Author + information
- aAab Cardiovascular Research Institute, Rochester, New York
- bDepartment of Medicine and Department of Pharmacology and Physiology, University of Rochester School of Medicine and Dentistry, Rochester, New York
- ↵∗Address for correspondence:
Dr. Eric M. Small, Aab Cardiovascular Research Institute, University of Rochester School of Medicine and Dentistry, 601 Elmwood Avenue, Box CVRI, Rochester, New York 14642.
- G protein–coupled receptor kinase
- transforming growth factor β
Coronary artery occlusion leads to myocardial infarction (MI) and subsequent cell death in ischemic cardiac tissue. Recent clinical advances in the treatment of MI, including rapid reperfusion and aggressive use of antiplatelet agents, limit the extent of cell death and save countless patients who would have died just a few decades ago (1). However, even after successful reperfusion, tissue lost during an MI is never repopulated with functioning cardiomyocytes. Instead, fibroblasts adjacent to the ischemic region proliferate, infiltrate the infarct, and become activated. Activated fibroblasts, also called myofibroblasts, secrete extracellular matrix (ECM) that partially restores structural integrity and prevents cardiac rupture. Although scar formation is initially a beneficial response to cardiac insult and is essential for cardiac repair, unrestrained fibroblast activity leads to expansion of fibrosis, culminating in heart failure (HF). Fueled by epidemics of obesity and diabetes that stimulate atherosclerotic narrowing of coronary vessels and ischemic heart disease, the incidence of HF has skyrocketed and is projected to affect 1 in 33 U.S. citizens by 2030 (2).
The negative effect of cardiac scarring on heart function is multifactorial (3). Interstitial fibrosis is associated with: 1) decreased tissue elasticity, impeding cardiac contraction and relaxation; 2) reduced left ventricular diastolic filling; 3) increased heart chamber size; 4) attenuation of the angiogenic response and reduced tissue oxygenation; and 5) obstruction of electrical propagation, altered heart rhythm, and sometimes lethal arrhythmias. Limiting pathological cardiac fibrosis should increase quality of life and reduce the likelihood of cardiac sudden death in heart disease patients and was the focus of the study by Travers et al. (4) in this issue of the Journal.
Cardiac fibroblasts have long been neglected in cardiovascular medicine, as they were thought to affect form more than function. Yet, accumulating evidence has revealed that fibroblasts actively participate in normal cardiac homeostasis and disease progression, and should be the focus of new therapeutic strategies to attenuate HF progression. Fibroblasts respond to neurohumoral, inflammatory, and mechanical signaling, which tune their activity to a changing environment (5,6). Ischemia-reperfusion (I/R) injury following acute MI treatment leads to significant cell death and a robust inflammatory response. Circulating cells populating the heart after MI deposit transforming growth factor beta (TGF)-β1 in the cardiac interstitium. Activation of TGF-β receptors on fibroblasts stimulates Smad-dependent transcription of genes encoding contractile and ECM proteins, generating replacement fibrosis and setting the stage for pathological remodeling.
Mechanical tension generated by myocardial loss during ischemia also leads to activation of latent TGF-β1, linking mechanical tension and inflammation to scar formation. Mechanical tension independently stimulates various pro-fibrotic signal transduction pathways, including Rho-Rho kinase (ROCK)-serum response factor signaling and noncanonical TGF-β1-p38/mitogen-activated protein kinase (MAPK) signaling (7,8). Thus, a feedforward loop can be envisioned in which fibroblast activation leads to ECM deposition and increased tissue rigidity, which is a mechanical substrate for sustained fibroblast activation.
Efforts to break this loop and attenuate myofibroblast activation include TGF-β1–blocking antibodies and chemical inhibitors of TGF-β or Rho-ROCK signaling. TGF-β1 inhibition with monoclonal antibodies has not proven useful at the clinical trial level, and induction of autoimmunity has complicated small molecule studies. Blocking Rho-ROCK signaling with chemical inhibitors (e.g., Y-27632, fasudil, or CCG-203971) attenuated fibrosis in animal models, but has been associated with rare side effects in the clinic, including ocular hyperemia. Given such challenges, an effective and safe antiscarring agent has not yet come to fruition (9).
The Gβγ-G protein–coupled receptor kinase 2 (GRK2 or βARK; gene name ADRBK1) signaling axis has previously been linked to heart disease progression (10). Gβγ is a component of a heterotrimeric G protein, which is activated by G protein–coupled receptors (GPCRs) such as the β-adrenergic receptor (βAR). Upon activation of the βAR, Gαs subunits stimulate adenylate cyclase to generate cyclic adenosine monophosphate, which activates protein kinase A. In addition to myriad cellular functions, protein kinase A drives GRK2-dependent recruitment of β-arrestin to the βAR, preventing further activation of Gαs subunits. Thus, GRK2 in normal physiology prevents overstimulation of βAR. Extended βAR stimulation in heart disease or after I/R injury leads to βAR down-regulation and desensitization, reducing cardiomyocyte contractility (11). The current standard of care for HF includes βAR blockers that restore βAR coupling and improve cardiomyocyte function.
Recent studies suggested that GRK2-based βAR uncoupling is also active in cardiac fibroblasts and may directly lead to myofibroblast activation and fibrosis (12). In this issue of the Journal, Travers et al. (4) investigated the relative contributions of Gβγ-GRK2 signaling in cardiomyocyte dysfunction and fibroblast activation in a mouse model of I/R injury (4). They also utilized pharmacogenetic epistasis experiments to probe the cellular mechanism of gallein, an investigational GRK2 inhibitor. Tissue-specific deletion of Adrbk1 in cardiomyocytes or activated fibroblasts attenuated maladaptive remodeling in response to I/R; cardiac fibrosis was significantly reduced, and both diastolic and systolic function were restored.
The striking improvement in post-I/R cardiac function upon deletion of Adrbk1 in activated fibroblasts led the authors to speculate whether the primary target of gallein was cardiomyocytes or fibroblasts. Restoration of cardiac function by deletion of Adrbk1 in cardiomyocytes was incomplete, but was further improved with an interventional regimen of gallein beginning 1 week post-I/R. In contrast, mice lacking Adrbk1 in activated fibroblasts displayed a complete reversal of maladaptive remodeling and exhibited no further improvement in post-I/R cardiac function with gallein treatment. These results were supported by in vitro data demonstrating reduced myofibroblast activation and increased cyclic adenosine monophosphate levels upon gallein treatment of cultured cardiac fibroblasts from human HF patients.
The authors interpreted the experimental results to indicate that gallein primarily acts through cardiac fibroblasts, although other cellular targets could certainly contribute. One caveat: functional restoration was complete in the fibroblast gene deletion experiments, reducing the ability to draw meaningful conclusions from an epistasis experiment that could not be further improved. Other considerations must include the mouse lines that express Cre recombinase in cardiomyocytes or activated fibroblasts. Efficiency and specificity of Adrbk1 deletion in cardiomyocytes and activated fibroblasts is difficult to directly compare. Furthermore, subtle differences in Cre and tamoxifen toxicity might preclude direct comparison of gallein on post-I/R cellular function. The positive results reported in this study warrant confirmation that dysfunctional Gβγ-GRK2 signaling is a dominant pathway in human cardiac fibroblast activation after MI.
Mechanistically, Gβγ signaling affects cytoskeletal organization and chemotactic migration, which are major factors underlying myofibroblast activation (13). It would be interesting to evaluate the effect of gallein or Gβγ-GRK2 signaling on Rho-ROCK or serum response factor myocardin-related transcription factor–dependent gene transcription, pathways linking actin cytoskeletal dynamics to pro-fibrotic gene programs (14). βAR also signals through Gβγ to activate p38-MAPK (15). Deletion of p38α (Mapk14) in activated fibroblasts attenuated fibrosis in response to βAR overstimulation or MI in mice (7). It may be fruitful to investigate potential crosstalk between Gβγ-GRK2 and p38/MAPK signaling in the context of cardiac fibrosis.
Scar formation is essential to cardiac integrity after MI, but excessive fibrosis is associated with left ventricular dysfunction and poor prognosis. Titrating therapies to attain optimal levels of scar tissue in the post-ischemic heart is of great clinical importance. Genetic deletion of Adrbk1 in activated fibroblasts elegantly accomplished this goal; however, the design of pharmacological strategies targeting Gβγ-GRK2 must spare normal GPCR signaling. In this study, gallein improved cardiac function to an equivalent extent as fibroblast-specific Adrbk1 knockout mice, although clinically relevant side effects remain to be explored. Development of cell-specific delivery methods, coupled with a comprehensive understanding of potential off-target effects of Gβγ inhibitors, are attainable goals that might pave the way for novel strategies to prevent or reverse pathological cardiac scarring.
↵∗ Editorials published in the Journal of the American College of Cardiology reflect the views of the authors and do not necessarily represent the views of JACC or the American College of Cardiology.
The authors are supported by National Institutes of Health/National Heart, Lung, and Blood Institute grants HL120919, HL133761, and HL136179 (to Dr. Small) and grant HL007937 (to Dr. Burke). Dr. Small has received an investigator initiated research project grant from Novartis Pharmaceuticals.
- 2017 American College of Cardiology Foundation
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