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA; Department of Cardiology, University Medical Center Utrecht, Utrecht, the Netherlands
| | - Wesley L McKeithan
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA; Sanford-Burnham-Prebys Medical Discovery Institute, La Jolla, CA, USA; Department of Bioengineering, University of California, San Diego, San Diego, CA, USA
| | - Arne A N Bruyneel
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Sean Spiering
- Sanford-Burnham-Prebys Medical Discovery Institute, La Jolla, CA, USA; Department of Bioengineering, University of California, San Diego, San Diego, CA, USA
| | - Larissa Hörmann
- Institute of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Bärbel Ulmer
- Institute of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Hui Zhang
- Department of Bioengineering, University of California, San Diego, San Diego, CA, USA
| | - Francesca Briganti
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Michaela Schweizer
- Electron Microscopy Unit, Center for Molecular Neurobiology Hamburg, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Bence Hegyi
- Department of Pharmacology, University of California, Davis, Davis, CA, USA
| | - Zhandi Liao
- Department of Pharmacology, University of California, Davis, Davis, CA, USA
| | | | - Kenneth S Ginsburg
- Department of Pharmacology, University of California, Davis, Davis, CA, USA
| | - Chi Keung Lam
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Ricardo Serrano
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Christine Wahlquist
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA; Sanford-Burnham-Prebys Medical Discovery Institute, La Jolla, CA, USA; Department of Bioengineering, University of California, San Diego, San Diego, CA, USA
| | - Alexander Kreymerman
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Michelle Vu
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Prashila L Amatya
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Charlotta S Behrens
- Institute of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Sara Ranjbarvaziri
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Renee G C Maas
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA; Department of Cardiology, University Medical Center Utrecht, Utrecht, the Netherlands
| | - Matthew Greenhaw
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Daniel Bernstein
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Joseph C Wu
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA
| | - Donald M Bers
- Department of Pharmacology, University of California, Davis, Davis, CA, USA
| | - Thomas Eschenhagen
- Institute of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Christian M Metallo
- Department of Bioengineering, University of California, San Diego, San Diego, CA, USA
| | - Mark Mercola
- Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA; Sanford-Burnham-Prebys Medical Discovery Institute, La Jolla, CA, USA; Department of Bioengineering, University of California, San Diego, San Diego, CA, USA.
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22
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Regulation of Cardiac PKA Signaling by cAMP and Oxidants. Antioxidants (Basel) 2021; 10:antiox10050663. [PMID: 33923287 PMCID: PMC8146537 DOI: 10.3390/antiox10050663] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2021] [Revised: 04/16/2021] [Accepted: 04/20/2021] [Indexed: 12/31/2022] Open
Abstract
Pathologies, such as cancer, inflammatory and cardiac diseases are commonly associated with long-term increased production and release of reactive oxygen species referred to as oxidative stress. Thereby, protein oxidation conveys protein dysfunction and contributes to disease progression. Importantly, trials to scavenge oxidants by systemic antioxidant therapy failed. This observation supports the notion that oxidants are indispensable physiological signaling molecules that induce oxidative post-translational modifications in target proteins. In cardiac myocytes, the main driver of cardiac contractility is the activation of the β-adrenoceptor-signaling cascade leading to increased cellular cAMP production and activation of its main effector, the cAMP-dependent protein kinase (PKA). PKA-mediated phosphorylation of substrate proteins that are involved in excitation-contraction coupling are responsible for the observed positive inotropic and lusitropic effects. PKA-actions are counteracted by cellular protein phosphatases (PP) that dephosphorylate substrate proteins and thus allow the termination of PKA-signaling. Both, kinase and phosphatase are redox-sensitive and susceptible to oxidation on critical cysteine residues. Thereby, oxidation of the regulatory PKA and PP subunits is considered to regulate subcellular kinase and phosphatase localization, while intradisulfide formation of the catalytic subunits negatively impacts on catalytic activity with direct consequences on substrate (de)phosphorylation and cardiac contractile function. This review article attempts to incorporate the current perception of the functionally relevant regulation of cardiac contractility by classical cAMP-dependent signaling with the contribution of oxidant modification.
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23
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Wu Y, Wang H, Li Z, Cheng J, Fang R, Cao H, Cui Y. Subtypes identification on heart failure with preserved ejection fraction via network enhancement fusion using multi-omics data. Comput Struct Biotechnol J 2021; 19:1567-1578. [PMID: 33868594 PMCID: PMC8039555 DOI: 10.1016/j.csbj.2021.03.010] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2020] [Revised: 03/03/2021] [Accepted: 03/06/2021] [Indexed: 11/24/2022] Open
Abstract
Heart failure with preserved ejection fraction (HFpEF) is associated with multiple etiologic and pathophysiologic factors. HFpEF leads to significant cardiovascular morbidity and mortality. There are various reasons that fail to identify effective therapeutic interventions for HFpEF, primarily due to its clinical heterogeneity causing significant difficulties in determining physiologic and prognostic implications for this syndrome. Thus, identifying clinical subtypes using multi-omics data has great implications for efficient treatment and prognosis of HFpEF patients. Here we proposed to integrate mRNA, DNA methylation and microRNA (miRNA) expression data of HFpEF with a similarity network fusion (SNF) method following a network enhancement (ne-SNF) denoising technique to form a fused network. A spectral clustering method was then used to obtain clusters of patient subtypes. Experiments on HFpEF datasets demonstrated that ne-SNF significantly outperforms single data subtype analysis and other integrated methods. The identified subgroups were shown to have statistically significant differences in survival. Two HFpEF subtypes were defined: a high-risk group (16.8%) and a low-risk group (83.2%). The 5-year mortality rates were 63.3% and 33.0% for the high- and low-risk group, respectively. After adjusting for the effects of clinical covariates, HFpEF patients in the high-risk group were 2.43 times more likely to die than the low-risk group. A total of 157 differentially expressed (DE) mRNAs, 2199 abnormal methylations and 121 DE miRNAs were identified between two subtypes. They were also enriched in many HFpEF-related biological processes or pathways. The ne-SNF method provides a novel pipeline for subtype identification in integrated analysis of multi-omics data.
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Affiliation(s)
- Yongqing Wu
- Division of Health Statistics, School of Public Health, Shanxi Medical University, Taiyuan, Shanxi 030001, PR China
| | - Huihui Wang
- Division of Health Statistics, School of Public Health, Shanxi Medical University, Taiyuan, Shanxi 030001, PR China
| | - Zhi Li
- Department of Hematology, Taiyuan Central Hospital of Shanxi Medical University, Taiyuan, Shanxi 030001, PR China
| | - Jinfang Cheng
- Department of Cardiology, Bethune Hospital, Shanxi Medical University, Taiyuan, Shanxi 030001, PR China
| | - Ruiling Fang
- Division of Health Statistics, School of Public Health, Shanxi Medical University, Taiyuan, Shanxi 030001, PR China
| | - Hongyan Cao
- Division of Health Statistics, School of Public Health, Shanxi Medical University, Taiyuan, Shanxi 030001, PR China.,Shanxi Provincial Key Laboratory of Major Disease Risk Assessment, Taiyuan, Shanxi 030001, PR China
| | - Yuehua Cui
- Department of Statistics and Probability, Michigan State University, East Lansing, MI 48824, USA
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24
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Schober KE, Rush JE, Luis Fuentes V, Glaus T, Summerfield NJ, Wright K, Lehmkuhl L, Wess G, Sayer MP, Loureiro J, MacGregor J, Mohren N. Effects of pimobendan in cats with hypertrophic cardiomyopathy and recent congestive heart failure: Results of a prospective, double-blind, randomized, nonpivotal, exploratory field study. J Vet Intern Med 2021; 35:789-800. [PMID: 33543810 PMCID: PMC7995419 DOI: 10.1111/jvim.16054] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/19/2020] [Revised: 01/19/2021] [Accepted: 01/20/2021] [Indexed: 12/11/2022] Open
Abstract
Background The benefits of pimobendan in the treatment of congestive heart failure (CHF) in cats with hypertrophic cardiomyopathy (HCM) have not been evaluated prospectively. Hypothesis/Objectives To investigate the effects of pimobendan in cats with HCM and recent CHF and to identify possible endpoints for a pivotal study. We hypothesized that pimobendan would be well‐tolerated and associated with improved outcome. Animals Eighty‐three cats with HCM and recently controlled CHF: 30 with and 53 without left ventricular outflow tract obstruction. Methods Prospective randomized placebo‐controlled double‐blind multicenter nonpivotal field study. Cats received either pimobendan (0.30 mg/kg q12h, n = 43), placebo (n = 39), or no medication (n = 1) together with furosemide (<10 mg/kg/d) with or without clopidogrel. The primary endpoint was a successful outcome (ie, completing the 180‐day study period without a dose escalation of furosemide). Results The proportion of cats in the full analysis set population with a successful outcome was not different between treatment groups (P = .75). For nonobstructive cats, the success rate was 32% in pimobendan‐treated cats versus 18.2% in the placebo group (odds ratio [OR], 2.12; 95% confidence interval [CI], 0.54‐8.34). For obstructive cats, the success rate was 28.6% and 60% in the pimobendan and placebo groups, respectively (OR, 0.27; 95% CI, 0.06‐1.26). No difference was found between treatments for the secondary endpoints of time to furosemide dose escalation or death (P = .89). Results were similar in the per‐protocol sets. Adverse events in both treatment groups were similar. Conclusions and Clinical Importance In this study of cats with HCM and recent CHF, no benefit of pimobendan on 180‐day outcome was identified.
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Affiliation(s)
- Karsten E Schober
- The Ohio State University, Department of Veterinary Clinical Sciences, Columbus, Ohio, USA
| | - John E Rush
- Cummings School of Veterinary Medicine, Tufts University, North Grafton, Massachusetts, USA
| | | | - Tony Glaus
- University of Zürich, Clinic for Small Animal Internal Medicine, Zürich, Switzerland
| | | | | | | | - Gerhard Wess
- University of Munich, Clinic of Small Animal Medicine, Munich, Germany
| | - Margaret P Sayer
- Charlotte Animal Referral & Emergency, Charlotte, North Carolina, USA
| | | | - John MacGregor
- New England Veterinary Cardiology, Durham, North Carolina, USA
| | - Nicole Mohren
- Boehringer Ingelheim Vetmedica GmbH, Ingelheim, Germany
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25
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Effects of High-Fat Diet Induced Obesity and Fructooligosaccharide Supplementation on Cardiac Protein Expression. Nutrients 2020; 12:nu12113404. [PMID: 33167590 PMCID: PMC7694524 DOI: 10.3390/nu12113404] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2020] [Revised: 10/29/2020] [Accepted: 11/03/2020] [Indexed: 01/12/2023] Open
Abstract
The mechanism by which high fat-diet induced obesity affects cardiac protein expression is unclear, and the extent to which this is modulated by prebiotic treatment is not known. These outcomes were assessed in rats initially fed a high-fat diet, then the top 40% weight gain group were randomly allocated to control (CON), high-fat (HF) and HF supplemented with fructooligosaccharide (32 g; HF-FOS) treatments for 12 weeks (n = 10/group). At sacrifice, left ventricles were either frozen or preserved in formalin. Serum was stored for glucose and insulin measurements. Protein spectra was obtained using an Orbitrap analyzer, processed with Sequest and fold changes assessed with Scaffold Q +. Treatment effects for body weights, glucose and insulin were assessed using one-way ANOVA, and the differential protein expression was assessed by a Mann-Whitney U test. The Database for Annotation, Visualization and Integrated Discovery and the Kyoto Encyclopedia of Genes and Genomes identified pathways containing overrepresented proteins. Hematoxylin and eosin sections were graded for hypertrophy and also quantified; differences were identified using Chi-square analyses and Mann-Whitney U tests. HF diet fed rats were significantly (p < 0.05) heavier than CON, and 23 proteins involved in mitochondrial function and lipid metabolism were differentially expressed between HF and CON. Between HF-FOS and HF, 117 proteins involved in contractility, lipid and carbohydrate metabolism were differentially expressed. HF cardiomyocytes were significantly (p < 0.05) more hypertrophic than CON. We conclude that high-fat feeding and FOS are associated with subcellular deviations in cardiac metabolism and contractility, which may influence myocardial function and alter the risk of cardiovascular disease.
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26
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Wacker C, Dams N, Schauer A, Ritzer A, Volk T, Wagner M. Region-specific mechanisms of corticosteroid-mediated inotropy in rat cardiomyocytes. Sci Rep 2020; 10:11604. [PMID: 32665640 PMCID: PMC7360564 DOI: 10.1038/s41598-020-68308-4] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/21/2020] [Accepted: 06/23/2020] [Indexed: 11/09/2022] Open
Abstract
Regional differences in ion channel activity in the heart control the sequence of repolarization and may contribute to differences in contraction. Corticosteroids such as aldosterone or corticosterone increase the L-type Ca2+ current (ICaL) in the heart via the mineralocorticoid receptor (MR). Here, we investigate the differential impact of corticosteroid-mediated increase in ICaL on action potentials (AP), ion currents, intracellular Ca2+ handling and contractility in endo- and epicardial myocytes of the rat left ventricle. Dexamethasone led to a similar increase in ICaL in endocardial and epicardial myocytes, while the K+ currents Ito and IK were unaffected. However, AP duration (APD) and AP-induced Ca2+ influx (QCa) significantly increased exclusively in epicardial myocytes, thus abrogating the normal differences between the groups. Dexamethasone increased Ca2+ transients, contractility and SERCA activity in both regions, the latter possibly due to a decrease in total phospholamban (PLB) and an increase PLBpThr17. These results suggest that corticosteroids are powerful modulators of ICaL, Ca2+ transients and contractility in both endo- and epicardial myocytes, while APD and QCa are increased in epicardial myocytes only. This indicates that increased ICaL and SERCA activity rather than QCa are the primary drivers of contractility by adrenocorticoids.
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Affiliation(s)
- Caroline Wacker
- Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Waldstraße 6, 91054, Erlangen, Germany
| | - Niklas Dams
- Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Waldstraße 6, 91054, Erlangen, Germany
| | - Alexander Schauer
- Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Waldstraße 6, 91054, Erlangen, Germany
| | - Anne Ritzer
- Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Waldstraße 6, 91054, Erlangen, Germany
| | - Tilmann Volk
- Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Waldstraße 6, 91054, Erlangen, Germany. .,Muscle Research Center Erlangen (MURCE), Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
| | - Michael Wagner
- Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Waldstraße 6, 91054, Erlangen, Germany. .,Abteilung für Rhythmologie, Herzzentrum Dresden, Fetscherstraße 76, 01307, Dresden, Germany.
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27
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Seeger T, Shrestha R, Lam CK, Chen C, McKeithan WL, Lau E, Wnorowski A, McMullen G, Greenhaw M, Lee J, Oikonomopoulos A, Lee S, Yang H, Mercola M, Wheeler M, Ashley EA, Yang F, Karakikes I, Wu JC. A Premature Termination Codon Mutation in MYBPC3 Causes Hypertrophic Cardiomyopathy via Chronic Activation of Nonsense-Mediated Decay. Circulation 2019; 139:799-811. [PMID: 30586709 DOI: 10.1161/circulationaha.118.034624] [Citation(s) in RCA: 96] [Impact Index Per Article: 16.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
BACKGROUND Hypertrophic cardiomyopathy (HCM) is frequently caused by mutations in myosin-binding protein C3 ( MYBPC3) resulting in a premature termination codon (PTC). The underlying mechanisms of how PTC mutations in MYBPC3 lead to the onset and progression of HCM are poorly understood. This study's aim was to investigate the molecular mechanisms underlying the pathogenesis of HCM associated with MYBPC3 PTC mutations by utilizing human isogenic induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). METHODS Isogenic iPSC lines were generated from HCM patients harboring MYBPC3 PTC mutations (p.R943x; p.R1073P_Fsx4) using genome editing. Comprehensive phenotypic and transcriptome analyses were performed in the iPSC-CMs. RESULTS We observed aberrant calcium handling properties with prolonged decay kinetics and elevated diastolic calcium levels in the absence of structural abnormalities or contracile dysfunction in HCM iPSC-CMs as compared to isogenic controls. The mRNA expression levels of MYBPC3 were significantly reduced in mutant iPSC-CMs, but the protein levels were comparable among isogenic iPSC-CMs, suggesting that haploinsufficiency of MYBPC3 does not contribute to the pathogenesis of HCM in vitro. Furthermore, truncated MYBPC3 peptides were not detected. At the molecular level, the nonsense-mediated decay pathway was activated, and a set of genes involved in major cardiac signaling pathways was dysregulated in HCM iPSC-CMs, indicating an HCM gene signature in vitro. Specific inhibition of the nonsense-mediated decay pathway in mutant iPSC-CMs resulted in reversal of the molecular phenotype and normalization of calcium-handling abnormalities. CONCLUSIONS iPSC-CMs carrying MYBPC3 PTC mutations displayed aberrant calcium signaling and molecular dysregulations in the absence of significant haploinsufficiency of MYBPC3 protein. Here we provided the first evidence of the direct connection between the chronically activated nonsense-mediated decay pathway and HCM disease development.
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Affiliation(s)
- Timon Seeger
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Rajani Shrestha
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Chi Keung Lam
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Caressa Chen
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Wesley L McKeithan
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Edward Lau
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Alexa Wnorowski
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Bioengineering (A.W., S.L., F.Y.), Stanford University School of Medicine, CA
| | - George McMullen
- Department of Cardiothoracic Surgery (G.M., M.G., I.K.), Stanford University School of Medicine, CA
| | - Matthew Greenhaw
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Cardiothoracic Surgery (G.M., M.G., I.K.), Stanford University School of Medicine, CA
| | - Jaecheol Lee
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Angelos Oikonomopoulos
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Soah Lee
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA.,Department of Bioengineering (A.W., S.L., F.Y.), Stanford University School of Medicine, CA.,Department of Orthopedic Surgery (S.L.), Stanford University School of Medicine, CA
| | - Huaxiao Yang
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Mark Mercola
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Matthew Wheeler
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Euan A Ashley
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA
| | - Fan Yang
- Department of Bioengineering (A.W., S.L., F.Y.), Stanford University School of Medicine, CA
| | - Ioannis Karakikes
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA
| | - Joseph C Wu
- Stanford Cardiovascular Institute (T.S., R.S., C.K.L., C.C., W.L.M., E.L., A.W., M.G, J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., I.K., J.C.W.), Stanford University School of Medicine, CA.,Department of Medicine, Division of Cardiology (T.S., R.S., C.K.L., C.C., W.L.M., E.L., J.L., A.O., S.L., H.Y., M.M., M.W., E.A.A., J.C.W.), Stanford University School of Medicine, CA.,Institute for Stem Cell Biology and Regenerative Medicine (J.C.W.) Stanford University School of Medicine, CA
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28
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Phosphomimetic cardiac myosin-binding protein C partially rescues a cardiomyopathy phenotype in murine engineered heart tissue. Sci Rep 2019; 9:18152. [PMID: 31796859 PMCID: PMC6890639 DOI: 10.1038/s41598-019-54665-2] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/11/2019] [Accepted: 11/15/2019] [Indexed: 12/20/2022] Open
Abstract
Phosphorylation of cardiac myosin-binding protein C (cMyBP-C), encoded by MYBPC3, increases the availability of myosin heads for interaction with actin thus enhancing contraction. cMyBP-C phosphorylation level is lower in septal myectomies of patients with hypertrophic cardiomyopathy (HCM) than in non-failing hearts. Here we compared the effect of phosphomimetic (D282) and wild-type (S282) cMyBP-C gene transfer on the HCM phenotype of engineered heart tissues (EHTs) generated from a mouse model carrying a Mybpc3 mutation (KI). KI EHTs showed lower levels of mutant Mybpc3 mRNA and protein, and altered gene expression compared with wild-type (WT) EHTs. Furthermore, KI EHTs exhibited faster spontaneous contractions and higher maximal force and sensitivity to external [Ca2+] under pacing. Adeno-associated virus-mediated gene transfer of D282 and S282 similarly restored Mybpc3 mRNA and protein levels and suppressed mutant Mybpc3 transcripts. Moreover, both exogenous cMyBP-C proteins were properly incorporated in the sarcomere. KI EHTs hypercontractility was similarly prevented by both treatments, but S282 had a stronger effect than D282 to normalize the force-Ca2+-relationship and the expression of dysregulated genes. These findings in an in vitro model indicate that S282 is a better choice than D282 to restore the HCM EHT phenotype. To which extent the results apply to human HCM remains to be seen.
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29
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Jungen C, Scherschel K, Flenner F, Jee H, Rajendran P, De Jong KA, Nikolaev V, Meyer C, Ardell JL, Tompkins JD. Increased arrhythmia susceptibility in type 2 diabetic mice related to dysregulation of ventricular sympathetic innervation. Am J Physiol Heart Circ Physiol 2019; 317:H1328-H1341. [PMID: 31625779 DOI: 10.1152/ajpheart.00249.2019] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/01/2023]
Abstract
Patients with type 2 diabetes mellitus (T2DM) have a greater risk of developing life-threatening cardiac arrhythmias. Because the underlying mechanisms and potential influence of diabetic autonomic neuropathy are not well understood, we aimed to assess the relevance of a dysregulation in cardiac autonomic tone. Ventricular arrhythmia susceptibility was increased in Langendorff-perfused hearts isolated from mice with T2DM (db/db). Membrane properties and synaptic transmission were similar at cardiac postganglionic parasympathetic neurons from diabetic and control mice; however, a greater asynchronous neurotransmitter release was present at sympathetic postganglionic neurons from the stellate ganglia of db/db mice. Western blot analysis showed a reduction of tyrosine hydroxylase (TH) from the ventricles of db/db mice, which was confirmed with confocal imaging as a heterogeneous loss of TH-immunoreactivity from the left ventricular wall but not the apex. In vivo stimulation of cardiac parasympathetic (vagus) or cardiac sympathetic (stellate ganglion) nerves induced similar changes in heart rate in control and db/db mice, and the kinetics of pacing-induced Ca2+ transients (recorded from isolated cardiomyocytes) were similar in control and db/db cells. Antagonism of cardiac muscarinic receptors did not affect the frequency or severity of arrhythmias in db/db mice, but sympathetic blockade with propranolol completely inhibited arrhythmogenicity. Collectively, these findings suggest that the increased ventricular arrhythmia susceptibility of type 2 diabetic mouse hearts is due to dysregulation of the sympathetic ventricular control.NEW & NOTEWORTHY Patients with type 2 diabetes mellitus have greater risk of suffering from sudden cardiac death. We found that the increased ventricular arrhythmia susceptibility in type 2 diabetic mouse hearts is due to cardiac sympathetic dysfunction. Sympathetic dysregulation is indicated by an increased asynchronous release at stellate ganglia, a heterogeneous loss of tyrosine hydroxylase from the ventricular wall but not apex, and inhibition of ventricular arrhythmias in db/db mice after β-sympathetic blockade.
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Affiliation(s)
- Christiane Jungen
- Department of Cardiology-Electrophysiology, cNEP, cardiac Neuro- and Electrophysiology research group, University Heart Center, University Hospital Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Katharina Scherschel
- Department of Cardiology-Electrophysiology, cNEP, cardiac Neuro- and Electrophysiology research group, University Heart Center, University Hospital Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Frederik Flenner
- DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Hamburg, Germany.,Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Centre, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany
| | - Haesung Jee
- University of California, Los Angeles Cardiac Arrhythmia Center, Neurocardiology Research Program of Excellence, Department of Medicine-Cardiology, Los Angeles, California
| | - Pradeep Rajendran
- University of California, Los Angeles Cardiac Arrhythmia Center, Neurocardiology Research Program of Excellence, Department of Medicine-Cardiology, Los Angeles, California
| | - Kirstie A De Jong
- DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Hamburg, Germany.,Institute of Experimental Cardiovascular Research, University Medical Center Hamburg-Eppendorf, University of Hamburg, Germany
| | - Viacheslav Nikolaev
- DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Hamburg, Germany.,Institute of Experimental Cardiovascular Research, University Medical Center Hamburg-Eppendorf, University of Hamburg, Germany
| | - Christian Meyer
- Department of Cardiology-Electrophysiology, cNEP, cardiac Neuro- and Electrophysiology research group, University Heart Center, University Hospital Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Center for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Jeffrey L Ardell
- University of California, Los Angeles Cardiac Arrhythmia Center, Neurocardiology Research Program of Excellence, Department of Medicine-Cardiology, Los Angeles, California
| | - John D Tompkins
- University of California, Los Angeles Cardiac Arrhythmia Center, Neurocardiology Research Program of Excellence, Department of Medicine-Cardiology, Los Angeles, California
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30
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Stathopoulou K, Schobesberger S, Bork NI, Sprenger JU, Perera RK, Sotoud H, Geertz B, David JP, Christ T, Nikolaev VO, Cuello F. Divergent off-target effects of RSK N-terminal and C-terminal kinase inhibitors in cardiac myocytes. Cell Signal 2019; 63:109362. [PMID: 31344438 DOI: 10.1016/j.cellsig.2019.109362] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/03/2019] [Revised: 07/18/2019] [Accepted: 07/18/2019] [Indexed: 12/15/2022]
Abstract
P90 ribosomal S6 kinases (RSK) are ubiquitously expressed and regulate responses to neurohumoral stimulation. To study the role of RSK signalling on cardiac myocyte function and protein phosphorylation, pharmacological RSK inhibitors were tested. Here, the ATP competitive N-terminal kinase domain-targeting compounds D1870 and SL0101 and the allosteric C-terminal kinase domain-targeting FMK were evaluated regarding their ability to modulate cardiac myocyte protein phosphorylation. Exposure to D1870 and SL0101 significantly enhanced phospholamban (PLN) Ser16 and cardiac troponin I (cTnI) Ser22/23 phosphorylation in response to D1870 and SL0101 upon exposure to phenylephrine (PE) that activates RSK. In contrast, FMK pretreatment significantly reduced phosphorylation of both proteins in response to PE. D1870-mediated enhancement of PLN Ser16 phosphorylation was also observed after exposure to isoprenaline or noradrenaline (NA) stimuli that do not activate RSK. Inhibition of β-adrenoceptors by atenolol or cAMP-dependent protein kinase (PKA) by H89 prevented the D1870-mediated increase in PLN phosphorylation, suggesting that PKA is the kinase responsible for the observed phosphorylation. Assessment of changes in cAMP formation by FRET measurements revealed increased cAMP formation in vicinity to PLN after exposure to D1870 and SL0101. D1870 inhibited phosphodiesterase activity similarly as established PDE inhibitors rolipram or 3-isobutyl-1-methylxanthine. Assessment of catecholamine-mediated force development in rat ventricular muscle strips revealed significantly reduced EC50 for NA after D1870 pretreatment (DMSO/NA: 2.33 μmol/L vs. D1870/NA: 1.30 μmol/L). The data reveal enhanced cardiac protein phosphorylation by D1870 and SL0101 that was not detectable in response to FMK. This disparate effect might be attributed to off-target inhibition of PDEs with impact on muscle function as demonstrated for D1870.
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Affiliation(s)
- Konstantina Stathopoulou
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Sophie Schobesberger
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Nadja I Bork
- DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; Institute of Experimental Cardiovascular Research, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Julia U Sprenger
- DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; Institute of Experimental Cardiovascular Research, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Ruwan K Perera
- DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; Institute of Experimental Cardiovascular Research, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Hannieh Sotoud
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Birgit Geertz
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Jean-Pierre David
- Institute of Osteology and Biomechanics, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Torsten Christ
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Viacheslav O Nikolaev
- DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; Institute of Experimental Cardiovascular Research, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany
| | - Friederike Cuello
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany; DZHK (German Center for Cardiovascular Research), Partner site Hamburg/Kiel/Lübeck, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany.
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31
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Verkaik M, Oranje M, Abdurrachim D, Goebel M, Gam Z, Prompers JJ, Helmes M, Ter Wee PM, van der Velden J, Kuster DW, Vervloet MG, Eringa EC. High Fibroblast Growth Factor 23 concentrations in experimental renal failure impair calcium handling in cardiomyocytes. Physiol Rep 2019; 6:e13591. [PMID: 29611320 PMCID: PMC5880876 DOI: 10.14814/phy2.13591] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2017] [Accepted: 12/13/2017] [Indexed: 12/14/2022] Open
Abstract
The overwhelming majority of patients with chronic kidney disease (CKD) die prematurely before reaching end‐stage renal disease, mainly due to cardiovascular causes, of which heart failure is the predominant clinical presentation. We hypothesized that CKD‐induced increases of plasma FGF23 impair cardiac diastolic and systolic function. To test this, mice were subjected to 5/6 nephrectomy (5/6Nx) or were injected with FGF23 for seven consecutive days. Six weeks after surgery, plasma FGF23 was higher in 5/6Nx mice compared to sham mice (720 ± 31 vs. 256 ± 3 pg/mL, respectively, P = 0.034). In cardiomyocytes isolated from both 5/6Nx and FGF23 injected animals the rise of cytosolic calcium during systole was slowed (−13% and −19%, respectively) as was the decay of cytosolic calcium during diastole (−15% and −21%, respectively) compared to controls. Furthermore, both groups had similarly decreased peak cytosolic calcium content during systole. Despite lower cytosolic calcium contents in CKD or FGF23 pretreated animals, no changes were observed in contractile parameters of cardiomyocytes between the groups. Expression of calcium handling proteins and cardiac troponin I phosphorylation were similar between groups. Blood pressure, the heart weight:tibia length ratio, α‐MHC/β‐MHC ratio and ANF mRNA expression, and systolic and diastolic function as measured by MRI did not differ between groups. In conclusion, the rapid, CKD‐induced rise in plasma FGF23 and the similar decrease in cardiomyocyte calcium transients in modeled kidney disease and following 1‐week treatment with FGF23 indicate that FGF23 partly mediates cardiomyocyte dysfunction in CKD.
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Affiliation(s)
- Melissa Verkaik
- Department of Nephrology, VU University Medical Center, Amsterdam, The Netherlands.,Department of Physiology, Amsterdam Cardiovascular Sciences, VU University Medical Center, Amsterdam, The Netherlands
| | - Maarten Oranje
- Department of Physiology, Amsterdam Cardiovascular Sciences, VU University Medical Center, Amsterdam, The Netherlands
| | - Desiree Abdurrachim
- Biomedical NMR, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands
| | - Max Goebel
- Department of Physiology, Amsterdam Cardiovascular Sciences, VU University Medical Center, Amsterdam, The Netherlands
| | - Zeineb Gam
- Department of Physiology, Amsterdam Cardiovascular Sciences, VU University Medical Center, Amsterdam, The Netherlands
| | - Jeanine J Prompers
- Biomedical NMR, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands
| | - Michiel Helmes
- Department of Physiology, Amsterdam Cardiovascular Sciences, VU University Medical Center, Amsterdam, The Netherlands
| | - Pieter M Ter Wee
- Department of Nephrology, VU University Medical Center, Amsterdam, The Netherlands
| | - Jolanda van der Velden
- Department of Physiology, Amsterdam Cardiovascular Sciences, VU University Medical Center, Amsterdam, The Netherlands
| | - Diederik W Kuster
- Department of Physiology, Amsterdam Cardiovascular Sciences, VU University Medical Center, Amsterdam, The Netherlands
| | - Marc G Vervloet
- Department of Nephrology, VU University Medical Center, Amsterdam, The Netherlands
| | - Etto C Eringa
- Department of Physiology, Amsterdam Cardiovascular Sciences, VU University Medical Center, Amsterdam, The Netherlands
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32
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Kresin N, Stücker S, Krämer E, Flenner F, Mearini G, Münch J, Patten M, Redwood C, Carrier L, Friedrich FW. Analysis of Contractile Function of Permeabilized Human Hypertrophic Cardiomyopathy Multicellular Heart Tissue. Front Physiol 2019; 10:239. [PMID: 30984009 PMCID: PMC6447666 DOI: 10.3389/fphys.2019.00239] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/03/2018] [Accepted: 02/22/2019] [Indexed: 01/08/2023] Open
Affiliation(s)
- Nico Kresin
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Sabrina Stücker
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Elisabeth Krämer
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Frederik Flenner
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Giulia Mearini
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Julia Münch
- University Heart Center Hamburg, Hamburg, Germany
| | | | - Charles Redwood
- Radcliffe Department of Medicine, University of Oxford, John Radcliffe Hospital, Oxford, United Kingdom
| | - Lucie Carrier
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Felix W Friedrich
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
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Viola HM, Hool LC. Impaired calcium handling and mitochondrial metabolic dysfunction as early markers of hypertrophic cardiomyopathy. Arch Biochem Biophys 2019; 665:166-174. [PMID: 30885674 DOI: 10.1016/j.abb.2019.03.006] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/21/2019] [Revised: 03/08/2019] [Accepted: 03/11/2019] [Indexed: 02/07/2023]
Abstract
Hypertrophic cardiomyopathy (HCM) is a primary myocardial disorder, characterised by myocyte remodeling, disorganisation of sarcomeric proteins, impaired energy metabolism and altered cardiac contractility. Gene mutations encoding cardiac contractile proteins account for 60% of HCM aetiology. Current drug therapy including L-type calcium channel antagonists, are used to manage symptoms in patients with overt HCM, but no treatment exists that can reverse or prevent the cardiomyopathy. Design of effective drug therapy will require a clear understanding of the early pathophysiological mechanisms of the disease. Numerous studies have investigated specific aspects of HCM pathophysiology. This review brings these findings together, in order to develop a holistic understanding of the early pathophysiological mechanisms of the disease. We focus on gene mutations in cardiac myosin binding protein-C, β-cardiac myosin heavy chain, cardiac troponin I, and cardiac troponin T, that comprise the majority of all HCM sarcomeric gene mutations. We find that although some similarities exist, each mutation leads to mutation-specific alterations in calcium handling, myofilament calcium sensitivity and mitochondrial metabolic function. This may contribute to the observed clinical phenotypic variability in sarcomeric-related HCM. An understanding of early mutation-specific mechanisms of the disease may provide useful markers of disease progression, and inform therapeutic design.
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Affiliation(s)
- Helena M Viola
- School of Human Sciences (Physiology), The University of Western Australia, Crawley, WA, Australia
| | - Livia C Hool
- School of Human Sciences (Physiology), The University of Western Australia, Crawley, WA, Australia; Victor Chang Cardiac Research Institute, Sydney, NSW, Australia.
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Mamidi R, Li J, Doh CY, Holmes JB, Stelzer JE. Lost in translation: Interpreting cardiac muscle mechanics data in clinical practice. Arch Biochem Biophys 2019; 662:213-218. [PMID: 30576628 PMCID: PMC6345594 DOI: 10.1016/j.abb.2018.12.021] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2018] [Revised: 10/25/2018] [Accepted: 12/17/2018] [Indexed: 01/10/2023]
Abstract
Current inotropic therapies improve systolic function in heart failure patients but also elicit undesirable side effects such as arrhythmias and increased intracellular Ca2+ transients. In order to maintain myocyte Ca2+ homeostasis, the increased cytosolic Ca2+ needs to be actively transported back to sarcoplasmic reticulum leading to depleted ATP reserves. Thus, an emerging approach is to design sarcomere-based treatments to correct impaired contractility via a direct and allosteric modulation of myosin's intrinsic force-generating behavior -a concept that potentially avoids the "off-target" effects. To achieve this goal, various biophysical approaches are utilized to investigate the mechanistic impact of sarcomeric modulators but information derived from diverse approaches is not fully integrated into therapeutic applications. This is in part due to the lack of information that provides a coherent connecting link between biophysical data to in vivo function. Hence, our ability to clearly discern the drug-mediated impact on whole-heart function is diminished. Reducing this translational barrier can significantly accelerate clinical progress related to sarcomere-based therapies by optimizing drug-dosing and treatment duration protocols based on information obtained from biophysical studies. Therefore, we attempt to link biophysical mechanical measurements obtained in isolated cardiac muscle and in vivo contractile function.
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Affiliation(s)
- Ranganath Mamidi
- Department of Physiology and Biophysics, School of Medicine, Case Western Reserve University, Cleveland, OH, 44106, USA
| | - Jiayang Li
- Department of Physiology and Biophysics, School of Medicine, Case Western Reserve University, Cleveland, OH, 44106, USA
| | - Chang Yoon Doh
- Department of Physiology and Biophysics, School of Medicine, Case Western Reserve University, Cleveland, OH, 44106, USA
| | - Joshua B Holmes
- Department of Physiology and Biophysics, School of Medicine, Case Western Reserve University, Cleveland, OH, 44106, USA
| | - Julian E Stelzer
- Department of Physiology and Biophysics, School of Medicine, Case Western Reserve University, Cleveland, OH, 44106, USA.
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35
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Toepfer CN, Wakimoto H, Garfinkel AC, McDonough B, Liao D, Jiang J, Tai AC, Gorham JM, Lunde IG, Lun M, Lynch TL, McNamara JW, Sadayappan S, Redwood CS, Watkins HC, Seidman JG, Seidman CE. Hypertrophic cardiomyopathy mutations in MYBPC3 dysregulate myosin. Sci Transl Med 2019; 11:eaat1199. [PMID: 30674652 PMCID: PMC7184965 DOI: 10.1126/scitranslmed.aat1199] [Citation(s) in RCA: 138] [Impact Index Per Article: 23.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2018] [Revised: 06/05/2018] [Accepted: 11/30/2018] [Indexed: 12/16/2022]
Abstract
The mechanisms by which truncating mutations in MYBPC3 (encoding cardiac myosin-binding protein C; cMyBPC) or myosin missense mutations cause hypercontractility and poor relaxation in hypertrophic cardiomyopathy (HCM) are incompletely understood. Using genetic and biochemical approaches, we explored how depletion of cMyBPC altered sarcomere function. We demonstrated that stepwise loss of cMyBPC resulted in reciprocal augmentation of myosin contractility. Direct attenuation of myosin function, via a damaging missense variant (F764L) that causes dilated cardiomyopathy (DCM), normalized the increased contractility from cMyBPC depletion. Depletion of cMyBPC also altered dynamic myosin conformations during relaxation, enhancing the myosin state that enables ATP hydrolysis and thin filament interactions while reducing the super relaxed conformation associated with energy conservation. MYK-461, a pharmacologic inhibitor of myosin ATPase, rescued relaxation deficits and restored normal contractility in mouse and human cardiomyocytes with MYBPC3 mutations. These data define dosage-dependent effects of cMyBPC on myosin that occur across the cardiac cycle as the pathophysiologic mechanisms by which MYBPC3 truncations cause HCM. Therapeutic strategies to attenuate cMyBPC activity may rescue depressed cardiac contractility in patients with DCM, whereas inhibiting myosin by MYK-461 should benefit the substantial proportion of patients with HCM with MYBPC3 mutations.
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Affiliation(s)
- Christopher N Toepfer
- Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, OX3 9DU, UK
- Wellcome Centre for Human Genetics, University of Oxford, OX3 7BN, UK
| | - Hiroko Wakimoto
- Department of Genetics, Harvard Medical School, Boston, MA 02115, USA
- Department of Cardiology, Children's Hospital Boston, Boston, MA 02115, USA
| | | | | | - Dan Liao
- Department of Biochemistry and Cardiovascular Research Institute (CVRI), Yong Loo Lin School of Medicine, National University of Singapore, Singapore 119228, Singapore
| | - Jianming Jiang
- Department of Biochemistry and Cardiovascular Research Institute (CVRI), Yong Loo Lin School of Medicine, National University of Singapore, Singapore 119228, Singapore
| | - Angela C Tai
- Department of Genetics, Harvard Medical School, Boston, MA 02115, USA
| | - Joshua M Gorham
- Department of Genetics, Harvard Medical School, Boston, MA 02115, USA
| | - Ida G Lunde
- Department of Genetics, Harvard Medical School, Boston, MA 02115, USA
- Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0318 Oslo, Norway
| | - Mingyue Lun
- Division of Genetics, Department of Medicine, Brigham and Women's Hospital, Boston, MA 02115, USA
| | - Thomas L Lynch
- Department of Molecular Pharmacology and Therapeutics, Health Sciences Division, Loyola University Chicago, Maywood, IL 60153, USA
| | - James W McNamara
- Heart, Lung and Vascular Institute, University of Cincinnati, Cincinnati, OH 45219, USA
| | - Sakthivel Sadayappan
- Heart, Lung and Vascular Institute, University of Cincinnati, Cincinnati, OH 45219, USA
| | - Charles S Redwood
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, OX3 9DU, UK
| | - Hugh C Watkins
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, OX3 9DU, UK
- Wellcome Centre for Human Genetics, University of Oxford, OX3 7BN, UK
| | | | - Christine E Seidman
- Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
- Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA
- Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA 02115, USA
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36
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Chakouri N, Reboul C, Boulghobra D, Kleindienst A, Nottin S, Gayrard S, Roubille F, Matecki S, Lacampagne A, Cazorla O. Stress-induced protein S-glutathionylation and phosphorylation crosstalk in cardiac sarcomeric proteins - Impact on heart function. Int J Cardiol 2018; 258:207-216. [PMID: 29544934 DOI: 10.1016/j.ijcard.2017.12.004] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/08/2017] [Revised: 11/16/2017] [Accepted: 12/01/2017] [Indexed: 11/25/2022]
Abstract
BACKGROUND The interplay between oxidative stress and other signaling pathways in the contractile machinery regulation during cardiac stress and its consequences on cardiac function remains poorly understood. We evaluated the effect of the crosstalk between β-adrenergic and redox signaling on post-translational modifications of sarcomeric regulatory proteins, Myosin Binding Protein-C (MyBP-C) and Troponin I (TnI). METHODS AND RESULTS We mimicked in vitro high level of physiological cardiac stress by forcing rat hearts to produce high levels of oxidized glutathione. This led to MyBP-C S-glutathionylation associated with lower protein kinase A (PKA) dependent phosphorylations of MyBP-C and TnI, increased myofilament Ca2+ sensitivity, and decreased systolic and diastolic properties of the isolated perfused heart. Moderate physiological cardiac stress achieved in vivo with a single 35 min exercise (Low stress induced by exercise, LSE) increased TnI and cMyBP-C phosphorylations and improved cardiac function in vivo (echocardiography) and ex-vivo (isolated perfused heart). High stress induced by exercise (HSE) altered strongly oxidative stress markers and phosphorylations were unchanged despite increased PKA activity. HSE led to in vivo intrinsic cardiac dysfunction associated with myofilament Ca2+ sensitivity defects. To limit protein S-glutathionylation after HSE, we treated rats with N-acetylcysteine (NAC). NAC restored the ability of PKA to modulate myofilament Ca2+ sensitivity and prevented cardiac dysfunction observed in HSE animals. CONCLUSION Under cardiac stress, adrenergic and oxidative signaling pathways work in concert to alter myofilament properties and are key regulators of cardiac function.
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Affiliation(s)
- Nourdine Chakouri
- PHYMEDEXP, INSERM U1046, CNRS UMR9214, Université de Montpellier, CHRU Montpellier, Montpellier, France
| | - Cyril Reboul
- EA 4278, Laboratoire de Pharm-Ecologie Cardiovasculaire, Avignon University, Avignon, France
| | - Doria Boulghobra
- EA 4278, Laboratoire de Pharm-Ecologie Cardiovasculaire, Avignon University, Avignon, France
| | - Adrien Kleindienst
- EA 4278, Laboratoire de Pharm-Ecologie Cardiovasculaire, Avignon University, Avignon, France
| | - Stéphane Nottin
- EA 4278, Laboratoire de Pharm-Ecologie Cardiovasculaire, Avignon University, Avignon, France
| | - Sandrine Gayrard
- EA 4278, Laboratoire de Pharm-Ecologie Cardiovasculaire, Avignon University, Avignon, France
| | - François Roubille
- PHYMEDEXP, INSERM U1046, CNRS UMR9214, Université de Montpellier, CHRU Montpellier, Montpellier, France
| | - Stefan Matecki
- PHYMEDEXP, INSERM U1046, CNRS UMR9214, Université de Montpellier, CHRU Montpellier, Montpellier, France
| | - Alain Lacampagne
- PHYMEDEXP, INSERM U1046, CNRS UMR9214, Université de Montpellier, CHRU Montpellier, Montpellier, France
| | - Olivier Cazorla
- PHYMEDEXP, INSERM U1046, CNRS UMR9214, Université de Montpellier, CHRU Montpellier, Montpellier, France.
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Eschenhagen T, Carrier L. Cardiomyopathy phenotypes in human-induced pluripotent stem cell-derived cardiomyocytes-a systematic review. Pflugers Arch 2018; 471:755-768. [PMID: 30324321 PMCID: PMC6475632 DOI: 10.1007/s00424-018-2214-0] [Citation(s) in RCA: 44] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/30/2018] [Revised: 09/19/2018] [Accepted: 10/02/2018] [Indexed: 12/12/2022]
Abstract
Human-induced pluripotent stem cells (hiPSC) can be differentiated to cardiomyocytes at high efficiency and are increasingly used to study cardiac disease in a human context. This review evaluated 38 studies on hypertrophic (HCM) and dilated cardiomyopathy (DCM) of different genetic causes asking to which extent published data allow the definition of an in vitro HCM/DCM hiPSC-CM phenotype. The data are put in context with the prevailing hypotheses on HCM/DCM dysfunction and pathophysiology. Relatively consistent findings in HCM not reported in DCM were larger cell size (156 ± 85%, n = 15), more nuclear localization of nuclear factor of activated T cells (NFAT; 175 ± 65%, n = 3), and higher β-myosin heavy chain gene expression levels (500 ± 547%, n = 8) than respective controls. Conversely, DCM lines showed consistently less force development than controls (47 ± 23%, n = 9), while HCM forces scattered without clear trend. Both HCM and DCM lines often showed sarcomere disorganization, higher NPPA/NPPB expression levels, and arrhythmic beating behaviour. The data have to be taken with the caveat that reporting frequencies of the various parameters (e.g. cell size, NFAT expression) differ widely between HCM and DCM lines, in which data scatter is large and that only 9/38 studies used isogenic controls. Taken together, the current data provide interesting suggestions for disease-specific phenotypes in HCM/DCM hiPSC-CM but indicate that the field is still in its early days. Systematic, quantitative comparisons and robust, high content assays are warranted to advance the field.
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Affiliation(s)
- Thomas Eschenhagen
- Institute of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany. .,Partner Site Hamburg/Kiel/Lübeck, DZHK (German Centre for Cardiovascular Research), Hamburg, Germany.
| | - Lucie Carrier
- Institute of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany. .,Partner Site Hamburg/Kiel/Lübeck, DZHK (German Centre for Cardiovascular Research), Hamburg, Germany.
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38
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Stanczyk PJ, Seidel M, White J, Viero C, George CH, Zissimopoulos S, Lai FA. Association of cardiac myosin-binding protein-C with the ryanodine receptor channel - putative retrograde regulation? J Cell Sci 2018; 131:jcs.210443. [PMID: 29930088 PMCID: PMC6104826 DOI: 10.1242/jcs.210443] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2017] [Accepted: 05/31/2018] [Indexed: 11/20/2022] Open
Abstract
The cardiac muscle ryanodine receptor-Ca2+ release channel (RyR2) constitutes the sarcoplasmic reticulum (SR) Ca2+ efflux mechanism that initiates myocyte contraction, while cardiac myosin-binding protein-C (cMyBP-C; also known as MYBPC3) mediates regulation of acto-myosin cross-bridge cycling. In this paper, we provide the first evidence for the presence of direct interaction between these two proteins, forming a RyR2-cMyBP-C complex. The C-terminus of cMyBP-C binds with the RyR2 N-terminus in mammalian cells and the interaction is not mediated by a fibronectin-like domain. Notably, we detected complex formation between both recombinant cMyBP-C and RyR2, as well as between the native proteins in cardiac tissue. Cellular Ca2+ dynamics in HEK293 cells is altered upon co-expression of cMyBP-C and RyR2, with lowered frequency of RyR2-mediated spontaneous Ca2+ oscillations, suggesting that cMyBP-C exerts a potential inhibitory effect on RyR2-dependent Ca2+ release. Discovery of a functional RyR2 association with cMyBP-C provides direct evidence for a putative mechanistic link between cytosolic soluble cMyBP-C and SR-mediated Ca2+ release, via RyR2. Importantly, this interaction may have clinical relevance to the observed cMyBP-C and RyR2 dysfunction in cardiac pathologies, such as hypertrophic cardiomyopathy.
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Affiliation(s)
- Paulina J Stanczyk
- Sir Geraint Evans Wales Heart Research Institute, Department of Cardiology, School of Medicine, Cardiff University, Cardiff CF14 4XN, UK.,School of Biosciences, Sir Martin Evans Building, College of Biomedical and Life Sciences, Cardiff University, Cardiff CF10 3AX, UK
| | - Monika Seidel
- Sir Geraint Evans Wales Heart Research Institute, Department of Cardiology, School of Medicine, Cardiff University, Cardiff CF14 4XN, UK.,Swansea University Medical School, Institute of Life Science, Swansea SA2 8PP, UK
| | - Judith White
- Sir Geraint Evans Wales Heart Research Institute, Department of Cardiology, School of Medicine, Cardiff University, Cardiff CF14 4XN, UK.,School of Biosciences, Sir Martin Evans Building, College of Biomedical and Life Sciences, Cardiff University, Cardiff CF10 3AX, UK
| | - Cedric Viero
- Sir Geraint Evans Wales Heart Research Institute, Department of Cardiology, School of Medicine, Cardiff University, Cardiff CF14 4XN, UK.,Institute of Pharmacology and Toxicology, Medical School, Saarland University, Homburg/Saar, Germany
| | - Christopher H George
- Sir Geraint Evans Wales Heart Research Institute, Department of Cardiology, School of Medicine, Cardiff University, Cardiff CF14 4XN, UK.,Swansea University Medical School, Institute of Life Science, Swansea SA2 8PP, UK
| | - Spyros Zissimopoulos
- Sir Geraint Evans Wales Heart Research Institute, Department of Cardiology, School of Medicine, Cardiff University, Cardiff CF14 4XN, UK .,Swansea University Medical School, Institute of Life Science, Swansea SA2 8PP, UK
| | - F Anthony Lai
- Sir Geraint Evans Wales Heart Research Institute, Department of Cardiology, School of Medicine, Cardiff University, Cardiff CF14 4XN, UK .,School of Biosciences, Sir Martin Evans Building, College of Biomedical and Life Sciences, Cardiff University, Cardiff CF10 3AX, UK.,College of Medicine, Member of QU Health, Qatar University, P.O. Box 2013, Doha, Qatar
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39
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Philipson DJ, DePasquale EC, Yang EH, Baas AS. Emerging pharmacologic and structural therapies for hypertrophic cardiomyopathy. Heart Fail Rev 2018; 22:879-888. [PMID: 28856513 DOI: 10.1007/s10741-017-9648-x] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/29/2023]
Abstract
Hypertrophic cardiomyopathy is the most common inherited heart disease. Although it was first described over 50 years ago, there has been little in the way of novel disease-specific therapeutic development for these patients. Current treatment practice largely aims at symptomatic control using old drugs made for other diseases and does little to modify the disease course. Septal reduction by surgical myectomy or percutaneous alcohol septal ablation are well-established treatments for pharmacologic-refractory left ventricular outflow tract obstruction in hypertrophic cardiomyopathy patients. In recent years, there has been a relative surge in the development of innovative therapeutics, which aim to target the complex molecular pathophysiology and resulting hemodynamics that underlie hypertrophic cardiomyopathy. Herein, we review the new and emerging therapeutics for hypertrophic cardiomyopathy, which include pharmacologic attenuation of sarcomeric calcium sensitivity, allosteric inhibition of cardiac myosin, myocardial metabolic modulation, and renin-angiotensin-aldosterone system inhibition, as well as structural intervention by percutaneous mitral valve plication and endocardial radiofrequency ablation of septal hypertrophy. In conclusion, while further development of these therapeutic strategies is ongoing, they each mark a significant and promising advancement in treatment for hypertrophic cardiomyopathy patients.
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Affiliation(s)
- Daniel J Philipson
- Department of Medicine, UCLA, 200 UCLA Medical Plaza Suite 420, Los Angeles, CA, 90095, USA.
| | - Eugene C DePasquale
- Ahmanson-UCLA Cardiomyopathy Center, Division of Cardiology, Department of Medicine, UCLA, Los Angeles, CA, USA
| | - Eric H Yang
- Division of Cardiology, Department of Medicine, UCLA, Los Angeles, CA, USA
| | - Arnold S Baas
- Ahmanson-UCLA Cardiomyopathy Center, Division of Cardiology, Department of Medicine, UCLA, Los Angeles, CA, USA
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40
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Point mutations in the tri-helix bundle of the M-domain of cardiac myosin binding protein-C influence systolic duration and delay cardiac relaxation. J Mol Cell Cardiol 2018; 119:116-124. [PMID: 29729251 DOI: 10.1016/j.yjmcc.2018.05.001] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/18/2017] [Revised: 04/06/2018] [Accepted: 05/01/2018] [Indexed: 11/23/2022]
Abstract
Cardiac myosin binding protein-C (cMyBP-C) is an essential regulatory protein required for proper systolic contraction and diastolic relaxation. We previously showed that N'-terminal domains of cMyBP-C stimulate contraction by binding to actin and activating the thin filament in vitro. In principle, thin filament activating effects of cMyBP-C could influence contraction and relaxation rates, or augment force amplitude in vivo. cMyBP-C binding to actin could also contribute to an internal load that slows muscle shortening velocity as previously hypothesized. However, the functional significance of cMyBP-C binding to actin has not yet been established in vivo. We previously identified an actin binding site in the regulatory M-domain of cMyBP-C and described two missense mutations that either increased (L348P) or decreased (E330K) binding affinity of recombinant cMyBP-C N'-terminal domains for actin in vitro. Here we created transgenic mice with either the L348P or E330K mutations to determine the functional significance of cMyBP-C binding to actin in vivo. Results showed that enhanced binding of cMyBP-C to actin in L348P-Tg mice prolonged the time to end-systole and slowed relaxation rates. Reduced interactions between cMyBP-C and actin in E330K-Tg mice had the opposite effect and significantly shortened the duration of ejection. Neither mouse model displayed overt systolic dysfunction, but L348P-Tg mice showed diastolic dysfunction presumably resulting from delayed relaxation. We conclude that cMyBP-C binding to actin contributes to sustained thin filament activation at the end of systole and during isovolumetric relaxation. These results provide the first functional evidence that cMyBP-C interactions with actin influence cardiac function in vivo.
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41
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Singh SR, Zech ATL, Geertz B, Reischmann-Düsener S, Osinska H, Prondzynski M, Krämer E, Meng Q, Redwood C, van der Velden J, Robbins J, Schlossarek S, Carrier L. Activation of Autophagy Ameliorates Cardiomyopathy in Mybpc3-Targeted Knockin Mice. Circ Heart Fail 2017; 10:CIRCHEARTFAILURE.117.004140. [PMID: 29021349 DOI: 10.1161/circheartfailure.117.004140] [Citation(s) in RCA: 37] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/04/2017] [Accepted: 07/26/2017] [Indexed: 12/14/2022]
Abstract
BACKGROUND Alterations in autophagy have been reported in hypertrophic cardiomyopathy (HCM) caused by Danon disease, Vici syndrome, or LEOPARD syndrome, but not in HCM caused by mutations in genes encoding sarcomeric proteins, which account for most of HCM cases. MYBPC3, encoding cMyBP-C (cardiac myosin-binding protein C), is the most frequently mutated HCM gene. METHODS AND RESULTS We evaluated autophagy in patients with HCM carrying MYBPC3 mutations and in a Mybpc3-targeted knockin HCM mouse model, as well as the effect of autophagy modulators on the development of cardiomyopathy in knockin mice. Microtubule-associated protein 1 light chain 3 (LC3)-II protein levels were higher in HCM septal myectomies than in nonfailing control hearts and in 60-week-old knockin than in wild-type mouse hearts. In contrast to wild-type, autophagic flux was blunted and associated with accumulation of residual bodies and glycogen in hearts of 60-week-old knockin mice. We found that Akt-mTORC1 (mammalian target of rapamycin complex 1) signaling was increased, and treatment with 2.24 mg/kg·d rapamycin or 40% caloric restriction for 9 weeks partially rescued cardiomyopathy or heart failure and restored autophagic flux in knockin mice. CONCLUSIONS Altogether, we found that (1) autophagy is altered in patients with HCM carrying MYBPC3 mutations, (2) autophagy is impaired in Mybpc3-targeted knockin mice, and (3) activation of autophagy ameliorated the cardiac disease phenotype in this mouse model. We propose that activation of autophagy might be an attractive option alone or in combination with another therapy to rescue HCM caused by MYBPC3 mutations.
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Affiliation(s)
- Sonia R Singh
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Antonia T L Zech
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Birgit Geertz
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Silke Reischmann-Düsener
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Hanna Osinska
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Maksymilian Prondzynski
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Elisabeth Krämer
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Qinghang Meng
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Charles Redwood
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Jolanda van der Velden
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Jeffrey Robbins
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Saskia Schlossarek
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.)
| | - Lucie Carrier
- From the Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany (S.R.S., A.T.L.Z., B.G., S.R.-D., M.P., E.K., S.S., L.C.); Department of Pediatrics, The Heart Institute, The Cincinnati Children's Hospital Medical Center, OH (S.R.S., H.O., Q.M., J.R.); Radcliffe Department of Medicine, University of Oxford, United Kingdom (C.R.); Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands (J.v.d.V.); and ICIN-Netherlands Heart Institute, Utrecht (J.v.d.V.).
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Stücker S, Kresin N, Carrier L, Friedrich FW. Nebivolol Desensitizes Myofilaments of a Hypertrophic Cardiomyopathy Mouse Model. Front Physiol 2017; 8:558. [PMID: 28824454 PMCID: PMC5539082 DOI: 10.3389/fphys.2017.00558] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2017] [Accepted: 07/17/2017] [Indexed: 01/19/2023] Open
Abstract
Background: Hypertrophic cardiomyopathy (HCM) patients often present with diastolic dysfunction and a normal to supranormal systolic function. To counteract this hypercontractility, guideline therapies advocate treatment with beta-adrenoceptor and Ca2+ channel blockers. One well established pathomechanism for the hypercontractile phenotype frequently observed in HCM patients and several HCM mouse models is an increased myofilament Ca2+ sensitivity. Nebivolol, a commonly used beta-adrenoceptor antagonist, has been reported to lower maximal force development and myofilament Ca2+ sensitivity in rabbit and human heart tissues. The aim of this study was to evaluate the effect of nebivolol in cardiac muscle strips of an established HCM Mybpc3 mouse model. Furthermore, we investigated actions of nebivolol and epigallocatechin-gallate, which has been shown to desensitize myofilaments for Ca2+ in mouse and human HCM models, in cardiac strips of HCM patients with a mutation in the most frequently mutated HCM gene MYBPC3. Methods and Results: Nebivolol effects were tested on contractile parameters and force-Ca2+ relationship of skinned ventricular muscle strips isolated from Mybpc3-targeted knock-in (KI), wild-type (WT) mice and cardiac strips of three HCM patients with MYBPC3 mutations. At baseline, KI strips showed no difference in maximal force development compared to WT mouse heart strips. Neither 1 nor 10 μM nebivolol had an effect on maximal force development in both genotypes. 10 μM nebivolol induced myofilament Ca2+ desensitization in WT strips and to a greater extent in KI strips. Neither 1 nor 10 μM nebivolol had an effect on Ca2+ sensitivity in cardiac muscle strips of three HCM patients with MYBPC3 mutations, whereas epigallocatechin-gallate induced a right shift in the force-Ca2+ curve. Conclusion: Nebivolol induced a myofilament Ca2+ desensitization in both WT and KI strips, which was more pronounced in KI muscle strips. In human cardiac muscle strips of three HCM patients nebivolol had no effect on myofilament Ca2+ sensitivity.
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Affiliation(s)
- Sabrina Stücker
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-EppendorfHamburg, Germany.,German Centre for Cardiovascular Research (DZHK)Hamburg, Germany
| | - Nico Kresin
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-EppendorfHamburg, Germany.,German Centre for Cardiovascular Research (DZHK)Hamburg, Germany
| | - Lucie Carrier
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-EppendorfHamburg, Germany.,German Centre for Cardiovascular Research (DZHK)Hamburg, Germany
| | - Felix W Friedrich
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-EppendorfHamburg, Germany.,German Centre for Cardiovascular Research (DZHK)Hamburg, Germany
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Flenner F, Geertz B, Reischmann-Düsener S, Weinberger F, Eschenhagen T, Carrier L, Friedrich FW. Diltiazem prevents stress-induced contractile deficits in cardiomyocytes, but does not reverse the cardiomyopathy phenotype in Mybpc3-knock-in mice. J Physiol 2017; 595:3987-3999. [PMID: 28090637 DOI: 10.1113/jp273769] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2016] [Accepted: 01/06/2017] [Indexed: 12/16/2022] Open
Abstract
KEY POINTS Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiac illness and can lead to diastolic dysfunction, sudden cardiac death and heart failure. Treatment of HCM patients is empirical and current pharmacological treatments are unable to stop disease progression or reverse hypertrophy. In this study, we tested if the non-dihydropyridine Ca2+ channel blocker diltiazem, which previously showed potential to stop disease progression, can improve the phenotype of a HCM mouse model (Mybpc3-targeted knock-in), which is based on a mutation commonly found in patients. Diltiazem improved contractile function of isolated ventricular cardiomyocytes acutely, but chronic application did not improve the phenotype of adult mice with a fully developed HCM. Our study shows that diltiazem has beneficial effects in HCM, but long-term treatment success is likely to depend on characteristics and cause of HCM and onset of treatment. ABSTRACT Left ventricular hypertrophy, diastolic dysfunction and fibrosis are the main features of hypertrophic cardiomyopathy (HCM). Guidelines recommend β-adrenoceptor or Ca2+ channel antagonists as pharmacological treatment. The Ca2+ channel blocker diltiazem recently showed promising beneficial effects in pre-clinical HCM, particularly in patients carrying MYBPC3 mutations. In the present study we evaluated whether diltiazem could ameliorate or reverse the disease phenotype in cells and in vivo in an Mybpc3-targeted knock-in (KI) mouse model of HCM. Sarcomere shortening and Ca2+ transients were measured in KI and wild-type (WT) cardiomyocytes in basal conditions (1-Hz pacing) and under stress conditions (30 nm isoprenaline, 5-Hz pacing) with or without pre-treatment with 1 μm diltiazem. KI cardiomyocytes exhibited lower diastolic sarcomere length (dSL) at baseline, a tendency to a stronger positive inotropic response to isoprenaline than WT, a marked reduction of dSL and a tendency towards arrhythmias under stress conditions. Pre-treatment of cardiomyocytes with 1 μm diltiazem reduced the drop in dSL and arrhythmia frequency in KI, and attenuated the positive inotropic effect of isoprenaline. Furthermore, diltiazem reduced the contraction amplitude at 5 Hz but did not affect diastolic Ca2+ load and Ca2+ transient amplitude. Six months of diltiazem treatment of KI mice did not reverse cardiac hypertrophy and dysfunction, activation of the fetal gene program or fibrosis. In conclusion, diltiazem blunted the response to isoprenaline in WT and KI cardiomyocytes and improved diastolic relaxation under stress conditions in KI cardiomyocytes. This beneficial effect of diltiazem in cells did not translate in therapeutic efficacy when applied chronically in KI mice.
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Affiliation(s)
- Frederik Flenner
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Centre, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Birgit Geertz
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Centre, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Silke Reischmann-Düsener
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Centre, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Florian Weinberger
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Centre, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Thomas Eschenhagen
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Centre, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Lucie Carrier
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Centre, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
| | - Felix W Friedrich
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Centre, University Medical Centre Hamburg-Eppendorf, Hamburg, Germany.,DZHK (German Centre for Cardiovascular Research), partner site Hamburg/Kiel/Lübeck, Germany
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Friedrich FW, Flenner F, Nasib M, Eschenhagen T, Carrier L. Epigallocatechin-3-Gallate Accelerates Relaxation and Ca 2+ Transient Decay and Desensitizes Myofilaments in Healthy and Mybpc3-Targeted Knock-in Cardiomyopathic Mice. Front Physiol 2016; 7:607. [PMID: 27994558 PMCID: PMC5136558 DOI: 10.3389/fphys.2016.00607] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2016] [Accepted: 11/22/2016] [Indexed: 11/13/2022] Open
Abstract
Background: Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiac muscle disease with left ventricular hypertrophy, interstitial fibrosis and diastolic dysfunction. Increased myofilament Ca2+ sensitivity could be the underlying cause of diastolic dysfunction. Epigallocatechin-3-gallate (EGCg), a catechin found in green tea, has been reported to decrease myofilament Ca2+ sensitivity in HCM models with troponin mutations. However, whether this is also the case for HCM-associated thick filament mutations is not known. Therefore, we evaluated whether EGCg affects the behavior of cardiomyocytes and myofilaments of an HCM mouse model carrying a gene mutation in cardiac myosin-binding protein C and exhibiting both increased myofilament Ca2+ sensitivity and diastolic dysfunction. Methods and Results: Acute effects of EGCg were tested on fractional sarcomere shortening and Ca2+ transients in intact ventricular myocytes and on force-Ca2+ relationship of skinned ventricular muscle strips isolated from Mybpc3-targeted knock-in (KI) and wild-type (WT) mice. Fractional sarcomere shortening and Ca2+ transients were analyzed at 37°C under 1-Hz pacing in the absence or presence of EGCg (1.8 μM). At baseline and in the absence of Fura-2, KI cardiomyocytes displayed lower diastolic sarcomere length, higher fractional sarcomere shortening, longer time to peak shortening and time to 50% relengthening than WT cardiomyocytes. In WT and KI neither diastolic sarcomere length nor fractional sarcomere shortening were influenced by EGCg treatment, but relaxation time was reduced, to a greater extent in KI cells. EGCg shortened time to peak Ca2+ and Ca2+ transient decay in Fura-2-loaded WT and KI cardiomyocytes. EGCg did not influence phosphorylation of phospholamban. In skinned cardiac muscle strips, EGCg (30 μM) decreased Ca2+ sensitivity in both groups. Conclusion: EGCg hastened relaxation and Ca2+ transient decay to a larger extent in KI than in WT cardiomyocytes. This effect could be partially explained by myofilament Ca2+ desensitization.
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Affiliation(s)
- Felix W Friedrich
- Cardiovascular Research Center, Department of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-EppendorfHamburg, Germany; German Centre for Cardiovascular Research (DZHK)Hamburg, Germany
| | - Frederik Flenner
- Cardiovascular Research Center, Department of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-EppendorfHamburg, Germany; German Centre for Cardiovascular Research (DZHK)Hamburg, Germany
| | - Mahtab Nasib
- Cardiovascular Research Center, Department of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-EppendorfHamburg, Germany; German Centre for Cardiovascular Research (DZHK)Hamburg, Germany
| | - Thomas Eschenhagen
- Cardiovascular Research Center, Department of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-EppendorfHamburg, Germany; German Centre for Cardiovascular Research (DZHK)Hamburg, Germany
| | - Lucie Carrier
- Cardiovascular Research Center, Department of Experimental Pharmacology and Toxicology, University Medical Center Hamburg-EppendorfHamburg, Germany; German Centre for Cardiovascular Research (DZHK)Hamburg, Germany
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Kobirumaki-Shimozawa F, Oyama K, Shimozawa T, Mizuno A, Ohki T, Terui T, Minamisawa S, Ishiwata S, Fukuda N. Nano-imaging of the beating mouse heart in vivo: Importance of sarcomere dynamics, as opposed to sarcomere length per se, in the regulation of cardiac function. ACTA ACUST UNITED AC 2016; 147:53-62. [PMID: 26712849 PMCID: PMC4692490 DOI: 10.1085/jgp.201511484] [Citation(s) in RCA: 42] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023]
Abstract
¡Vive la différence! In cardiac contraction, the reduction in sarcomere length—rather than length itself—determines contractile force. Sarcomeric contraction in cardiomyocytes serves as the basis for the heart’s pump functions in mammals. Although it plays a critical role in the circulatory system, myocardial sarcomere length (SL) change has not been directly measured in vivo under physiological conditions because of technical difficulties. In this study, we developed a high speed (100–frames per second), high resolution (20-nm) imaging system for myocardial sarcomeres in living mice. Using this system, we conducted three-dimensional analysis of sarcomere dynamics in left ventricular myocytes during the cardiac cycle, simultaneously with electrocardiogram and left ventricular pressure measurements. We found that (a) the working range of SL was on the shorter end of the resting distribution, and (b) the left ventricular–developed pressure was positively correlated with the SL change between diastole and systole. The present findings provide the first direct evidence for the tight coupling of sarcomere dynamics and ventricular pump functions in the physiology of the heart.
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Affiliation(s)
- Fuyu Kobirumaki-Shimozawa
- Department of Cell Physiology and Department of Anesthesiology, The Jikei University School of Medicine, Minato-ku, Tokyo 105-8461, Japan
| | - Kotaro Oyama
- Department of Cell Physiology and Department of Anesthesiology, The Jikei University School of Medicine, Minato-ku, Tokyo 105-8461, Japan Department of Physics and Department of Life Science and Medical Bioscience, School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 169-8555, Japan
| | - Togo Shimozawa
- Department of Physics and Department of Life Science and Medical Bioscience, School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 169-8555, Japan
| | - Akari Mizuno
- Department of Physics and Department of Life Science and Medical Bioscience, School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 169-8555, Japan
| | - Takashi Ohki
- Department of Physics and Department of Life Science and Medical Bioscience, School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 169-8555, Japan
| | - Takako Terui
- Department of Cell Physiology and Department of Anesthesiology, The Jikei University School of Medicine, Minato-ku, Tokyo 105-8461, Japan
| | - Susumu Minamisawa
- Department of Cell Physiology and Department of Anesthesiology, The Jikei University School of Medicine, Minato-ku, Tokyo 105-8461, Japan
| | - Shin'ichi Ishiwata
- Department of Physics and Department of Life Science and Medical Bioscience, School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 169-8555, Japan Waseda Bioscience Research Institute in Singapore, Waseda University, Helios, Singapore 138667
| | - Norio Fukuda
- Department of Cell Physiology and Department of Anesthesiology, The Jikei University School of Medicine, Minato-ku, Tokyo 105-8461, Japan
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Wijnker PJM, Friedrich FW, Dutsch A, Reischmann S, Eder A, Mannhardt I, Mearini G, Eschenhagen T, van der Velden J, Carrier L. Comparison of the effects of a truncating and a missense MYBPC3 mutation on contractile parameters of engineered heart tissue. J Mol Cell Cardiol 2016; 97:82-92. [PMID: 27108529 DOI: 10.1016/j.yjmcc.2016.03.003] [Citation(s) in RCA: 52] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/18/2015] [Revised: 03/08/2016] [Accepted: 03/09/2016] [Indexed: 12/25/2022]
Abstract
Hypertrophic cardiomyopathy (HCM) is a cardiac genetic disease characterized by left ventricular hypertrophy, diastolic dysfunction and myocardial disarray. The most frequently mutated gene is MYBPC3, encoding cardiac myosin-binding protein-C (cMyBP-C). We compared the pathomechanisms of a truncating mutation (c.2373_2374insG) and a missense mutation (c.1591G>C) in MYBPC3 in engineered heart tissue (EHT). EHTs enable to study the direct effects of mutants without interference of secondary disease-related changes. EHTs were generated from Mybpc3-targeted knock-out (KO) and wild-type (WT) mouse cardiac cells. MYBPC3 WT and mutants were expressed in KO EHTs via adeno-associated virus. KO EHTs displayed higher maximal force and sensitivity to external [Ca(2+)] than WT EHTs. Expression of WT-Mybpc3 at MOI-100 resulted in ~73% cMyBP-C level but did not prevent the KO phenotype, whereas MOI-300 resulted in ≥95% cMyBP-C level and prevented the KO phenotype. Expression of the truncating or missense mutation (MOI-300) or their combination with WT (MOI-150 each), mimicking the homozygous or heterozygous disease state, respectively, failed to restore force to WT level. Immunofluorescence analysis revealed correct incorporation of WT and missense, but not of truncated cMyBP-C in the sarcomere. In conclusion, this study provides evidence in KO EHTs that i) haploinsufficiency affects EHT contractile function if WT cMyBP-C protein levels are ≤73%, ii) missense or truncating mutations, but not WT do not fully restore the disease phenotype and have different pathogenic mechanisms, e.g. sarcomere poisoning for the missense mutation, iii) the direct impact of (newly identified) MYBPC3 gene variants can be evaluated.
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Affiliation(s)
- Paul J M Wijnker
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany; Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands
| | - Felix W Friedrich
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Alexander Dutsch
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Silke Reischmann
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Alexandra Eder
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Ingra Mannhardt
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Giulia Mearini
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Thomas Eschenhagen
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Jolanda van der Velden
- Department of Physiology, Institute for Cardiovascular Research, VU University Medical Center, Amsterdam, The Netherlands
| | - Lucie Carrier
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany.
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Stathopoulou K, Wittig I, Heidler J, Piasecki A, Richter F, Diering S, van der Velden J, Buck F, Donzelli S, Schröder E, Wijnker PJM, Voigt N, Dobrev D, Sadayappan S, Eschenhagen T, Carrier L, Eaton P, Cuello F. S-glutathiolation impairs phosphoregulation and function of cardiac myosin-binding protein C in human heart failure. FASEB J 2016; 30:1849-64. [PMID: 26839380 DOI: 10.1096/fj.201500048] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2015] [Accepted: 01/13/2016] [Indexed: 11/11/2022]
Abstract
Cardiac myosin-binding protein C (cMyBP-C) regulates actin-myosin interaction and thereby cardiac myocyte contraction and relaxation. This physiologic function is regulated by cMyBP-C phosphorylation. In our study, reduced site-specific cMyBP-C phosphorylation coincided with increased S-glutathiolation in ventricular tissue from patients with dilated or ischemic cardiomyopathy compared to nonfailing donors. We used redox proteomics, to identify constitutive and disease-specific S-glutathiolation sites in cMyBP-C in donor and patient samples, respectively. Among those, a cysteine cluster in the vicinity of the regulatory phosphorylation sites within the myosin S2 interaction domain C1-M-C2 was identified and showed enhanced S-glutathiolation in patients. In vitro S-glutathiolation of recombinant cMyBP-C C1-M-C2 occurred predominantly at Cys(249), which attenuated phosphorylation by protein kinases. Exposure to glutathione disulfide induced cMyBP-C S-glutathiolation, which functionally decelerated the kinetics of Ca(2+)-activated force development in ventricular myocytes from wild-type, but not those from Mybpc3-targeted knockout mice. These oxidation events abrogate protein kinase-mediated phosphorylation of cMyBP-C and therefore potentially contribute to the reduction of its phosphorylation and the contractile dysfunction observed in human heart failure.-Stathopoulou, K., Wittig, I., Heidler, J., Piasecki, A., Richter, F., Diering, S., van der Velden, J., Buck, F., Donzelli, S., Schröder, E., Wijnker, P. J. M., Voigt, N., Dobrev, D., Sadayappan, S., Eschenhagen, T., Carrier, L., Eaton, P., Cuello, F. S-glutathiolation impairs phosphoregulation and function of cardiac myosin-binding protein C in human heart failure.
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Affiliation(s)
- Konstantina Stathopoulou
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Frankfurt, Germany
| | - Ilka Wittig
- Functional Proteomics, SFB 815 Core Unit, Faculty of Medicine, Goethe University, Frankfurt am Main, Germany; Cluster of Excellence "Macromolecular Complexes," Goethe University, Frankfurt am Main, Germany; Partner Site Rhein/Main, Frankfurt, Germany
| | - Juliana Heidler
- Functional Proteomics, SFB 815 Core Unit, Faculty of Medicine, Goethe University, Frankfurt am Main, Germany; Cluster of Excellence "Macromolecular Complexes," Goethe University, Frankfurt am Main, Germany; Partner Site Rhein/Main, Frankfurt, Germany
| | - Angelika Piasecki
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Frankfurt, Germany
| | - Florian Richter
- Functional Proteomics, SFB 815 Core Unit, Faculty of Medicine, Goethe University, Frankfurt am Main, Germany; Cluster of Excellence "Macromolecular Complexes," Goethe University, Frankfurt am Main, Germany; Partner Site Rhein/Main, Frankfurt, Germany
| | - Simon Diering
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Frankfurt, Germany
| | - Jolanda van der Velden
- Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Amsterdam, The Netherlands; ICIN-The Netherlands Heart Institute, Utrecht, The Netherlands
| | - Friedrich Buck
- Department of Clinical Chemistry/Central Laboratories, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Sonia Donzelli
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Frankfurt, Germany
| | - Ewald Schröder
- King's British Heart Foundation Centre, King's College London, London, United Kingdom
| | - Paul J M Wijnker
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Frankfurt, Germany; Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Amsterdam, The Netherlands
| | - Niels Voigt
- Institute of Pharmacology, Faculty of Medicine, University Duisburg-Essen, Essen, Germany; and
| | - Dobromir Dobrev
- Institute of Pharmacology, Faculty of Medicine, University Duisburg-Essen, Essen, Germany; and
| | - Sakthivel Sadayappan
- Department of Cell and Molecular Physiology, Loyola University, Chicago, Maywood, Illinois, USA
| | - Thomas Eschenhagen
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Frankfurt, Germany
| | - Lucie Carrier
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Frankfurt, Germany
| | - Philip Eaton
- King's British Heart Foundation Centre, King's College London, London, United Kingdom
| | - Friederike Cuello
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Frankfurt, Germany;
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Najafi A, Sequeira V, Helmes M, Bollen IAE, Goebel M, Regan JA, Carrier L, Kuster DWD, Van Der Velden J. Selective phosphorylation of PKA targets after β-adrenergic receptor stimulation impairs myofilament function in Mybpc3-targeted HCM mouse model. Cardiovasc Res 2016; 110:200-14. [PMID: 26825555 DOI: 10.1093/cvr/cvw026] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/08/2015] [Accepted: 01/22/2016] [Indexed: 12/19/2022] Open
Abstract
AIMS Hypertrophic cardiomyopathy (HCM) has been associated with reduced β-adrenergic receptor (β-AR) signalling, leading downstream to a low protein kinase A (PKA)-mediated phosphorylation. It remained undefined whether all PKA targets will be affected similarly by diminished β-AR signalling in HCM. We aimed to investigate the role of β-AR signalling on regulating myofilament and calcium handling in an HCM mouse model harbouring a gene mutation (G > A transition on the last nucleotide of exon 6) in Mybpc3 encoding cardiac myosin-binding protein C. METHODS AND RESULTS Cardiomyocyte contractile properties and phosphorylation state were measured in left ventricular permeabilized and intact cardiomyocytes isolated from heterozygous (HET) or homozygous (KI) Mybpc3-targeted knock-in mice. Significantly higher myofilament Ca²⁺sensitivity and passive tension were detected in KI mice, which were normalized after PKA treatment. Loaded intact cardiomyocyte force-sarcomere length relation was impaired in both HET and KI mice, suggesting a reduced length-dependent activation. Unloaded cardiomyocyte function revealed an impaired myofilament contractile response to isoprenaline (ISO) in KI, whereas the calcium-handling response to ISO was maintained. This disparity was explained by an attenuated increase in cardiac troponin I (cTnI) phosphorylation in KI, whereas the increase in phospholamban (PLN) phosphorylation was maintained to wild-type values. CONCLUSION These data provide evidence that in the KI HCM mouse model, β-AR stimulation leads to preferential PKA phosphorylation of PLN over cTnI, resulting in an impaired inotropic and lusitropic response.
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Affiliation(s)
- Aref Najafi
- Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Netherlands ICIN-Netherlands Heart Institute, Utrecht, The Netherlands
| | - Vasco Sequeira
- Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Netherlands
| | - Michiel Helmes
- Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Netherlands
| | - Ilse A E Bollen
- Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Netherlands
| | - Max Goebel
- Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Netherlands
| | - Jessica A Regan
- Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Netherlands
| | - Lucie Carrier
- Institute of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Germany
| | - Diederik W D Kuster
- Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Netherlands
| | - Jolanda Van Der Velden
- Department of Physiology, Institute for Cardiovascular Research (ICaR-VU), VU University Medical Center Amsterdam, Netherlands ICIN-Netherlands Heart Institute, Utrecht, The Netherlands
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ADP-stimulated contraction: A predictor of thin-filament activation in cardiac disease. Proc Natl Acad Sci U S A 2015; 112:E7003-12. [PMID: 26621701 DOI: 10.1073/pnas.1513843112] [Citation(s) in RCA: 30] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Diastolic dysfunction is general to all idiopathic dilated (IDCM) and hypertrophic cardiomyopathy (HCM) patients. Relaxation deficits may result from increased actin-myosin formation during diastole due to altered tropomyosin position, which blocks myosin binding to actin in the absence of Ca(2+). We investigated whether ADP-stimulated force development (without Ca(2+)) can be used to reveal changes in actin-myosin blockade in human cardiomyopathy cardiomyocytes. Cardiac samples from HCM patients, harboring thick-filament (MYH7mut, MYBPC3mut) and thin-filament (TNNT2mut, TNNI3mut) mutations, and IDCM were compared with sarcomere mutation-negative HCM (HCMsmn) and nonfailing donors. Myofilament ADP sensitivity was higher in IDCM and HCM compared with donors, whereas it was lower for MYBPC3. Increased ADP sensitivity in IDCM, HCMsmn, and MYH7mut was caused by low phosphorylation of myofilament proteins, as it was normalized to donors by protein kinase A (PKA) treatment. Troponin exchange experiments in a TNNT2mut sample corrected the abnormal actin-myosin blockade. In MYBPC3trunc samples, ADP sensitivity highly correlated with cardiac myosin-binding protein-C (cMyBP-C) protein level. Incubation of cardiomyocytes with cMyBP-C antibody against the actin-binding N-terminal region reduced ADP sensitivity, indicative of cMyBP-C's role in actin-myosin regulation. In the presence of Ca(2+), ADP increased myofilament force development and sarcomere stiffness. Enhanced sarcomere stiffness in sarcomere mutation-positive HCM samples was irrespective of the phosphorylation background. In conclusion, ADP-stimulated contraction can be used as a tool to study how protein phosphorylation and mutant proteins alter accessibility of myosin binding on actin. In the presence of Ca(2+), pathologic [ADP] and low PKA-phosphorylation, high actin-myosin formation could contribute to the impaired myocardial relaxation observed in cardiomyopathies.
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50
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Flenner F, Friedrich FW, Ungeheuer N, Christ T, Geertz B, Reischmann S, Wagner S, Stathopoulou K, Söhren KD, Weinberger F, Schwedhelm E, Cuello F, Maier LS, Eschenhagen T, Carrier L. Ranolazine antagonizes catecholamine-induced dysfunction in isolated cardiomyocytes, but lacks long-term therapeutic effects in vivo in a mouse model of hypertrophic cardiomyopathy. Cardiovasc Res 2015; 109:90-102. [PMID: 26531128 DOI: 10.1093/cvr/cvv247] [Citation(s) in RCA: 36] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/13/2015] [Accepted: 10/27/2015] [Indexed: 12/19/2022] Open
Abstract
AIMS Hypertrophic cardiomyopathy (HCM) is often accompanied by increased myofilament Ca(2+) sensitivity and diastolic dysfunction. Recent findings indicate increased late Na(+) current density in human HCM cardiomyocytes. Since ranolazine has the potential to decrease myofilament Ca(2+) sensitivity and late Na(+) current, we investigated its effects in an Mybpc3-targeted knock-in (KI) mouse model of HCM. METHODS AND RESULTS Unloaded sarcomere shortening and Ca(2+) transients were measured in KI and wild-type (WT) cardiomyocytes. Measurements were performed at baseline (1 Hz) and under increased workload (30 nM isoprenaline (ISO), 5 Hz) in the absence or presence of 10 µM ranolazine. KI myocytes showed shorter diastolic sarcomere length at baseline, stronger inotropic response to ISO, and drastic drop of diastolic sarcomere length under increased workload. Ranolazine attenuated ISO responses in WT and KI cells and prevented workload-induced diastolic failure in KI. Late Na(+) current density was diminished and insensitive to ranolazine in KI cardiomyocytes. Ca(2+) sensitivity of skinned KI trabeculae was slightly decreased by ranolazine. Phosphorylation analysis of cAMP-dependent protein kinase A-target proteins and ISO concentration-response measurements on muscle strips indicated antagonism at β-adrenoceptors with 10 µM ranolazine shifting the ISO response by 0.6 log units. Six-month treatment with ranolazine (plasma level >20 µM) demonstrated a β-blocking effect, but did not reverse cardiac hypertrophy or dysfunction in KI mice. CONCLUSION Ranolazine improved tolerance to high workload in mouse HCM cardiomyocytes, not by blocking late Na(+) current, but by antagonizing β-adrenergic stimulation and slightly desensitizing myofilaments to Ca(2+). This effect did not translate in therapeutic efficacy in vivo.
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Affiliation(s)
- Frederik Flenner
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Felix W Friedrich
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Nele Ungeheuer
- Clinic for Cardiology and Pneumology, Georg-August-University Göttingen, Göttingen, Germany
| | - Torsten Christ
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Birgit Geertz
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Silke Reischmann
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Stefan Wagner
- Clinic for Cardiology and Pneumology, Georg-August-University Göttingen, Göttingen, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Göttingen, Göttingen, Germany Department for Internal Medicine II, University Hospital Regensburg, Regensburg, Germany
| | - Konstantina Stathopoulou
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Klaus-Dieter Söhren
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany
| | - Florian Weinberger
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Edzard Schwedhelm
- DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany Department of Clinical Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Friederike Cuello
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Lars S Maier
- Clinic for Cardiology and Pneumology, Georg-August-University Göttingen, Göttingen, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Göttingen, Göttingen, Germany Department for Internal Medicine II, University Hospital Regensburg, Regensburg, Germany
| | - Thomas Eschenhagen
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
| | - Lucie Carrier
- Department of Experimental Pharmacology and Toxicology, Cardiovascular Research Center, University Medical Center Hamburg-Eppendorf, Martinistraße 52, D-20246 Hamburg, Germany DZHK (German Centre for Cardiovascular Research), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany
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