Phosphomimetic cardiac myosin-binding protein C partially rescues a cardiomyopathy phenotype in murine engineered heart tissue

Dutsch, Alexander; Wijnker, Paul J.M.; Schlossarek, Saskia; Friedrich, Felix W.; Krämer, Elisabeth; Braren, Ingke; Hirt, Marc N.; Brenière‐Letuffe, David; Rhoden, Alexandra; Mannhardt, Ingra; Eschenhagen, Thomas; Carrier, Lucie; Mearini, Giulia

doi:10.1038/s41598-019-54665-2

Cited by 16 publications

(12 citation statements)

References 56 publications

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“…On the other hand, human MYBPC3hom EHTs exhibited lower beating frequency and higher RR-scatter that reflects arrhythmic behaviors, which are in agreement with previous findings in mouse Hom-KI EHTs [28], suggesting that a low level of mutant cMyBP-C protein as found in both homozygous human and mouse models is associated with this phenotype [28]. Similarly, intact or skinned MYBPC3hom EHTs developed higher maximal force and faster relaxation time, in agreement with previous findings obtained in EHTs derived from neonatal Hom-KI or Hom-knock-out (KO) cardiac cells [28,29,48]. The shorter relaxation time in all these models with a low amount of cMyBP-C is consistent with the shortening of the ejection phase of systole in Hom-KO mice [57] and supports the role of cMyBP-C in sustaining the dynamic range of myosin-actin interaction during contraction twitch [31,[57][58][59].…”

Section: Discussionsupporting

confidence: 91%

“…This supports previous data obtained for other human HCM EHTs or septal myectomies [22,47]. Relaxation time was shorter in MYBPC3hom EHTs, exhibiting a low amount of cMyBP-C, which is in agreement with previous findings in EHTs derived from neonatal Hom-KI cardiac cells [28,48].…”

Section: Functional Measurements In Intact and Skinned Human Engineered Heart Tissuessupporting

confidence: 93%

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Translational investigation of electrophysiology in hypertrophic cardiomyopathy

Flenner

Jungen

Küpker

et al. 2021

Journal of Molecular and Cellular Cardiology

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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 Ca 2+ 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 ( MYBPC3 hom) 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 Ca 2+ 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 MYBPC3 hom 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-Ca 2+ relation in skinned EHTs. Furthermore, MYBPC3 hom 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 Ca 2+ 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 fro...

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Section: Discussionsupporting

confidence: 91%

Section: Functional Measurements In Intact and Skinned Human Engineered Heart Tissuessupporting

confidence: 93%

Translational investigation of electrophysiology in hypertrophic cardiomyopathy

Flenner

Jungen

Küpker

et al. 2021

Journal of Molecular and Cellular Cardiology

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“…Moreover, it is possible to genetically modify each of these cellular components to gain mechanistic insights into the pathogenesis of various diseases including COVID-19 myocarditis. These observations add to an emerging field that have successfully utilized EHT systems for testing of compounds for cardiotoxicity and efficacy 47,48 . From a broader perspective, our findings are consistent with the emerging utility of human organoid systems in the investigation of SARS-CoV-2 38,39,49,50 .…”

Section: Discussionmentioning

confidence: 79%

“…These observations add to an emerging field that have successfully utilized EHT systems for testing of compounds for cardiotoxicity and efficacy 47,48 . known mutations associated with heart disease and the stem cells are pluripotent as assessed by immunofluorescence staining .…”

Section: Discussionmentioning

confidence: 83%

SARS-CoV-2 Infects Human Engineered Heart Tissues and Models COVID-19 Myocarditis

Bailey

Dmytrenko

Greenberg

et al. 2020

Preprint

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Epidemiological studies of the COVID-19 pandemic have revealed evidence of cardiac involvement and documented that myocardial injury and myocarditis are predictors of poor outcomes. Nonetheless, little is understood regarding SARS-CoV-2 tropism within the heart and whether cardiac complications result directly from myocardial infection. Here, we develop a human engineered heart tissue model and demonstrate that SARS-CoV-2 selectively infects cardiomyocytes. Viral infection is dependent on expression of angiotensin-I converting enzyme 2 (ACE2) and endosomal cysteine proteases, suggesting an endosomal mechanism of cell entry. After infection with SARS-CoV-2, engineered tissues display typical features of myocarditis, including cardiomyocyte cell death, impaired cardiac contractility, and innate immune cell activation. Consistent with these findings, autopsy tissue obtained from individuals with COVID-19 myocarditis demonstrated cardiomyocyte infection, cell death, and macrophage-predominate immune cell infiltrate. These findings establish human cardiomyocyte tropism for SARS-CoV-2 and provide an experimental platform for interrogating and mitigating cardiac complications of COVID-19.

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“…It has been reported that a single systemic dose of AAV9-carrying MYBPC3 cDNA was able to restore MYBPC3 mRNA and protein levels and prevent the development of left ventricular hypertrophy (LVH) in MYBPC3-targeted knock-in mice with only 20% cMyBP-C protein 30 . Another study reported that AAV6-carrying MYBPC3 cDNA was able to achieve similar improvement in engineered 3D heart tissue 31,32 . In addition, AAV9-carrying MYBPC3 cDNA has shown to be able to suppress the development of hypertrophy in human iPSC-derived cardiomyocytes from HCM patients 33 .…”

Section: Discussionmentioning

confidence: 97%

Modulation of Myosin by Cardiac Myosin Binding Protein-C Peptides Improves Cardiac Contractility in Ex-Vivo Experimental Heart Failure Models

Hou¹,

Kumar²,

Anand³

et al. 2021

Preprint

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Cardiac myosin binding protein-C (cMyBP-C) is an important regulator of sarcomeric function. Although reduced phosphorylation of cMyBP-C has been linked to compromised contractility in heart failure patients, direct modulation of cMyBP-C to myosin using small molecules or peptides has not been reported to improve cardiac performance. Here we used previously published cMyBP-C peptides 302A and 302S (surrogates to the regulatory phosphorylation site serine 302) as tool molecules to investigate the role of cMyBP-C in modulating cardiac contraction and relaxation in experimental heart failure (HF) models in vitro. cMyBP-C peptides 302A and 302S were able to increase contractility of papillary muscle fibers isolated from a cMyBP-C phospho-ablation (cMyBP-CAAA) mouse model. In addition, 302A was able to improve the force redevelopment rate (ktr) in papillary muscle fibers from cMyBP-CAAA mice. Consistent with above findings, cMyBP-C peptides 302A and 302S were able to increase the ATPase rates in myofibrils isolated from MI rats but not from sham rats. Furthermore, in cMyBP-CAAA mouse and myocardial infarction (MI) HF models, both cMyBP-C peptides 302A and 302S were able to improve ATPase hydrolysis rates. These changes were not observed in non-transgenic (NTG) mice or sham rats, indicating the specific effects of these peptides in regulating the reduced or unphosphorylated state of cMyBP-C only under pathological conditions of heart failure. Taken together, these studies demonstrate that modulation of cMyBP-C in a reduced phosphorylation or unphosphorylated state can be a therapeutic approach to improve myosin function, sarcomere contractility and relaxation. Therefore, targeting cMyBP-C can be a differentiated approach to improve overall cardiac performance on top of standard care drugs in HF patients.

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Phosphomimetic cardiac myosin-binding protein C partially rescues a cardiomyopathy phenotype in murine engineered heart tissue

Cited by 16 publications

References 56 publications

Translational investigation of electrophysiology in hypertrophic cardiomyopathy

Translational investigation of electrophysiology in hypertrophic cardiomyopathy

SARS-CoV-2 Infects Human Engineered Heart Tissues and Models COVID-19 Myocarditis

Modulation of Myosin by Cardiac Myosin Binding Protein-C Peptides Improves Cardiac Contractility in Ex-Vivo Experimental Heart Failure Models

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