Abstract
Myocardial infarction causes sympathetic activation and parasympathetic dysfunction, which increase risk of sudden death due to ventricular arrhythmias. Mechanisms underlying parasympathetic dysfunction are unclear. The aim of this study was to delineate consequences of myocardial infarction on parasympathetic myocardial neurotransmitter levels and the function of parasympathetic cardiac ganglia neurons, and to assess electrophysiological effects of vagal nerve stimulation on ventricular arrhythmias in a chronic porcine infarct model. While norepinephrine levels decreased, cardiac acetylcholine levels remained preserved in border zones and viable myocardium of infarcted hearts. In vivo neuronal recordings demonstrated abnormalities in firing frequency of parasympathetic neurons of infarcted animals. Neurons that were activated by parasympathetic stimulation had low basal firing frequency, while neurons that were suppressed by left vagal nerve stimulation had abnormally high basal activity. Myocardial infarction increased sympathetic inputs to parasympathetic convergent neurons. However, the underlying parasympathetic cardiac neuronal network remained intact. Augmenting parasympathetic drive with vagal nerve stimulation reduced ventricular arrhythmia inducibility by decreasing ventricular excitability and heterogeneity of repolarization of infarct border zones, an area with known proarrhythmic potential. Preserved acetylcholine levels and intact parasympathetic neuronal pathways can explain the electrical stabilization of infarct border zones with vagal nerve stimulation, providing insight into its antiarrhythmic benefit.
Authors
Marmar Vaseghi, Siamak Salavatian, Pradeep S. Rajendran, Daigo Yagishita, William R. Woodward, David Hamon, Kentaro Yamakawa, Tadanobu Irie, Beth A. Habecker, Kalyanam Shivkumar
Abstract
Widespread changes in cardiac gene expression occur during heart failure, yet the mechanisms responsible for coordinating these changes remain poorly understood. The Mediator complex represents a nodal point for modulating transcription by bridging chromatin-bound transcription factors with RNA polymerase II activity; it is reversibly regulated by its cyclin-dependent kinase 8 (Cdk8) kinase submodule. Here, we identified increased Cdk8 protein expression in human failing heart explants and determined the consequence of this increase in cardiac-specific Cdk8-expressing mice. Transgenic Cdk8 overexpression resulted in progressive dilated cardiomyopathy, heart failure, and premature lethality. Prior to functional decline, left ventricular cardiomyocytes were dramatically elongated, with disorganized transverse tubules and dysfunctional calcium handling. RNA sequencing results showed that myofilament gene isoforms not typically expressed in adult cardiomyocytes were enriched, while oxidative phosphorylation and fatty acid biosynthesis genes were downregulated. Interestingly, candidate upstream transcription factor expression levels and MAPK signaling pathways thought to determine cardiomyocyte size remained relatively unaffected, suggesting that Cdk8 functions within a novel growth regulatory pathway. Our findings show that manipulating cardiac gene expression through increased Cdk8 levels is detrimental to the heart by establishing a transcriptional program that induces pathological remodeling and eccentric hypertrophy culminating in heart failure.
Authors
Duane D. Hall, Jessica M. Ponce, Biyi Chen, Kathryn M. Spitler, Adrianne Alexia, Gavin Y. Oudit, Long-Sheng Song, Chad E. Grueter
Abstract
Among children with the most severe presentation of Marfan syndrome (MFS), an inherited disorder of connective tissue caused by a deficiency of extracellular fibrillin-1, heart failure is the leading cause of death. Here, we show that, while MFS mice (Fbn1C1039G/+ mice) typically have normal cardiac function, pressure overload (PO) induces an acute and severe dilated cardiomyopathy in association with fibrosis and myocyte enlargement. Failing MFS hearts show high expression of TGF-β ligands, with increased TGF-β signaling in both nonmyocytes and myocytes; pathologic ERK activation is restricted to the nonmyocyte compartment. Informatively, TGF-β, angiotensin II type 1 receptor (AT1R), or ERK antagonism (with neutralizing antibody, losartan, or MEK inhibitor, respectively) prevents load-induced cardiac decompensation in MFS mice, despite persistent PO. In situ analyses revealed an unanticipated axis of activation in nonmyocytes, with AT1R-dependent ERK activation driving TGF-β ligand expression that culminates in both autocrine and paracrine overdrive of TGF-β signaling. The full compensation seen in wild-type mice exposed to mild PO correlates with enhanced deposition of extracellular fibrillin-1. Taken together, these data suggest that fibrillin-1 contributes to cardiac reserve in the face of hemodynamic stress, critically implicate nonmyocytes in disease pathogenesis, and validate ERK as a therapeutic target in MFS-related cardiac decompensation.
Authors
Rosanne Rouf, Elena Gallo MacFarlane, Eiki Takimoto, Rahul Chaudhary, Varun Nagpal, Peter P. Rainer, Julia G. Bindman, Elizabeth E. Gerber, Djahida Bedja, Christopher Schiefer, Karen L. Miller, Guangshuo Zhu, Loretha Myers, Nuria Amat-Alarcon, Dong I. Lee, Norimichi Koitabashi, Daniel P. Judge, David A. Kass, Harry C. Dietz
Abstract
Myocardial atrophy is a wasting of cardiac muscle due to hemodynamic unloading. Doxorubicin is a highly effective anticancer agent but also induces myocardial atrophy through a largely unknown mechanism. Here, we demonstrate that inhibiting transient receptor potential canonical 3 (TRPC3) channels abolishes doxorubicin-induced myocardial atrophy in mice. Doxorubicin increased production of ROS in rodent cardiomyocytes through hypoxic stress–mediated upregulation of NADPH oxidase 2 (Nox2), which formed a stable complex with TRPC3. Cardiomyocyte-specific expression of TRPC3 C-terminal minipeptide inhibited TRPC3-Nox2 coupling and suppressed doxorubicin-induced reduction of myocardial cell size and left ventricular (LV) dysfunction, along with its upregulation of Nox2 and oxidative stress, without reducing hypoxic stress. Voluntary exercise, an effective treatment to prevent doxorubicin-induced cardiotoxicity, also downregulated the TRPC3-Nox2 complex and promoted volume load–induced LV compliance, as demonstrated in TRPC3-deficient hearts. These results illustrate the impact of TRPC3 on LV compliance and flexibility and, focusing on the TRPC3-Nox2 complex, provide a strategy for prevention of doxorubicin-induced cardiomyopathy.
Authors
Tsukasa Shimauchi, Takuro Numaga-Tomita, Tomoya Ito, Akiyuki Nishimura, Ryosuke Matsukane, Sayaka Oda, Sumio Hoka, Tomomi Ide, Norimichi Koitabashi, Koji Uchida, Hideki Sumimoto, Yasuo Mori, Motohiro Nishida
Abstract
The Mediator complex regulates gene transcription by linking basal transcriptional machinery with DNA-bound transcription factors. The activity of the Mediator complex is mainly controlled by a kinase submodule that is composed of 4 proteins, including MED12. Although ubiquitously expressed, Mediator subunits can differentially regulate gene expression in a tissue-specific manner. Here, we report that MED12 is required for normal cardiac function, such that mice with conditional cardiac-specific deletion of MED12 display progressive dilated cardiomyopathy. Loss of MED12 perturbs expression of calcium-handling genes in the heart, consequently altering calcium cycling in cardiomyocytes and disrupting cardiac electrical activity. We identified transcription factors that regulate expression of calcium-handling genes that are downregulated in the heart in the absence of MED12, and we found that MED12 localizes to transcription factor consensus sequences within calcium-handling genes. We showed that MED12 interacts with one such transcription factor, MEF2, in cardiomyocytes and that MED12 and MEF2 co-occupy promoters of calcium-handling genes. Furthermore, we demonstrated that MED12 enhances MEF2 transcriptional activity and that overexpression of both increases expression of calcium-handling genes in cardiomyocytes. Our data support a role for MED12 as a coordinator of transcription through MEF2 and other transcription factors. We conclude that MED12 is a regulator of a network of calcium-handling genes, consequently mediating contractility in the mammalian heart.
Authors
Kedryn K. Baskin, Catherine A. Makarewich, Susan M. DeLeon, Wenduo Ye, Beibei Chen, Nadine Beetz, Heinrich Schrewe, Rhonda Bassel-Duby, Eric N. Olson
Abstract
Pediatric dilated cardiomyopathy (DCM) is the most common indication for heart transplantation in children. Despite similar genetic etiologies, medications routinely used in adult heart failure patients do not improve outcomes in the pediatric population. The mechanistic basis for these observations is unknown. We hypothesized that pediatric and adult DCM comprise distinct pathological entities, in that children do not undergo adverse remodeling, the target of adult heart failure therapies. To test this hypothesis, we examined LV specimens obtained from pediatric and adult donor controls and DCM patients. Consistent with the established pathophysiology of adult heart failure, adults with DCM displayed marked cardiomyocyte hypertrophy and myocardial fibrosis compared with donor controls. In contrast, pediatric DCM specimens demonstrated minimal cardiomyocyte hypertrophy and myocardial fibrosis compared with both age-matched controls and adults with DCM. Strikingly, RNA sequencing uncovered divergent gene expression profiles in pediatric and adult patients, including enrichment of transcripts associated with adverse remodeling and innate immune activation in adult DCM specimens. Collectively, these findings reveal that pediatric and adult DCM represent distinct pathological entities, provide a mechanistic basis to explain why children fail to respond to adult heart failure therapies, and suggest the need to develop new approaches for pediatric DCM.
Authors
Meghna D. Patel, Jayaram Mohan, Caralin Schneider, Geetika Bajpai, Enkhsaikhan Purevjav, Charles E. Canter, Jeffrey Towbin, Andrea Bredemeyer, Kory J. Lavine
Abstract
Molecular chaperones regulate quality control in the human proteome, pathways that have been implicated in many diseases, including heart failure. Mutations in the BAG3 gene, which encodes a co-chaperone protein, have been associated with heart failure due to both inherited and sporadic dilated cardiomyopathy. Familial BAG3 mutations are autosomal dominant and frequently cause truncation of the coding sequence, suggesting a heterozygous loss-of-function mechanism. However, heterozygous knockout of the murine BAG3 gene did not cause a detectable phenotype. To model BAG3 cardiomyopathy in a human system, we generated an isogenic series of human induced pluripotent stem cells (iPSCs) with loss-of-function mutations in BAG3. Heterozygous BAG3 mutations reduced protein expression, disrupted myofibril structure, and compromised contractile function in iPSC-derived cardiomyocytes (iPS-CMs). BAG3-deficient iPS-CMs were particularly sensitive to further myofibril disruption and contractile dysfunction upon exposure to proteasome inhibitors known to cause cardiotoxicity. We performed affinity tagging of the endogenous BAG3 protein and mass spectrometry proteomics to further define the cardioprotective chaperone complex that BAG3 coordinates in the human heart. Our results establish a model for evaluating protein quality control pathways in human cardiomyocytes and their potential as therapeutic targets and susceptibility factors for cardiac drug toxicity.
Authors
Luke M. Judge, Juan A. Perez-Bermejo, Annie Truong, Alexandre J.S. Ribeiro, Jennie C. Yoo, Christina L. Jensen, Mohammad A. Mandegar, Nathaniel Huebsch, Robyn M. Kaake, Po-Lin So, Deepak Srivastava, Beth L. Pruitt, Nevan J. Krogan, Bruce R. Conklin
Abstract
Our previous work showed myocellular differences in pediatric and adult dilated cardiomyopathy (DCM). However, a thorough characterization of the molecular pathways involved in pediatric DCM does not exist, limiting the development of age-specific therapies. To characterize this patient population, we investigated the transcriptome profile of pediatric patients. RNA-Seq from 7 DCM and 7 nonfailing (NF) explanted age-matched pediatric left ventricles (LV) was performed. Changes in gene expression were confirmed by real-time PCR (RT-PCR) in 36 DCM and 21 NF pediatric hearts and in 20 DCM and 10 NF adult hearts. The degree of myocyte hypertrophy was investigated in 4 DCM and 7 NF pediatric hearts and in 4 DCM and 9 NF adult hearts. Changes in gene expression in response to pluripotency-inducing factors were investigated in neonatal rat ventricular myocytes (NRVMs). Transcriptome analysis identified a gene expression profile in children compared with adults with DCM. Additionally, myocyte hypertrophy was not observed in pediatric hearts but was present in adult hearts. Furthermore, treatment of NRVMs with pluripotency-inducing factors recapitulated changes in gene expression observed in the pediatric DCM heart. Pediatric DCM is characterized by unique changes in gene expression that suggest maintenance of an undifferentiated state.
Authors
Philip D. Tatman, Kathleen C. Woulfe, Anis Karimpour-Fard, Danielle A. Jeffrey, James Jaggers, Joseph C. Cleveland, Karin Nunley, Matthew R.G. Taylor, Shelley D. Miyamoto, Brian L. Stauffer, Carmen C. Sucharov
Abstract
Increasing NAD+ levels by supplementing with the precursor nicotinamide mononucleotide (NMN) improves cardiac function in multiple mouse models of disease. While NMN influences several aspects of mitochondrial metabolism, the molecular mechanisms by which increased NAD+ enhances cardiac function are poorly understood. A putative mechanism of NAD+ therapeutic action exists via activation of the mitochondrial NAD+-dependent protein deacetylase sirtuin 3 (SIRT3). We assessed the therapeutic efficacy of NMN and the role of SIRT3 in the Friedreich’s ataxia cardiomyopathy mouse model (FXN-KO). At baseline, the FXN-KO heart has mitochondrial protein hyperacetylation, reduced Sirt3 mRNA expression, and evidence of increased NAD+ salvage. Remarkably, NMN administered to FXN-KO mice restores cardiac function to near-normal levels. To determine whether SIRT3 is required for NMN therapeutic efficacy, we generated SIRT3-KO and SIRT3-KO/FXN-KO (double KO [dKO]) models. The improvement in cardiac function upon NMN treatment in the FXN-KO is lost in the dKO model, demonstrating that the effects of NMN are dependent upon cardiac SIRT3. Coupled with cardio-protection, SIRT3 mediates NMN-induced improvements in both cardiac and extracardiac metabolic function and energy metabolism. Taken together, these results serve as important preclinical data for NMN supplementation or SIRT3 activator therapy in Friedreich’s ataxia patients.
Authors
Angelical S. Martin, Dennis M. Abraham, Kathleen A. Hershberger, Dhaval P. Bhatt, Lan Mao, Huaxia Cui, Juan Liu, Xiaojing Liu, Michael J. Muehlbauer, Paul A. Grimsrud, Jason W. Locasale, R. Mark Payne, Matthew D. Hirschey
Abstract
We previously showed that angiotensin II (Ang II) increases T cell production of IL-17A, and that mice deficient in IL-17A have blunted hypertension and attenuated renal and vascular dysfunction. It was recently shown that salt enhances IL-17A production from CD4+ T cells via a serum- and glucocorticoid-regulated kinase 1–dependent (SGK1-dependent) pathway. Thus, we tested the hypothesis that SGK1 signaling in T cells promotes hypertension and contributes to end-organ damage. We show that loss of T cell SGK1 results in a blunted hypertensive response to Ang II infusion by 25 mmHg. Importantly, renal and vascular inflammation is abrogated in these mice compared with control mice. Furthermore, mice lacking T cell SGK1 are protected from Ang II–induced endothelial dysfunction and renal injury. Loss of T cell SGK1 also blunts blood pressure and vascular inflammation in response to deoxycorticosterone acetate–salt (DOCA-salt) hypertension. Finally, we demonstrate that the Na+-K+-2Cl– cotransporter 1 (NKCC1) is upregulated in Th17 cells and is necessary for the salt-induced increase in SGK1 and the IL-23 receptor. These studies demonstrate that T cell SGK1 and NKCC1 may be novel therapeutic targets for the treatment of hypertension and identify a potentially new mechanism by which salt contributes to hypertension.
Authors
Allison E. Norlander, Mohamed A. Saleh, Arvind K. Pandey, Hana A. Itani, Jing Wu, Liang Xiao, Jooeun Kang, Bethany L. Dale, Slavina B. Goleva, Fanny Laroumanie, Liping Du, David G. Harrison, Meena S. Madhur
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