bims-cesiha 2021-02-28 papers

bims-cesiha

Biomed News

on Cell signalling in the heart

Issue of 2021–02–28
thirty papers selected by
Danae Angelidaki, Max Planck Institute for Biology of Ageing

Cx43 hemichannel microdomain signaling at the intercalated disc enhances cardiac excitability.
Metformin-Enhanced Cardiac AMP-Activated Protein Kinase/Atrogin-1 Pathways Inhibit Charged Multivesicular Body Protein 2B Accumulation in Ischemia-Reperfusion Injury.
Activation of the cardiac non-neuronal cholinergic system prevents the development of diabetes-associated cardiovascular complications.
Dapagliflozin attenuates hypoxia/reoxygenation-caused cardiac dysfunction and oxidative damage through modulation of AMPK.
Meteorin-like/Meteorin-β protects heart against cardiac dysfunction.
Interleukin 35 protects cardiomyocytes following ischemia/reperfusion-induced apoptosis via activation of mitochondrial STAT3.
PHD Finger Protein 19 Promotes Cardiac Hypertrophy via Epigenetically Regulating SIRT2.
High glucose induces Drp1-mediated mitochondrial fission via the Orai1 calcium channel to participate in diabetic cardiomyocyte hypertrophy.
Single Nuclei Sequencing Reveals Novel Insights into the Regulation of Cellular Signatures in Children with Dilated Cardiomyopathy.
Functional analysis of a gene-edited mouse model to gain insights into the disease mechanisms of a titin missense variant.
MiR-200a-3p Aggravates DOX-Induced Cardiotoxicity by Targeting PEG3 Through SIRT1/NF-κB Signal Pathway.
Endoplasmic reticulum stress and unfolded protein response in cardiovascular diseases.
Paclitaxel mitigates structural alterations and cardiac conduction system defects in a mouse model of Hutchinson-Gilford progeria syndrome.
Exosomes Derived From Hypertrophic Cardiomyocytes Induce Inflammation in Macrophages via miR-155 Mediated MAPK Pathway.
Cardiovascular phenotype of the Dmdmdx rat - a suitable animal model for Duchenne muscular dystrophy.
Consistent Long-Term Therapeutic Efficacy of Human Umbilical Cord Matrix-Derived Mesenchymal Stromal Cells After Myocardial Infarction Despite Individual Differences and Transient Engraftment.
Global identification of S-palmitoylated proteins and detection of palmitoylating (DHHC) enzymes in heart.
CircSLC8A1 and circNFIX can be used as auxiliary diagnostic markers for sudden cardiac death caused by acute ischemic heart disease.
Mapping genetic changes in the cAMP-signaling cascade in human atria.
Maturation of Pluripotent Stem Cell-Derived Cardiomyocytes Enables Modeling of Human Hypertrophic Cardiomyopathy.
Use of stable isotope-tagged thymidine and multi-isotope imaging mass spectrometry (MIMS) for quantification of human cardiomyocyte division.
Asymmetric Hapln1a drives regionalised cardiac ECM expansion and promotes heart morphogenesis in zebrafish development.
Effect of doxorubicin on cardiac lipid metabolism-related transcriptome and the protective activity of Alda-1.
Peritransplant Cardiometabolic and Mitochondrial Function: The Missing Piece in Donor Heart Dysfunction and Graft Failure.
Epigenetic regulation of cardiac electrophysiology in atrial fibrillation: HDAC2 determines action potential duration and suppresses NRSF in cardiomyocytes.
Cateslytin abrogates lipopolysaccharide-induced cardiomyocyte injury by reducing inflammation and oxidative stress through toll like receptor 4 interaction.
Predictive values of multiple non-invasive markers for myocardial fibrosis in hypertrophic cardiomyopathy patients with preserved ejection fraction.
Deletion of Orphan G Protein-Coupled Receptor GPR37L1 in Mice Alters Cardiovascular Homeostasis in a Sex-Specific Manner.
Dexpramipexole attenuates myocardial ischemia/reperfusion injury through upregulation of mitophagy.
Optogenetic current in myofibroblasts acutely alters electrophysiology and conduction of co-cultured cardiomyocytes.

J Clin Invest. 2021 Feb 23. pii: 137752. [Epub ahead of print]

Cx43 hemichannel microdomain signaling at the intercalated disc enhances cardiac excitability.

Maarten Aj De Smet, Alessio Lissoni, Timur Nezlobinsky, Nan Wang, Eef Dries, Marta Pérez-Hernández, Xianming Lin, Matthew Amoni, Tim Vervliet, Katja Witschas, Eli Rothenberg, Geert Bultynck, Rainer Schulz, Alexander V Panfilov, Mario Delmar, Karin R Sipido, Luc Leybaert.

  Cx43, a major cardiac connexin, forms precursor hemichannels that accrue at the intercalated disc to assemble as gap junctions. While gap junctions are crucial for electrical conduction in the heart, little is known on potential roles of hemichannels. Recent evidence suggests that inhibiting Cx43 hemichannel opening with Gap19 has antiarrhythmic effects. Here, we used multiple electrophysiology, imaging and super-resolution techniques to understand and define the conditions underlying Cx43 hemichannel activation in ventricular cardiomyocytes, their contribution to diastolic Ca2+ release from the sarcoplasmic reticulum, and their impact on electrical stability. We showed that Cx43 hemichannels are activated during diastolic Ca2+ release in single ventricular cardiomyocytes and cardiomyocyte cell pairs from mouse and pig. This activation involved Cx43 hemichannel Ca2+ entry and coupling to Ca2+ release microdomains at the intercalated disc resulting in enhanced Ca2+ dynamics. Hemichannel opening furthermore contributed to delayed afterdepolarizations and triggered action potentials. In single cardiomyocytes, cardiomyocyte cell pairs and arterially perfused tissue wedges from failing human hearts, increased hemichannel activity contributed to electrical instability as compared to non-failing rejected donor hearts. We conclude that microdomain coupling between Cx43 hemichannels and Ca2+ release is a novel, targetable, mechanism of cardiac arrhythmogenesis in heart failure.

Keywords:  Arrhythmias; Calcium signaling; Cardiology; Cell Biology; Peptides

DOI:  https://doi.org/10.1172/JCI137752
Front Cell Dev Biol. 2020 ;8 621509

Metformin-Enhanced Cardiac AMP-Activated Protein Kinase/Atrogin-1 Pathways Inhibit Charged Multivesicular Body Protein 2B Accumulation in Ischemia-Reperfusion Injury.

Tian Li, Yue Yin, Nan Mu, Yishi Wang, Manling Liu, Mai Chen, Wenhua Jiang, Lu Yu, Yan Li, Heng Ma.

  Background: Cardiac autophagic flux is impaired during myocardial ischemia/reperfusion (MI/R). Impaired autophagic flux may exacerbate MI/R injury. Charged multivesicular body protein 2B (CHMP2B) is a subunit of the endosomal sorting complex required for transport (ESCRT-III) complex that is required for autophagy. However, the reverse role of CHMP2B accumulation in autophagy and MI/R injury has not been established. The objective of this article is to elucidate the roles of AMP-activated protein kinase (AMPK)/atrogin-1 pathways in inhibiting CHMP2B accumulation in ischemia-reperfusion injury. Methods: Male C57BL/6 mice (3-4 months) and H9c2 cardiomyocytes were used to evaluate MI/R and hypoxia/reoxygenation (H/R) injury in vivo and in vitro, respectively. MI/R was built by a left lateral thoracotomy and occluded the left anterior descending artery. H9c2 cells were firstly treated in 95% N2 and 5% CO2 for 15 h and reoxygenation for 1 h. Metformin (100 mg/kg/d) and CHMP2B (Ad-CHMP2B) transfected adenoviruses were administered to the mice. The H9c2 cells were treated with metformin (2.5 mM), MG-132 (10 μM), bafilomycin A1 (10 nM), and compound C (20 μM). Results: Autophagic flux was found to be inhibited in H/R-treated cardiomyocytes and MI/R mice, with elevated cardiac CHMP2B accumulation. Upregulated CHMP2B levels in the in vivo and in vitro experiments were shown to inhibit autophagic flux leading to the deterioration of H/R-cardiomyocytes and MI/R injury. This finding implies that CHMP2B accumulation increases the risk of myocardial ischemia. Metformin suppressed CHMP2B accumulation and ameliorated H/R-induced autophagic dysfunction by activating AMPK. Activated AMPK upregulated the messenger RNA expression and protein levels of atrogin-1, a muscle-specific ubiquitin ligase, in the myocardium. Atrogin-1 significantly enhanced the interaction between atrogin-1 and CHMP2B, therefore, promoting CHMP2B degradation in the MI/R myocardium. Finally, this study revealed that metformin-inhibited CHMP2B accumulation induced autophagic impairment and ischemic susceptibility in vivo through the AMPK-regulated CHMP2B degradation by atrogin-1. Conclusion: Impaired CHMP2B clearance in vitro and in vivo inhibits autophagic flux and weakens the myocardial ischemic tolerance. Metformin treatment degrades CHMP2B through the AMPK-atrogin-1-dependent pathway to maintain the homeostasis of autophagic flux. This is a novel mechanism that enriches the understanding of cardioprotection.

Keywords:  AMPK; CHMP2B; atrogin-1; autophagic flux; metformin; myocardial ischemia/reperfusion

DOI:  https://doi.org/10.3389/fcell.2020.621509
Cardiovasc Diabetol. 2021 Feb 22. 20(1): 50

Activation of the cardiac non-neuronal cholinergic system prevents the development of diabetes-associated cardiovascular complications.

Eng Leng Saw, James T Pearson, Daryl O Schwenke, Pujika Emani Munasinghe, Hirotsugu Tsuchimochi, Shruti Rawal, Sean Coffey, Philip Davis, Richard Bunton, Isabelle Van Hout, Yuko Kai, Michael J A Williams, Yoshihiko Kakinuma, Martin Fronius, Rajesh Katare.

   BACKGROUND: Acetylcholine (ACh) plays a crucial role in the function of the heart. Recent evidence suggests that cardiomyocytes possess a non-neuronal cholinergic system (NNCS) that comprises of choline acetyltransferase (ChAT), choline transporter 1 (CHT1), vesicular acetylcholine transporter (VAChT), acetylcholinesterase (AChE) and type-2 muscarinic ACh receptors (M2AChR) to synthesize, release, degrade ACh as well as for ACh to transduce a signal. NNCS is linked to cardiac cell survival, angiogenesis and glucose metabolism. Impairment of these functions are hallmarks of diabetic heart disease (DHD). The role of the NNCS in DHD is unknown. The aim of this study was to examine the effect of diabetes on cardiac NNCS and determine if activation of cardiac NNCS is beneficial to the diabetic heart.
METHODS: Ventricular samples from type-2 diabetic humans and db/db mice were used to measure the expression pattern of NNCS components (ChAT, CHT1, VAChT, AChE and M2AChR) and glucose transporter-4 (GLUT-4) by western blot analysis. To determine the function of the cardiac NNCS in the diabetic heart, a db/db mouse model with cardiac-specific overexpression of ChAT gene was generated (db/db-ChAT-tg). Animals were followed up serially and samples collected at different time points for molecular and histological analysis of cardiac NNCS components and prosurvival and proangiogenic signaling pathways.
RESULTS: Immunoblot analysis revealed alterations in the components of cardiac NNCS and GLUT-4 in the type-2 diabetic human and db/db mouse hearts. Interestingly, the dysregulation of cardiac NNCS was followed by the downregulation of GLUT-4 in the db/db mouse heart. Db/db-ChAT-tg mice exhibited preserved cardiac and vascular function in comparison to db/db mice. The improved function was associated with increased cardiac ACh and glucose content, sustained angiogenesis and reduced fibrosis. These beneficial effects were associated with upregulation of the PI3K/Akt/HIF1α signaling pathway, and increased expression of its downstream targets-GLUT-4 and VEGF-A.
CONCLUSION: We provide the first evidence for dysregulation of the cardiac NNCS in DHD. Increased cardiac ACh is beneficial and a potential new therapeutic strategy to prevent or delay the development of DHD.

Keywords:  Acetylcholine; Angiogenesis; Diabetic heart disease; Glucose metabolism; Non-neuronal cholinergic system

DOI:  https://doi.org/10.1186/s12933-021-01231-8
Cell Biosci. 2021 Feb 26. 11(1): 44

Dapagliflozin attenuates hypoxia/reoxygenation-caused cardiac dysfunction and oxidative damage through modulation of AMPK.

Kun-Ling Tsai, Pei-Ling Hsieh, Wan-Ching Chou, Hui-Ching Cheng, Yu-Ting Huang, Shih-Hung Chan.

   BACKGROUND: Emerging evidence demonstrated dapagliflozin (DAPA), a sodium-glucose cotransporter 2 inhibitor, prevented various cardiovascular events. However, the detailed mechanisms underlying its cardioprotective properties remained largely unknown.
RESULTS: In the present study, we sought to investigate the effects of DAPA on the cardiac ischemia/reperfusion (I/R) injury. Results from in vitro experiments showed that DAPA induced the phosphorylation of AMPK, resulting in the downregulation of PKC in the cardiac myoblast H9c2 cells following hypoxia/reoxygenation (H/R) condition. We demonstrated that DAPA treatment diminished the H/R-elicited oxidative stress via the AMPK/ PKC/ NADPH oxidase pathway. In addition, DAPA prevented the H/R-induced abnormality of PGC-1α expression, mitochondrial membrane potential, and mitochondrial DNA copy number through AMPK/ PKC/ NADPH oxidase signaling. Besides, DAPA reversed the H/R-induced apoptosis. Furthermore, we demonstrated that DAPA improved the I/R-induced cardiac dysfunction by echocardiography and abrogated the I/R-elicited apoptosis in the myocardium of rats. Also, the administration of DAPA mitigated the production of myocardial infarction markers.
CONCLUSIONS: In conclusion, our data suggested that DAPA treatment holds the potential to ameliorate the I/R-elicited oxidative stress and the following cardiac apoptosis via modulation of AMPK, which attenuates the cardiac dysfunction caused by I/R injury.

Keywords:  AMPK; Dapagliflozin; Ischemia/reperfusion injury

DOI:  https://doi.org/10.1186/s13578-021-00547-y
J Exp Med. 2021 May 03. pii: e20201206. [Epub ahead of print]218(5):

Meteorin-like/Meteorin-β protects heart against cardiac dysfunction.

Celia Rupérez, Gemma Ferrer-Curriu, Aina Cervera-Barea, Laura Florit, Mariona Guitart-Mampel, Gloria Garrabou, Mònica Zamora, Fàtima Crispi, Joaquim Fernandez-Solà, Josep Lupón, Antoni Bayes-Genis, Francesc Villarroya, Anna Planavila.

Meteorin-like/Meteorin-β (Metrnl/Metrnβ) is a secreted protein produced by skeletal muscle and adipose tissue that exerts metabolic actions that improve glucose metabolism. The role of Metrnβ in cardiac disease is completely unknown. Here, we show that Metrnβ-null mice exhibit asymmetrical cardiac hypertrophy, fibrosis, and enhanced signs of cardiac dysfunction in response to isoproterenol-induced cardiac hypertrophy and aging. Conversely, adeno-associated virus-mediated specific overexpression of Metrnβ in the heart prevents the development of cardiac remodeling. Furthermore, Metrnβ inhibits cardiac hypertrophy development in cardiomyocytes in vitro, indicating a direct effect on cardiac cells. Antibody-mediated blockage of Metrnβ in cardiomyocyte cell cultures indicated an autocrine action of Metrnβ on the heart, in addition to an endocrine action. Moreover, Metrnβ is highly produced in the heart, and analysis of circulating Metrnβ concentrations in a large cohort of patients reveals that it is a new biomarker of heart failure with an independent prognostic value.

DOI: https://doi.org/10.1084/jem.20201206
Acta Biochim Biophys Sin (Shanghai). 2021 Feb 23. pii: gmab007. [Epub ahead of print]

Interleukin 35 protects cardiomyocytes following ischemia/reperfusion-induced apoptosis via activation of mitochondrial STAT3.

Fengyun Zhou, Ting Feng, Xiangqi Lu, Huicheng Wang, Yangping Chen, Qiuxia Zhang, Xinlu Zhang, Jiancheng Xiu.

  Mitochondrial reactive oxygen species (mtROS)-induced apoptosis has been suggested to contribute to myocardial ischemia/reperfusion injury. Interleukin 35 (IL-35), a novel anti-inflammatory cytokine, has been shown to protect the myocardium and inhibit mtROS production. However, its effect on cardiomyocytes upon exposure to hypoxia/reoxygenation (H/R) damage has not yet been elucidated. The present study aimed to investigate the potential protective role and underlying mechanisms of IL-35 in H/R-induced mouse neonatal cardiomyocyte injury. Mouse neonatal cardiomyocytes were challenged to H/R in the presence of IL-35, and we found that IL-35 dose dependently promotes cell viability, diminishes mtROS, maintains mitochondrial membrane potential, and decreases the number of apoptotic cardiomyocytes. Meanwhile, IL-35 remarkably activates mitochondrial STAT3 (mitoSTAT3) signaling, inhibits cytochrome c release, and reduces apoptosis signaling. Furthermore, co-treatment of the cardiomyocytes with the STAT3 inhibitor AG490 abrogates the IL-35-induced cardioprotective effects. Our study identified the protective role of IL-35 in cardiomyocytes following H/R damage and revealed that IL-35 protects cardiomyocytes against mtROS-induced apoptosis through the mitoSTAT3 signaling pathway during H/R.

Keywords:  IL-35; ROS; STAT3; apoptosis; hypoxia/reoxygenation

DOI:  https://doi.org/10.1093/abbs/gmab007
Cardiovasc Toxicol. 2021 Feb 21.

PHD Finger Protein 19 Promotes Cardiac Hypertrophy via Epigenetically Regulating SIRT2.

Wei Gu, Yutong Cheng, Su Wang, Tao Sun, Zhizhong Li.

  Epigenetic regulations essentially participate in the development of cardiomyocyte hypertrophy. PHD finger protein 19 (PHF19) is a polycomb protein that controls H3K36me3 and H3K27me3. However, the roles of PHF19 in cardiac hypertrophy remain unknown. Here in this work, we observed that PHF19 promoted cardiac hypertrophy via epigenetically targeting SIRT2. In angiotensin II (Ang II)-induced cardiomyocyte hypertrophy, adenovirus-mediated knockdown of Phf19 reduced the increase in cardiomyocyte size, repressed the expression of hypertrophic marker genes Anp and Bnp, as well as inhibited protein synthesis. By contrast, Phf19 overexpression promoted Ang II-induced cardiomyocyte hypertrophy in vitro. We also knocked down Phf19 expression in mouse hearts in vivo. The results demonstrated that Phf19 knockdown reduced Ang II-induced decline in cardiac fraction shortening and ejection fraction. Phf19 knockdown also inhibited Ang II-mediated increase in heart weight, reduced cardiomyocyte size, and repressed the expression of hypertrophic marker genes in mouse hearts. Further mechanism studies showed that PHF19 suppressed the expression of SIRT2, which contributed to the function of PHF19 during cardiomyocyte hypertrophy. PHF19 bound the promoter of SIRT2 and regulated the balance between H3K27me3 and H3K36me3 to repress the expression of SIRT2 in vitro and in vivo. In human hypertrophic hearts, the overexpression of PHF19 and downregulation of SIRT2 were observed. Of importance, PHF19 expression was positively correlated with hypertrophic marker genes ANP and BNP but negatively correlated with SIRT2 in human hypertrophic hearts. Therefore, our findings demonstrated that PHF19 promoted the development of cardiac hypertrophy via epigenetically regulating SIRT2.

Keywords:  Cardiac hypertrophy; Epigenetics; Histone methylation; PHF19; SIRT2

DOI:  https://doi.org/10.1007/s12012-021-09639-0
Cell Death Dis. 2021 Feb 26. 12(2): 216

High glucose induces Drp1-mediated mitochondrial fission via the Orai1 calcium channel to participate in diabetic cardiomyocyte hypertrophy.

Qing-Rui Wu, Dan-Lin Zheng, Pei-Ming Liu, Hui Yang, Lu-An Li, Su-Juan Kuang, Ying-Yu Lai, Fang Rao, Yu-Mei Xue, Ji-Jin Lin, Shuang-Xin Liu, Chun-Bo Chen, Chun-Yu Deng.

Mitochondrial dysfunction and impaired Ca2+ handling are involved in the development of diabetic cardiomyopathy (DCM). Dynamic relative protein 1 (Drp1) regulates mitochondrial fission by changing its level of phosphorylation, and the Orai1 (Ca2+ release-activated calcium channel protein 1) calcium channel is important for the increase in Ca2+ entry into cardiomyocytes. We aimed to explore the mechanism of Drp1 and Orai1 in cardiomyocyte hypertrophy caused by high glucose (HG). We found that Zucker diabetic fat rats induced by administration of a high-fat diet develop cardiac hypertrophy and impaired cardiac function, accompanied by the activation of mitochondrial dynamics and calcium handling pathway-related proteins. Moreover, HG induces cardiomyocyte hypertrophy, accompanied by abnormal mitochondrial morphology and function, and increased Orai1-mediated Ca2+ influx. Mechanistically, the Drp1 inhibitor mitochondrial division inhibitor 1 (Mdivi-1) prevents cardiomyocyte hypertrophy induced by HG by reducing phosphorylation of Drp1 at serine 616 (S616) and increasing phosphorylation at S637. Inhibition of Orai1 with single guide RNA (sgOrai1) or an inhibitor (BTP2) not only suppressed Drp1 activity and calmodulin-binding catalytic subunit A (CnA) and phosphorylated-extracellular signal-regulated kinase (p-ERK1/2) expression but also alleviated mitochondrial dysfunction and cardiomyocyte hypertrophy caused by HG. In addition, the CnA inhibitor cyclosporin A and p-ERK1/2 inhibitor U0126 improved HG-induced cardiomyocyte hypertrophy by promoting and inhibiting phosphorylation of Drp1 at S637 and S616, respectively. In summary, we identified Drp1 as a downstream target of Orai1-mediated Ca2+ entry, via activation by p-ERK1/2-mediated phosphorylation at S616 or CnA-mediated dephosphorylation at S637 in DCM. Thus, the Orai1-Drp1 axis is a novel target for treating DCM.

DOI: https://doi.org/10.1038/s41419-021-03502-4
Circulation. 2021 Feb 23.

Single Nuclei Sequencing Reveals Novel Insights into the Regulation of Cellular Signatures in Children with Dilated Cardiomyopathy.

Luka Nicin, Wesley T Abplanalp, Anne Schänzer, Anke Sprengel, David John, Hannah Mellentin, Lukas Tombor, Matthias Keuper, Evelyn Ullrich, Karin Klingel, Reinhard B Dettmeyer, Jedrzej Hoffmann, Hakan Akintuerk, Christian Jux, Dietmar Schranz, Andreas M Zeiher, Stefan Rupp, Stefanie Dimmeler.

  Background: Dilated cardiomyopathy (DCM) is a leading cause of death in children with heart failure. The outcome of pediatric heart failure treatment is inconsistent and large cohort studies are lacking. Progress may be achieved through personalized therapy that takes age- and disease-related pathophysiology, pathology and molecular fingerprints into account. We present snRNA-seq from pediatric DCM patients as the next step in identifying cellular signatures. Methods: We performed single nuclei RNA sequencing with heart tissues from six children with DCM with an age of 0.5, 0.75, 5, 6, 12 and 13 years. Unsupervised clustering of 18,211 nuclei led to the identification of 14 distinct clusters with 6 major cell types. Results: The number of nuclei in fibroblast clusters increased with age in DCM patients, a finding that was confirmed by histological analysis and was consistent with an age-related increase in cardiac fibrosis quantified by cardiac magnetic resonance imaging. Fibroblasts of DCM patients over 6 years of age showed a profoundly altered gene expression pattern with enrichment of genes encoding fibrillary collagens, modulation of proteoglycans, switch in thrombospondin isoforms and signatures of fibroblast activation. Additionally, a population of cardiomyocytes with a high pro-regenerative profile was identified in infant DCM patients, but was absent in > 6-year-old children. This cluster showed high expression of cell cycle activators such as cyclin D family members, increased glycolytic metabolism and antioxidative genes and alterations in ß-adrenergic signaling genes. Conclusions: Novel insights into the cellular transcriptomes of hearts from pediatric DCM patients provide remarkable age-dependent changes in the expression patterns of fibroblast and cardiomyocyte genes with less fibrotic but enriched pro-regenerative signatures in infants.

Keywords:  Pediatric dilated cardiomyopathy; cardiomyocyte proliferation; cellular heterogeneity; single nuclei RNA sequencing

DOI:  https://doi.org/10.1161/CIRCULATIONAHA.120.051391
Basic Res Cardiol. 2021 Feb 26. 116(1): 14

Functional analysis of a gene-edited mouse model to gain insights into the disease mechanisms of a titin missense variant.

He Jiang, Charlotte Hooper, Matthew Kelly, Violetta Steeples, Jillian N Simon, Julia Beglov, Amar J Azad, Lisa Leinhos, Pauline Bennett, Elisabeth Ehler, Jacinta I Kalisch-Smith, Duncan B Sparrow, Roman Fischer, Raphael Heilig, Henrik Isackson, Mehroz Ehsan, Giannino Patone, Norbert Huebner, Benjamin Davies, Hugh Watkins, Katja Gehmlich.

  Titin truncating variants are a well-established cause of cardiomyopathy; however, the role of titin missense variants is less well understood. Here we describe the generation of a mouse model to investigate the underlying disease mechanism of a previously reported titin A178D missense variant identified in a family with non-compaction and dilated cardiomyopathy. Heterozygous and homozygous mice carrying the titin A178D missense variant were characterised in vivo by echocardiography. Heterozygous mice had no detectable phenotype at any time point investigated (up to 1 year). By contrast, homozygous mice developed dilated cardiomyopathy from 3 months. Chronic adrenergic stimulation aggravated the phenotype. Targeted transcript profiling revealed induction of the foetal gene programme and hypertrophic signalling pathways in homozygous mice, and these were confirmed at the protein level. Unsupervised proteomics identified downregulation of telethonin and four-and-a-half LIM domain 2, as well as the upregulation of heat shock proteins and myeloid leukaemia factor 1. Loss of telethonin from the cardiac Z-disc was accompanied by proteasomal degradation; however, unfolded telethonin accumulated in the cytoplasm, leading to a proteo-toxic response in the mice.We show that the titin A178D missense variant is pathogenic in homozygous mice, resulting in cardiomyopathy. We also provide evidence of the disease mechanism: because the titin A178D variant abolishes binding of telethonin, this leads to its abnormal cytoplasmic accumulation. Subsequent degradation of telethonin by the proteasome results in proteasomal overload, and activation of a proteo-toxic response. The latter appears to be a driving factor for the cardiomyopathy observed in the mouse model.

Keywords:  Cardiomyopathy; Mouse model; Proteasome; Proteo-toxic response; Telethonin; Titin missense variant

DOI:  https://doi.org/10.1007/s00395-021-00853-z
Cardiovasc Toxicol. 2021 Feb 27.

MiR-200a-3p Aggravates DOX-Induced Cardiotoxicity by Targeting PEG3 Through SIRT1/NF-κB Signal Pathway.

Qinghua Fu, Hongwei Pan, Yi Tang, Jingjing Rong, Zhaofen Zheng.

  Doxorubicin (DOX) is a widely used cytotoxic drug whose application is limited by its severe side effects. Little was known regarding how to offset its side effects. Therefore this study aims to explore the role of miR-200a-3p in DOX-induced cardiotoxicity and its possible mechanism. DOX-induced myocardial injury rat models were established, which were then injected with miR-200a-3p inhibitor (miR-200a-3p suppression) to observe the effects of miR-200a-3p on cell proliferation, and apoptosis. Heart function and weights of rat models were also measured. Cardiomyocytes were induced by DOX, in which PEG3 knockdown or corresponding plasmids were transfected to assess the possible effect of PEG3 on cell activity. Dual luciferase reporter assay was applied to verify the binding of PEG3 with miR-200a-3p. Elevated levels of lactate dehydrogenase (LDH), creatine kinase-MB (CK-MB) and left ventricular end-diastolic pressure (LVEDP), as well as suppressed left ventricular systolic pressure (LVSP) and ± dp/dt max were showed in myocardial injury rat models. DOX induced myocardial injury and increased miR-200a-3p expression levels. miR-200a-3p inhibitor could partially attenuate DOX-induced cardiotoxicity in rat models, while PEG3 could regulate myocardial injury in DOX-treated cell models. miR-200a-3p, by targeting PEG3 through SIRT1/NF-κB signal pathway, regulated cell proliferation, inflammation and apoptosis of myocardiocytes. The results in current study demonstrated that miR-200a-3p regulates cell proliferation and apoptosis of cardiomyocytes by targeting PEG3 through SIRT1/NF-κB signal pathway. This result may provide a potential clue for the treatment of DOX-induced cardiotoxicity.

Keywords:  Apoptosis; Doxorubicin-induced myocardial injury; NF-κB; PEG3; Proliferation; SIRT1; miR-200a-3p

DOI:  https://doi.org/10.1007/s12012-020-09620-3
Nat Rev Cardiol. 2021 Feb 22.

Endoplasmic reticulum stress and unfolded protein response in cardiovascular diseases.

Jun Ren, Yaguang Bi, James R Sowers, Claudio Hetz, Yingmei Zhang.

Cardiovascular diseases (CVDs), such as ischaemic heart disease, cardiomyopathy, atherosclerosis, hypertension, stroke and heart failure, are among the leading causes of morbidity and mortality worldwide. Although specific CVDs and the associated cardiometabolic abnormalities have distinct pathophysiological and clinical manifestations, they often share common traits, including disruption of proteostasis resulting in accumulation of unfolded or misfolded proteins in the endoplasmic reticulum (ER). ER proteostasis is governed by the unfolded protein response (UPR), a signalling pathway that adjusts the protein-folding capacity of the cell to sustain the cell's secretory function. When the adaptive UPR fails to preserve ER homeostasis, a maladaptive or terminal UPR is engaged, leading to the disruption of ER integrity and to apoptosis. ER stress functions as a double-edged sword, with long-term ER stress resulting in cellular defects causing disturbed cardiovascular function. In this Review, we discuss the distinct roles of the UPR and ER stress response as both causes and consequences of CVD. We also summarize the latest advances in our understanding of the importance of the UPR and ER stress in the pathogenesis of CVD and discuss potential therapeutic strategies aimed at restoring ER proteostasis in CVDs.

DOI: https://doi.org/10.1038/s41569-021-00511-w
Cardiovasc Res. 2021 Feb 24. pii: cvab055. [Epub ahead of print]

Paclitaxel mitigates structural alterations and cardiac conduction system defects in a mouse model of Hutchinson-Gilford progeria syndrome.

Álvaro Macías, J Jaime Díaz-Larrosa, Yaazan Blanco, Víctor Fanjul, Cristina González-Gómez, Pilar Gonzalo, María Jesús Andrés-Manzano, Andre Monteiro da Rocha, Daniela Ponce-Balbuena, Andrew Allan, David Filgueiras-Rama, José Jalife, Vicente Andrés.

   AIMS: Hutchinson-Gilford progeria syndrome (HGPS) is an ultrarare laminopathy caused by expression of progerin, a lamin A variant, also present at low levels in non-HGPS individuals. HGPS patients age and die prematurely, predominantly from cardiovascular complications. Progerin-induced cardiac repolarization defects have been described previously, although the underlying mechanisms are unknown.
METHODS AND RESULTS: We conducted studies in heart tissue from progerin-expressing LmnaG609G/G609G (G609G) mice, including microscopy, intracellular calcium dynamics, patch-clamping, in vivo magnetic resonance imaging, and electrocardiography. Mouse G609G cardiomyocytes showed tubulin-cytoskeleton disorganization, t-tubular system disruption, sarcomere shortening, altered excitation-contraction coupling, and reductions in ventricular thickening and cardiac index. G609G mice exhibited severe bradycardia, and significant alterations of atrio-ventricular conduction and repolarization. Most importantly, 50% of G609G mice had altered heart rate variability, and sinoatrial block, both significant signs of premature cardiac aging. G609G cardiomyocytes had electrophysiological alterations which result in an elevated action potential plateau and early afterdepolarization bursting, reflecting slower sodium current inactivation and long Ca+2 transient duration, which may also help explain the mild QT prolongation in some HGPS patients. Chronic treatment with low-dose paclitaxel ameliorated structural and functional alterations in G609G hearts.
CONCLUSIONS: Our results demonstrate that tubulin-cytoskeleton disorganization in progerin-expressing cardiomyocytes causes structural, cardiac conduction and excitation-contraction coupling defects, all of which can be partially corrected by chronic treatment with low-dose of paclitaxel.

Keywords:  Hutchinson-Gilford progeria syndrome; animal model of cardiovascular disease; cardiomyocytes; electrophysiology; lamin A/C; progerin

DOI:  https://doi.org/10.1093/cvr/cvab055
Front Immunol. 2020 ;11 606045

Exosomes Derived From Hypertrophic Cardiomyocytes Induce Inflammation in Macrophages via miR-155 Mediated MAPK Pathway.

Hui Yu, Lei Qin, Yunzhi Peng, Wenhui Bai, Zhanli Wang.

  The inflammatory immune microenvironment plays an important role in the development of cardiac hypertrophy. Exosomes have emerged as the potent modulators of inflammatory responses. This study aimed to determine how exosomes derived from angiotensin II (Ang II)-induced hypertrophic cardiomyocytes (HCs) interfere with the inflammatory signal pathways in macrophages. Herein, we showed that increased exosome release was observed in HCs when compared to normal cardiomyocytes (NCs). Incubation of the murine macrophage cell line RAW264.7 in the presence of exosomes isolated from the culture media of HCs triggers the secretion of inflammatory cytokines interleukin (IL)-6 and IL-8. Cytokines release induced by HCs-derived exosomes was prevented by down-regulation of Argonaute2 (AGO2), suggesting that the non-coding RNAs were involved in exosome-induced inflammatory responses in RAW 264.7 macrophages. RNA sequencing assays further demonstrated that a total of seven microRNAs were differentially expressed between NCs-derived and HCs-derived exosomes. Importantly, miR-155 played a crucial role in the initiation of inflammation in macrophages. Further analyses demonstrated that HCs-derived exosomes induced the phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 via miR-155. Our results support the concept that exosomal microRNAs have emerged as important inflammatory response modulators regulating cardiac hypertrophy.

Keywords:  cardiac hypertrophy; exosome; inflammation; macrophage; microRNA

DOI:  https://doi.org/10.3389/fimmu.2020.606045
Dis Model Mech. 2021 Feb 22. pii: dmm047704. [Epub ahead of print]14(2):

Cardiovascular phenotype of the Dmdmdx rat - a suitable animal model for Duchenne muscular dystrophy.

Petra Lujza Szabó, Janine Ebner, Xaver Koenig, Ouafa Hamza, Simon Watzinger, Sandra Trojanek, Dietmar Abraham, Hannes Todt, Helmut Kubista, Klaus Schicker, Séverine Remy, Ignacio Anegon, Attila Kiss, Bruno K Podesser, Karlheinz Hilber.

  Besides skeletal muscle abnormalities, Duchenne muscular dystrophy (DMD) patients present with dilated cardiomyopathy development, which considerably contributes to morbidity and mortality. Because the mechanisms responsible for the cardiac complications in the context of DMD are largely unknown, evidence-based therapy approaches are still lacking. This has increased the need for basic research efforts into animal models for DMD. Here, we characterized in detail the cardiovascular abnormalities of Dmdmdx rats, with the aim of determining the suitability of this recently established dystrophin-deficient small animal as a model for DMD.Various methods were applied to compare cardiovascular properties between wild-type and Dmdmdx rats, and to characterize the Dmdmdx cardiomyopathy. These methods comprised echocardiography, invasive assessment of left ventricular hemodynamics, examination of adverse remodeling and endothelial cell inflammation, and evaluation of vascular function, employing wire myography. Finally, intracellular Ca2+ transient measurements, and recordings of currents through L-type Ca2+ channels were performed in isolated single ventricular cardiomyocytes. We found that, similar to respective observations in DMD patients, the hearts of Dmdmdx rats show significantly impaired cardiac function, fibrosis and inflammation, consistent with the development of a dilated cardiomyopathy. Moreover, in Dmdmdx rats, vascular endothelial function is impaired, which may relate to inflammation and oxidative stress, and Ca2+ handling in Dmdmdx cardiomyocytes is abnormal.These findings indicate that Dmdmdx rats represent a promising small-animal model to elucidate mechanisms of cardiomyopathy development in the dystrophic heart, and to test mechanism-based therapies aiming to combat cardiovascular complications in DMD.

Keywords:  Cardiomyocyte; Cardiovascular dysfunction; Muscular dystrophy; Rat; Remodeling

DOI:  https://doi.org/10.1242/dmm.047704
Front Cell Dev Biol. 2021 ;9 624601

Consistent Long-Term Therapeutic Efficacy of Human Umbilical Cord Matrix-Derived Mesenchymal Stromal Cells After Myocardial Infarction Despite Individual Differences and Transient Engraftment.

Tiago L Laundos, Francisco Vasques-Nóvoa, Rita N Gomes, Vasco Sampaio-Pinto, Pedro Cruz, Hélder Cruz, Jorge M Santos, Rita N Barcia, Perpétua Pinto-do-Ó, Diana S Nascimento.

  Human mesenchymal stem cells gather special interest as a universal and feasible add-on therapy for myocardial infarction (MI). In particular, human umbilical cord matrix-derived mesenchymal stromal cells (UCM-MSC) are advantageous since can be easily obtained and display high expansion potential. Using isolation protocols compliant with cell therapy, we previously showed UCM-MSC preserved cardiac function and attenuated remodeling 2 weeks after MI. In this study, UCM-MSC from two umbilical cords, UC-A and UC-B, were transplanted in a murine MI model to investigate consistency and durability of the therapeutic benefits. Both cellular products improved cardiac function and limited adverse cardiac remodeling 12 weeks post-ischemic injury, supporting sustained and long-term beneficial therapeutic effect. Donor associated variability was found in the modulation of cardiac remodeling and activation of the Akt-mTOR-GSK3β survival pathway. In vitro, the two cell products displayed similar ability to induce the formation of vessel-like structures and comparable transcriptome in normoxia and hypoxia, apart from UCM-MSCs proliferation and expression differences in a small subset of genes associated with MHC Class I. These findings support that UCM-MSC are strong candidates to assist the treatment of MI whilst calling for the discussion on methodologies to characterize and select best performing UCM-MSC before clinical application.

Keywords:  Wharton's jelly; cardiac fibrosis; cell therapy; donor variability; mesenchymal stromal (or stem) cells; myocardial infarction; regeneration/repair; umbilical cord matrix derived mesenchymal stromal cells (hUCM-MSCs)

DOI:  https://doi.org/10.3389/fcell.2021.624601
J Mol Cell Cardiol. 2021 Feb 23. pii: S0022-2828(21)00042-0. [Epub ahead of print]

Global identification of S-palmitoylated proteins and detection of palmitoylating (DHHC) enzymes in heart.

Madeleine R Miles, John Seo, Min Jiang, Zachary T Wilson, Janay Little, Jon Hao, Joshua Andrade, Beatrix Ueberheide, Gea-Ny Tseng.

  High-throughput experiments suggest that almost 20% of human proteins may be S-palmitoylatable, a post-translational modification (PTM) whereby fatty acyl chains, most commonly palmitoyl chain, are linked to cysteine thiol groups that impact on protein trafficking, distribution and function. In human, protein S-palmitoylation is mediated by a group of 23 palmitoylating 'Asp-His-His-Cys' domain-containing (DHHC) enzymes. There is no information on the scope of protein S-palmitoylation, or the pattern of DHHC enzyme expression, in the heart. We used resin-assisted capture to pull down S-palmitoylated proteins from human, dog, and rat hearts, followed by proteomic search to identify proteins in the pulldowns. We identified 454 proteins present in at least 2 species-specific pulldowns. These proteins are operationally called 'cardiac palmitoylome'. Enrichment analysis based on Gene Ontology terms 'cellular component' indicated that cardiac palmitoylome is involved in cell-cell and cell-substrate junctions, plasma membrane microdomain organization, vesicular trafficking, and mitochondrial enzyme organization. Importantly, cardiac palmitoylome is uniquely enriched in proteins participating in the organization and function of t-tubules, costameres and intercalated discs, three microdomains critical for excitation-contraction coupling and intercellular communication of cardiomyocytes. We validated antibodies targeting DHHC enzymes, and detected eleven of them expressed in hearts across species. In conclusion, we provide resources useful for investigators interested in studying protein S-palmitoylation and its regulation by DHHC enzymes in the heart. We also discuss challenges in these efforts, and suggest methods and tools that should be developed to overcome these challenges.

Keywords:  Cardiac disease; Palmitoylation; Protein palmitoyl transferase; Proteomics

DOI:  https://doi.org/10.1016/j.yjmcc.2021.02.007
Sci Rep. 2021 Feb 25. 11(1): 4695

CircSLC8A1 and circNFIX can be used as auxiliary diagnostic markers for sudden cardiac death caused by acute ischemic heart disease.

Meihui Tian, Jiajia Xue, Cuiyun Dai, Enzhu Jiang, Baoli Zhu, Hao Pang.

Sudden cardiac death (SCD) caused by acute ischemic heart disease (IHD) is a major cause of sudden death worldwide. Circular RNAs (circRNAs) are abundant in the heart and play important roles in cardiovascular diseases, but the role of circRNAs as biomarkers in the forensic diagnosis of SCD caused by acute IHD remains poorly characterized. To investigate the potential of two heart-enriched circRNAs, circNFIX and circSLC8A1, we explored the expression of these two circRNAs in different kinds of commonly used IHD models, and further verified their expressions in forensic autopsy cases. The results from both the IHD rat and H9c2 cell models revealed that circSlc8a1 level was upregulated, while the circNfix level was elevated in the early stage of ischemia and subsequently downregulated. The time-dependent expression patterns of the two circRNAs suggested their potential as SCD biomarkers. In autopsy cases, the results showed that the expression of these two circRNAs in the myocardium with acute IHD-related SCDs corresponded to the observations in the ischemic models. Further analysis related to myocardial ischemia indicated that circSLC8A1 showed high sensitivity and specificity for myocardial infarction and was positively correlated with creatine kinase MB in pericardial fluid. Downregulated circNFIX level could indicate the ischemic myocardial damage, and it was negatively correlated with the coronary artery stenosis grade. The combination of circSLC8A1 and circNFIX had better performance to discriminate IHD-related SCDs. The results suggested that circSLC8A1 and circNFIX may be used as auxiliary diagnostic markers for SCD caused by acute IHD in forensic medicine.

DOI: https://doi.org/10.1038/s41598-021-84056-5
J Mol Cell Cardiol. 2021 Feb 22. pii: S0022-2828(21)00035-3. [Epub ahead of print]

Mapping genetic changes in the cAMP-signaling cascade in human atria.

Anne Garnier, Nadja I Bork, Eric Jacquet, Svante Zipfel, Christian Muñoz-Guijosa, Istvan Baczkó, Hermann Reichenspurner, Patrick Donzeau-Gouge, Lars S Maier, Dobromir Dobrev, Evaldas Girdauskas, Viacheslav O Nikolaev, Rodolphe Fischmeister, Cristina E Molina.

   AIM: To obtain a quantitative expression profile of the main genes involved in the cAMP-signaling cascade in human control atria and in different cardiac pathologies.
METHODS AND RESULTS: Expression of 48 target genes playing a relevant role in the cAMP-signaling cascade was assessed by RT-qPCR. 113 samples were obtained from right atrial appendages (RAA) of patients in sinus rhythm (SR) with or without atrium dilation, paroxysmal atrial fibrillation (AF), persistent AF or heart failure (HF); and left atrial appendages (LAA) from patients in SR or with AF. Our results show that right and left atrial appendages in donor hearts or from SR patients have similar expression values except for AC7 and PDE2A. Despite the enormous chamber-dependent variability in the gene-expression changes between pathologies, several distinguishable patterns could be identified. PDE8A, PKI3G and EPAC2 were upregulated in AF. Different phosphodiesterase (PDE) families showed specific pathology-dependent changes.
CONCLUSION: By comparing mRNA-expression patterns of the cAMP-signaling cascade related genes in right and left atrial appendages of human hearts and across different pathologies, we show that 1) gene expression is not significantly affected by cardioplegic solution content, 2) it is appropriate to use SR atrial samples as controls, and 3) many genes in the cAMP-signaling cascade are affected in AF and HF but only few of them appear to be chamber (right or left) specific.
TOPIC: AC, AKAP, PDE, Epac, PKA, CaMKII TRANSLATIONAL PERSPECTIVE: The cyclic AMP signaling pathway is important for atrial function. However, expression patterns of the genes involved in the atria of healthy and diseased hearts are still unclear. We give here a general overview of how different pathologies affect the expression of key genes in the cAMP signaling pathway in human right and left atria appendages. Our study may help identifying new genes of interest as potential therapeutic targets or clinical biomarkers for these pathologies and could serve as a guide in future gene therapy studies.

Keywords:  Atrial fibrillation; Heart failure; Left atria; Right atria; cAMP-signaling cascade; β-Adrenergic pathway

DOI:  https://doi.org/10.1016/j.yjmcc.2021.02.006
Stem Cell Reports. 2021 Feb 12. pii: S2213-6711(21)00048-5. [Epub ahead of print]

Maturation of Pluripotent Stem Cell-Derived Cardiomyocytes Enables Modeling of Human Hypertrophic Cardiomyopathy.

Walter E Knight, Yingqiong Cao, Ying-Hsi Lin, Congwu Chi, Betty Bai, Genevieve C Sparagna, Yuanbiao Zhao, Yanmei Du, Pilar Londono, Julie A Reisz, Benjamin C Brown, Matthew R G Taylor, Amrut V Ambardekar, Joseph C Cleveland, Timothy A McKinsey, Mark Y Jeong, Lori A Walker, Kathleen C Woulfe, Angelo D'Alessandro, Kathryn C Chatfield, Hongyan Xu, Michael R Bristow, Peter M Buttrick, Kunhua Song.

  Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are a powerful platform for biomedical research. However, they are immature, which is a barrier to modeling adult-onset cardiovascular disease. Here, we sought to develop a simple method that could drive cultured hiPSC-CMs toward maturity across a number of phenotypes, with the aim of utilizing mature hiPSC-CMs to model human cardiovascular disease. hiPSC-CMs were cultured in fatty acid-based medium and plated on micropatterned surfaces. These cells display many characteristics of adult human cardiomyocytes, including elongated cell morphology, sarcomeric maturity, and increased myofibril contractile force. In addition, mature hiPSC-CMs develop pathological hypertrophy, with associated myofibril relaxation defects, in response to either a pro-hypertrophic agent or genetic mutations. The more mature hiPSC-CMs produced by these methods could serve as a useful in vitro platform for characterizing cardiovascular disease.

Keywords:  HIF1α; cardiomyocyte maturation; disease modeling; hiPSC-CM; hiPSC-CM maturation; hypertrophic cardiomyopathy

DOI:  https://doi.org/10.1016/j.stemcr.2021.01.018
Nat Protoc. 2021 Feb 24.

Use of stable isotope-tagged thymidine and multi-isotope imaging mass spectrometry (MIMS) for quantification of human cardiomyocyte division.

Jessie W Yester, Honghai Liu, Frank Gyngard, Niyatie Ammanamanchi, Kathryn C Little, Dawn Thomas, Mara L G Sullivan, Sean Lal, Matthew L Steinhauser, Bernhard Kühn.

Quantification of cellular proliferation in humans is important for understanding biology and responses to injury and disease. However, existing methods require administration of tracers that cannot be ethically administered in humans. We present a protocol for the direct quantification of cellular proliferation in human hearts. The protocol involves administration of non-radioactive, non-toxic stable isotope 15Nitrogen-enriched thymidine (15N-thymidine), which is incorporated into DNA during S-phase, in infants with tetralogy of Fallot, a common form of congenital heart disease. Infants with tetralogy of Fallot undergo surgical repair, which requires the removal of pieces of myocardium that would otherwise be discarded. This protocol allows for the quantification of cardiomyocyte proliferation in this discarded tissue. We quantitatively analyzed the incorporation of 15N-thymidine with multi-isotope imaging spectrometry (MIMS) at a sub-nuclear resolution, which we combined with correlative confocal microscopy to quantify formation of binucleated cardiomyocytes and cardiomyocytes with polyploid nuclei. The entire protocol spans 3-8 months, which is dependent on the timing of surgical repair, and 3-4.5 researcher days. This protocol could be adapted to study cellular proliferation in a variety of human tissues.

DOI: https://doi.org/10.1038/s41596-020-00477-y
Cardiovasc Res. 2021 Feb 22. pii: cvab004. [Epub ahead of print]

Asymmetric Hapln1a drives regionalised cardiac ECM expansion and promotes heart morphogenesis in zebrafish development.

Christopher J Derrick, Juliana Sánchez-Posada, Farah Hussein, Federico Tessadori, Eric J G Pollitt, Aaron M Savage, Robert N Wilkinson, Timothy J Chico, Fredericus J van Eeden, Jeroen Bakkers, Emily S Noël.

AIMS: Vertebrate heart development requires the complex morphogenesis of a linear tube to form the mature organ, a process essential for correct cardiac form and function requiring coordination of embryonic laterality, cardiac growth, and regionalised cellular changes. While previous studies have demonstrated broad requirements for extracellular matrix (ECM) components in cardiac morphogenesis, we hypothesised that ECM regionalisation may fine tune cardiac shape during heart development.
METHODS AND RESULTS: Using live in vivo light sheet imaging of zebrafish embryos we describe a left-sided expansion of the ECM between the myocardium and endocardium prior to the onset of heart looping and chamber ballooning. Analysis using an ECM sensor revealed the cardiac ECM is further regionalised along the atrioventricular axis. Spatial transcriptomic analysis of gene expression in the heart tube identified candidate genes that may drive ECM expansion. This approach identified regionalised expression of hapln1a, encoding an ECM cross-linking protein. Validation of transcriptomic data by in situ hybridisation confirmed regionalised hapln1a expression in the heart, with highest levels of expression in the future atrium and on the left side of the tube, overlapping with the observed ECM expansion. Analysis of CRISPR-Cas9-generated hapln1a mutants revealed a reduction in atrial size and reduced chamber ballooning. Loss-of-function analysis demonstrated that ECM expansion is dependent upon Hapln1a, together supporting a role for Hapln1a in regionalised ECM modulation and cardiac morphogenesis. Analysis of hapln1a expression in zebrafish mutants with randomised or absent embryonic left-right asymmetry revealed that laterality cues position hapln1a-expressing cells asymmetrically in the left side of the heart tube.
CONCLUSIONS: We identify a regionalised ECM expansion in the heart tube which promotes correct heart development, and propose a novel model whereby embryonic laterality cues orient the axis of ECM asymmetry in the heart, suggesting these two pathways interact to promote robust cardiac morphogenesis.
TRANSLATIONAL PERSPECTIVE: This study reveals that the cardiac ECM exhibits regional specialisation required for heart morphogenesis, and sheds light on how embryonic left-right asymmetry acts in concert with ECM regionalisation to fine tune heart shape. This work can help us understand the origins of congenital heart defects, and in particular the nature of morphological heart abnormalities in patients with heterotaxia-associated heart malformations. Furthermore, recent studies suggest the ECM is a key regulator of regenerative potential in the heart, thus defining how distinct ECM composition impacts upon heart form and function has implications for developing regenerative therapies in the future.

DOI: https://doi.org/10.1093/cvr/cvab004
Eur J Pharmacol. 2021 Feb 19. pii: S0014-2999(21)00108-4. [Epub ahead of print] 173955

Effect of doxorubicin on cardiac lipid metabolism-related transcriptome and the protective activity of Alda-1.

Leonardo da Cunha Menezes Souza, Fábio Henrique Fernandes, Paula Torres Presti, Ana Lucia Anjos Ferreira, Daisy Maria Fávero Salvadori.

  The use of doxorubicin (DOX) as an antineoplastic drug is compromised by its cardiotoxicity risk. Although several mechanisms have been proposed for DOX-induced cardiac dysfunction, there is still increased interest in assessing its effects. Likewise, it is important to find protocols that can prevent or minimize the side effects of DOX without hindering its antitumor activity. Thus, this study was designed to investigate the molecular mechanisms underlying DOX cardiotoxicity, with a special focus on cardiac energy metabolism and the ability of Alda-1 (ALDH2 agonist) to prevent DOX-induced cardiac alterations. We explored the effects of DOX on the histological morphology of the myocardium, on lipid profile, and on the expression of genes related to fatty acid metabolism, in the presence and absence of Alda-1 (8 mg/kg body weight; b.wt.). Two DOX treatment protocols were used: a single dose of DOX (4 mg/kg b.wt.); four doses of DOX (4 mg/kg b.wt.), one dose/week, for 4 weeks. Treatment with DOX caused a progressive injury in the cardiac tissue and an increase in the blood total cholesterol, high-density lipoproteins, very low-density lipoproteins and triglyceride, as well as an up-regulation of FABP4 (DOX and DOX + Alda-1 groups) and Slc27a2 (in DOX-treated animals). Alda-1 administration promoted reduction in the severity of the histopathological injuries (after single dose of DOX) and Slc27a2 overexpression was restored. In conclusion, the study revealed novel insights regarding the development of DOX-mediated cardiomyopathy, indicating a relationship between DOX exposure and FABP4 and Slc27a2 overexpression, and confirmed the cardioprotective effect of Alda-1.

Keywords:  ALDH2 activator; Cardioprotection; Cardiotoxicity; Fatty acid binding protein

DOI:  https://doi.org/10.1016/j.ejphar.2021.173955
Transplantation. 2021 Mar 01. 105(3): 496-508

Peritransplant Cardiometabolic and Mitochondrial Function: The Missing Piece in Donor Heart Dysfunction and Graft Failure.

Matthew A Wells, Louise E See Hoe, Lisa C Heather, Peter Molenaar, Jacky Y Suen, Jason Peart, David McGiffin, John F Fraser.

Primary graft dysfunction is an important cause of morbidity and mortality after cardiac transplantation. Donor brain stem death (BSD) is a significant contributor to donor heart dysfunction and primary graft dysfunction. There remain substantial gaps in the mechanistic understanding of peritransplant cardiac dysfunction. One of these gaps is cardiac metabolism and metabolic function. The healthy heart is an "omnivore," capable of utilizing multiple sources of nutrients to fuel its enormous energetic demand. When this fails, metabolic inflexibility leads to myocardial dysfunction. Data have hinted at metabolic disturbance in the BSD donor and subsequent heart transplantation; however, there is limited evidence demonstrating specific metabolic or mitochondrial dysfunction. This review will examine the literature surrounding cardiometabolic and mitochondrial function in the BSD donor, organ preservation, and subsequent cardiac transplantation. A more comprehensive understanding of this subject may then help to identify important cardioprotective strategies to improve the number and quality of donor hearts.

DOI: https://doi.org/10.1097/TP.0000000000003368
Basic Res Cardiol. 2021 Feb 25. 116(1): 13

Epigenetic regulation of cardiac electrophysiology in atrial fibrillation: HDAC2 determines action potential duration and suppresses NRSF in cardiomyocytes.

Patrick Lugenbiel, Katharina Govorov, Pascal Syren, Ann-Kathrin Rahm, Teresa Wieder, Maximilian Wunsch, Nadine Weiberg, Emili Manolova, Dominik Gramlich, Rasmus Rivinius, Daniel Finke, Lorenz H Lehmann, Patrick A Schweizer, Derk Frank, Fadwa A El Tahry, Claus Bruehl, Tanja Heimberger, Steffi Sandke, Tanja Weis, Patrick Most, Bastian Schmack, Arjang Ruhparwar, Matthias Karck, Norbert Frey, Hugo A Katus, Dierk Thomas.

  Atrial fibrillation (AF) is associated with electrical remodeling, leading to cellular electrophysiological dysfunction and arrhythmia perpetuation. Emerging evidence suggests a key role for epigenetic mechanisms in the regulation of ion channel expression. Histone deacetylases (HDACs) control gene expression through deacetylation of histone proteins. We hypothesized that class I HDACs in complex with neuron-restrictive silencer factor (NRSF) determine atrial K+ channel expression. AF was characterized by reduced atrial HDAC2 mRNA levels and upregulation of NRSF in humans and in a pig model, with regional differences between right and left atrium. In vitro studies revealed inverse regulation of Hdac2 and Nrsf in HL-1 atrial myocytes. A direct association of HDAC2 with active regulatory elements of cardiac K+ channels was revealed by chromatin immunoprecipitation. Specific knock-down of Hdac2 and Nrsf induced alterations of K+ channel expression. Hdac2 knock-down resulted in prolongation of action potential duration (APD) in neonatal rat cardiomyocytes, whereas inactivation of Nrsf induced APD shortening. Potential AF-related triggers were recapitulated by experimental tachypacing and mechanical stretch, respectively, and exerted differential effects on the expression of class I HDACs and K+ channels in cardiomyocytes. In conclusion, HDAC2 and NRSF contribute to AF-associated remodeling of APD and K+ channel expression in cardiomyocytes via direct interaction with regulatory chromatin regions. Specific modulation of these factors may provide a starting point for the development of more individualized treatment options for atrial fibrillation.

Keywords:  Atrial fibrillation; Electrophysiology; Epigenetics; Histone deacetylase; K+ channel

DOI:  https://doi.org/10.1007/s00395-021-00855-x
Int Immunopharmacol. 2021 Feb 23. pii: S1567-5769(21)00123-5. [Epub ahead of print]94 107487

Cateslytin abrogates lipopolysaccharide-induced cardiomyocyte injury by reducing inflammation and oxidative stress through toll like receptor 4 interaction.

Carmine Rocca, Anna De Bartolo, Fedora Grande, Bruno Rizzuti, Teresa Pasqua, Francesca Giordano, Maria Concetta Granieri, Maria Antonietta Occhiuzzi, Antonio Garofalo, Nicola Amodio, Maria Carmela Cerra, Francis Schneider, Maria Luisa Panno, Marie Hélène Metz-Boutigue, Tommaso Angelone.

  Global public health is threatened by new pathogens, antimicrobial resistant microorganisms and a rapid decline of conventional antimicrobials efficacy. Thus, numerous medical procedures become life-threating. Sepsis can lead to tissue damage such as myocardium inflammation, associated with reduction of contractility and diastolic dysfunction, which may cause death. In this perspective, growing interest and attention are paid on host defence peptides considered as new potential antimicrobials. In the present study, we investigated the physiological and biochemical properties of Cateslytin (Ctl), an endogenous antimicrobial chromogranin A-derived peptide, in H9c2 cardiomyocytes exposed to lipopolysaccharide (LPS) infection. We showed that both Ctl (L and D) enantiomers, but not their scrambled counterparts, significantly increased cardiomyocytes viability following LPS, even if L-Ctl was effective at lower concentration (1 nM) compared to D-Ctl (10 nM). L-Ctl mitigated LPS-induced LDH release and oxidative stress, as visible by a reduction of MDA and protein carbonyl groups content, and by an increase of SOD activity. Molecular docking simulations strongly suggested that L-Ctl modulates TLR4 through a direct binding to the partner protein MD-2. Molecular analyses indicated that the protection mediated by L-Ctl against LPS-evoked sepsis targeted the TLR4/ERK/JNK/p38-MAPK pathway, regulating NFkB p65, NFkB p52 and COX2 expression and repressing the mRNA expression levels of the LPS-induced proinflammatory factors IL-1β, IL-6, TNF-α and NOS2. These findings indicate that Ctl could be considered as a possible candidate for the development of new antimicrobials strategies in the treatment of myocarditis. Interestingly, L-enantiomeric Ctl showed remarkable properties in strengthening the anti-inflammatory and anti-oxidant effects on cardiomyocytes.

Keywords:  Antimicrobial peptides; Cardiomyocytes; Cateslytin; Inflammation; Oxidative stress; Toll-like receptor 4

DOI:  https://doi.org/10.1016/j.intimp.2021.107487
Sci Rep. 2021 Feb 22. 11(1): 4297

Predictive values of multiple non-invasive markers for myocardial fibrosis in hypertrophic cardiomyopathy patients with preserved ejection fraction.

Yumin Li, Jia Liu, Yukun Cao, Xiaoyu Han, Guozhu Shao, Xiaoyue Zhou, Jin Gu, Tong Liu, Yue Cui, Heshui Shi.

Myocardial fibrosis assessed by late gadolinium enhancement (LGE) on cardiovascular magnetic resonance (CMR) is associated with cardiovascular outcomes in hypertrophic cardiomyopathy (HCM) patients, but little is known about the utility of non-invasive markers for detecting LGE. This study aims to explore the association between cardiac-specific biomarkers, CMR myocardial strain, left ventricular (LV) hypertrophy and LGE in HCM patients with preserved ejection fraction (EF) and investigate the predictive values of these indexes for LGE. We recruited 33 healthy volunteers and 86 HCM patients with preserved EF to undergo contrast-enhanced CMR examinations. In total, 48 of 86 HCM patients had the presence of LGE. The LGE-positive patients had significant higher serum high-sensitivity cardiac troponin I (hs-cTnI) and N-terminal pro b-type natriuretic peptide (Nt-proBNP) levels and lower global longitudinal (GLS) and circumferential (GCS) strains than the LGE-negative group. The LGE% was independently associated with the Nt-proBNP levels, GCS, LV end-diastolic maximum wall thickness (MWT) and beta-blocker treatment. In the receiver operating characteristic curve analysis, the combined parameters of Nt-proBNP ≥ 108.00 pg/mL and MWT ≥ 17.30 mm had good diagnostic performance for LGE, with a specificity of 81.25% and sensitivity of 70.00%. These data indicate that serum Nt-proBNP is a potential biomarker associated with LGE% and, combined with MWT, were useful for identifying myocardial fibrosis in HCM patients with preserved EF. Additionally, LV GCS may be a more sensitive indicator for reflecting the presence of myocardial fibrosis than GLS.

DOI: https://doi.org/10.1038/s41598-021-83678-z
Front Pharmacol. 2020 ;11 600266

Deletion of Orphan G Protein-Coupled Receptor GPR37L1 in Mice Alters Cardiovascular Homeostasis in a Sex-Specific Manner.

Margaret A Mouat, Kristy L Jackson, James L J Coleman, Madeleine R Paterson, Robert M Graham, Geoffrey A Head, Nicola J Smith.

  GPR37L1 is a family A orphan G protein-coupled receptor (GPCR) with a putative role in blood pressure regulation and cardioprotection. In mice, genetic ablation of Gpr37l1 causes sex-dependent effects; female mice lacking Gpr37l1 (GPR37L1-/-) have a modest but significant elevation in blood pressure, while male GPR37L1-/- mice are more susceptible to cardiovascular dysfunction following angiotensin II-induced hypertension. Given that this receptor is highly expressed in the brain, we hypothesize that the cardiovascular phenotype of GPR37L1-/- mice is due to changes in autonomic regulation of blood pressure and heart rate. To investigate this, radiotelemetry was employed to characterize baseline cardiovascular variables in GPR37L1-/- mice of both sexes compared to wildtype controls, followed by power spectral analysis to quantify short-term fluctuations in blood pressure and heart rate attributable to alterations in autonomic homeostatic mechanisms. Additionally, pharmacological ganglionic blockade was performed to determine vasomotor tone, and environmental stress tests were used to assess whether cardiovascular reactivity was altered in GPR37L1-/- mice. We observed that mean arterial pressure was significantly lower in female GPR37L1-/- mice compared to wildtype counterparts, but was unchanged in male GPR37L1-/- mice. GPR37L1-/- genotype had a statistically significant positive chronotropic effect on heart rate across both sexes when analyzed by two-way ANOVA. Power spectral analysis of these data revealed a reduction in power in the heart rate spectrum between 0.5 and 3 Hz in female GPR37L1-/- mice during the diurnal active period, which indicates that GPR37L1-/- mice may have impaired cardiac vagal drive. GPR37L1-/- mice of both sexes also exhibited attenuated depressor responses to ganglionic blockade with pentolinium, indicating that GPR37L1 is involved in maintaining sympathetic vasomotor tone. Interestingly, when these mice were subjected to aversive and appetitive behavioral stressors, the female GPR37L1-/- mice exhibited an attenuation of cardiovascular reactivity to aversive, but not appetitive, environmental stimuli. Together, these results suggest that loss of GPR37L1 affects autonomic maintenance of blood pressure, giving rise to sex-specific cardiovascular changes in GPR37L1-/- mice.

Keywords:  G protein-coupled receptor; blood pressure; heart rate variability; hypertension; radiotelemetry; sex differences

DOI:  https://doi.org/10.3389/fphar.2020.600266
Eur J Pharmacol. 2021 Feb 18. pii: S0014-2999(21)00115-1. [Epub ahead of print] 173962

Dexpramipexole attenuates myocardial ischemia/reperfusion injury through upregulation of mitophagy.

Lu Tang, Yun-Peng Li, Juan Hu, Ai-Hua Chen, Yingli Mo.

  Reperfusion causes undesirable damage to the ischemic myocardium while restoring the blood flow. In this study, we evaluated the effects of dexpramipexole (DPX) on myocardial injury induced by ischemia/reperfusion (I/R) in-vivo and the hypoxia/reoxygenation (HR) in-vitro and examined the functional mechanisms of DPX. DPX protected cells against H/R-induced mitochondrial dysfunction and prevented H/R damage. Both myocardial infarct size and tissue damage due to I/R was reduced upon DPX treatment. We discovered that DPX enhanced mitophagy in-vivo and in-vitro, which was accompanied by enhanced expression of PINK1 and Parkin. Knock-down of PINK1 and Parkin by specific siRNAs reversed DPX-induced inhibition of myocardial I/R injury. These findings suggest that DPX might protect against myocardial injury via PINK1 and Parkin.

Keywords:  dexpramipexole; ischemia; mitophagy; myocardial; reperfusion

DOI:  https://doi.org/10.1016/j.ejphar.2021.173962
Sci Rep. 2021 Feb 24. 11(1): 4430

Optogenetic current in myofibroblasts acutely alters electrophysiology and conduction of co-cultured cardiomyocytes.

Geran M Kostecki, Yu Shi, Christopher S Chen, Daniel H Reich, Emilia Entcheva, Leslie Tung.

Interactions between cardiac myofibroblasts and myocytes may slow conduction and generate spontaneous beating in fibrosis, increasing the chance of life-threatening arrhythmia. While co-culture studies have shown that myofibroblasts can affect cardiomyocyte electrophysiology in vitro, the extent of myofibroblast-myocyte electrical conductance in a syncytium is unknown. In this neonatal rat study, cardiac myofibroblasts were transduced with Channelrhodopsin-2, which allowed acute and selective increase of myofibroblast current, and plated on top of cardiomyocytes. Optical mapping revealed significantly decreased conduction velocity (- 27 ± 6%, p < 10-3), upstroke rate (- 13 ± 4%, p = 0.002), and action potential duration (- 14 ± 7%, p = 0.004) in co-cultures when 0.017 mW/mm2 light was applied, as well as focal spontaneous beating in 6/7 samples and a decreased cycle length (- 36 ± 18%, p = 0.002) at 0.057 mW/mm2 light. In silico modeling of the experiments reproduced the experimental findings and suggested the light levels used in experiments produced excess current similar in magnitude to endogenous myofibroblast current. Fitting the model to experimental data predicted a tissue-level electrical conductance across the 3-D interface between myofibroblasts and cardiomyocytes of ~ 5 nS/cardiomyocyte, and showed how increased myofibroblast-myocyte conductance, increased myofibroblast/myocyte capacitance ratio, and increased myofibroblast current, which occur in fibrosis, can work in tandem to produce pro-arrhythmic increases in conduction and spontaneous beating.

DOI: https://doi.org/10.1038/s41598-021-83398-4