Cardiac fibrosis begins as an intrinsic response to injury or ageing that functions to preserve the tissue from further damage. Fibrosis results from activated cardiac myofibroblasts, which secrete extracellular matrix (ECM) proteins in an effort to replace damaged tissue; however, excessive ECM deposition leads to pathological fibrotic remodeling. At this extent, fibrosis gravely disturbs myocardial compliance, and ultimately leads to adverse outcomes like heart failure with heightened mortality. As such, understanding the complexity behind fibrotic remodeling has been a focal point of cardiac research in recent years. Resident cardiac fibroblasts and activated myofibroblasts have been proven integral to the fibrotic response; however, several findings point to additional cell types that may contribute to the development of pathological fibrosis. For one, leukocytes expand in number after injury and exhibit high plasticity, thus their distinct role(s) in cardiac fibrosis is an ongoing and controversial field of study. This review summarizes current findings, focusing on both direct and indirect leukocyte-mediated mechanisms of fibrosis, which may provide novel targeted strategies against fibrotic remodeling.

Aims Myocardial infarction (MI) is the most common cause of heart failure (HF) worldwide. G protein-coupled receptor kinase 5 (GRK5) is upregulated in failing human myocardium and promotes maladaptive cardiac hypertrophy in animal models. However, the role of GRK5 in ischemic heart disease is still unknown. In this study, we evaluated whether myocardial GRK5 plays a critical role post-MI in mice and included examination of specific cardiac immune and inflammatory responses. Methods and results Cardiomyocyte-specific GRK5 overexpressing transgenic mice (TgGRK5) and non-transgenic littermate control (NLC) mice as well as cardiomyocyte-specific GRK5 knockout mice (GRK5cKO) and wild type (WT) were subjected to MI and, functional as well as structural changes together with outcomes were studied. TgGRK5 post-MI mice showed decreased cardiac function, augmented left ventricular dimension and decreased survival rate compared to NLC post-MI mice. Cardiac hypertrophy and fibrosis as well as fetal gene expression were increased post-MI in TgGRK5 compared to NLC mice. In TgGRK5 mice, GRK5 elevation produced immuno-regulators that contributed to the elevated and long-lasting leukocyte recruitment into the injured heart and ultimately to chronic cardiac inflammation. We found an increased presence of pro-inflammatory neutrophils and macrophages as well as neutrophils, macrophages and T- lymphocytes at 4- days and 8- weeks respectively post-MI in TgGRK5 hearts. Conversely, GRK5cKO mice were protected from ischemic injury and showed reduced early immune cell recruitment (predominantly monocytes) to the heart, improved contractility and reduced mortality compared to WT post-MI mice. Interestingly, cardiomyocyte-specific GRK2 transgenic mice did not share the same phenotype of TgGRK5 mice and did not have increased cardiac leukocyte migration and cytokine or chemokine production post-MI. Conclusions Our study shows that myocyte GRK5 has a crucial and GRK-selective role on the regulation of leucocyte infiltration into the heart, cardiac function and survival in a murine model of post-ischemic HF, supporting GRK5 inhibition as a therapeutic target for HF. TRANSLATIONAL PERSPECTIVE GRK5 is upregulated in failing human myocardium and associated with heart failure development. In this study, we evaluated whether cardiomyocyte GRK5 plays a critical role during ischemic heart disease in a mouse animal model. We discovered that GRK5 overexpression in cardiomyocyte affects cardiac function, remodeling, immune cell recruitment, and ultimately survival in ischemic heart failure. Conversely, cardiomyocyte-specific GRK5 ablation diminished the early immune cell infiltration in the heart, improved contractility and reduced mortality post-myocardial infarction. The overall translational significance of these findings is substantial, as selective small molecule inhibitors of GRK5 have begun to emerge as novel therapeutic treatment in heart disease.

Aims Epidermal growth factor receptor (EGFR) is essential to the development of multiple tissues and organs and is a target of cancer therapeutics. Due to the embryonic lethality of global EGFR deletion and conflicting reports of cardiac-overexpressed EGFR mutants, its specific impact on the adult heart, normally or in response to chronic stress, has not been established. Using complimentary genetic strategies to modulate cardiomyocyte-specific EGFR expression, we aim to define its role in the regulation of cardiac function and remodelling. Methods and results A floxed EGFR mouse model with α-myosin heavy chain-Cre-mediated cardiomyocyte-specific EGFR downregulation (CM-EGFR-KD mice) developed contractile dysfunction by 9 weeks of age, marked by impaired diastolic relaxation, as monitored via echocardiographic, haemodynamic, and isolated cardiomyocyte contractility analyses. This contractile defect was maintained over time without overt cardiac remodelling until 10 months of age, after which the mice ultimately developed severe heart failure and reduced lifespan. Acute downregulation of EGFR in adult floxed EGFR mice with adeno-associated virus 9 (AAV9)-encoded Cre with a cardiac troponin T promoter (AAV9-cTnT-Cre) recapitulated the CM-EGFR-KD phenotype, while AAV9-cTnT-EGFR treatment of adult CM-EGFR-KD mice rescued the phenotype. Notably, chronic administration of the β-adrenergic receptor agonist isoproterenol effectively and reversibly compensated for the contractile dysfunction in the absence of cardiomyocyte hypertrophy in CM-EGFR-KD mice. Mechanistically, EGFR downregulation reduced the expression of protein phosphatase 2A regulatory subunit Ppp2r3a/PR72, which was associated with decreased phosphorylation of phospholamban and Ca2+ clearance, and whose re-expression via AAV9-cTnT-PR72 rescued the CM-EGFR-KD phenotype. Conclusions Altogether, our study highlights a previously unrecognized role for EGFR in maintaining contractile homeostasis under physiologic conditions in the adult heart via regulation of PR72 expression.

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Fish has been an important source of proteins, essential vitamins, and low saturated fats for centuries. However, improperly handled fish can expose consumers to infectious bacteria, including difficult to treat multidrug-resistant pathogens. With the goal to investigate the existence of disease-causing and antibiotic-resistant bacteria, we examined bacterial communities present on various types of fish purchased from supermarkets in Houston, Texas, USA. The bacterial communities were characterized by selective phenotypic culture methods, 16S ribosomal RNA gene sequencing, and antibiotic susceptibility testing. The results revealed the presence of different bacterial communities on the fish samples examined. The bacterial communities were not significantly different between the supermarkets sampled. The following presumptive human pathogens were isolated on the fish samples: Escherichia coli (67%), enterohemorrhagic E. coli (31%), Shigella and Salmonella species (28%), Listeria species (29%), and Staphylococcus aureus (28%). Drug sensitivity assays showed resistance to commonly prescribed antibiotics ciprofloxacin, gentamicin, and vancomycin. Out of a total of 99 E. coli samples tested, 41.4% were resistant to ciprofloxacin, whereas 33.3% were resistant to gentamicin. Of the total of 31 S. aureus isolates tested, 87% were resistant to ciprofloxacin, whereas 61.3% were resistant to vancomycin. Moreover, some of the E. coli strains were resistant to both ciprofloxacin and gentamicin (28%), whereas 49% of the S. aureus isolates were resistant to both ciprofloxacin and vancomycin. These results highlight the prevalence of antimicrobial-resistant foodborne pathogens on fish purchased from the supermarkets and underscore the risk associated with improper handling of fish.

A mountain of evidence suggests paramount roles for myeloid cells in cardiac physiology. For one, these leukocytes have been reported to play major roles in the maintenance of functional and structural homeostasis. Additionally, following an injury, myeloid cell processes can enormously influence both short‐ and long‐term remodeling and repair outcomes, ultimately informing heart failure. Given these significant roles, a lot of recent work has focused on examining exactly how these leukocytes might be regulated. Epidermal growth factor receptor (EGFR) is a tyrosine kinase receptor that is known to critically govern cell function through for instance proliferation, migration, and survival. Interestingly, recent reports suggest that EGFR regulates macrophage activation and function, however it is currently unknown if, and how EGFR might influence myeloid cell responses within the heart. Thus, our group has generated myeloid cell‐specific EGFR knockout mice (EGFRmylKO) to determine the impact of such deletion on cardiac homeostasis and post injury outcomes. We hypothesize that myeloid cell‐specific EGFR is a central regulator of cardiovascular inflammation, and is key in influencing post injury outcomes. To date, we have identified that in the absence of myeloid expressed EGFR, 12‐16‐week‐old mice exhibit increased cardiomyocyte size and “fetal gene program” transcripts when compared to both age matched floxed EGFR (EGFRf/f) and LysM cre controls. Further, we have subjected EGFRmylKOand controls to experimental myocardial infarction (MI), and have observed decreased systolic function inEGFRmylKO. To begin understanding these phenotypes, we have analyzed basal and post injury cardiacmyeloid populations. We have observed that EGFRmylKO mice exhibit increased c‐c chemokine receptor type2 (CCR2+) resident macrophages, and an increase in monocytes and macrophages 1 week following MI. Altogether, these results suggest novel roles for EGFR in the failing heart.

During heart failure, chronically decreased cardiac output can be treated with positive inotropes, but classic inotropes such as β-adrenergic receptor (βAR) agonists that increase cAMP-dependent Ca 2+ mobilization and contractility ultimately enhance patient mortality. Thus, an alternate approach would be to enhance cardiomyocyte contractility without alterations in cAMP and Ca 2+ levels, such as regulation of sarcomeric proteins. Recently, we demonstrated that a small lipidated pepducin designed from the 1 st intracellular loop of β2AR (ICL1-9) enhanced cardiomyocyte contractility in a Ca 2+ -independent, β-arrestin-dependent manner, yet the complete mechanisms remained unclear. We also showed that β2AR stimulation in hearts in vivo or neonatal rat ventricular myocytes (NRVM) in vitro activated RhoA in a βarr-dependent manner. Therefore, we sought to determine both the proximal and distal mechanisms by which ICL1-9 enhances cardiomyocyte contractility. Using adult murine cardiomyocytes isolated from wild-type C57Bl/6J mice, we measured basal, ICL1-9- and isoproterenol (ISO, as a positive control)-promoted contractility either alone or in the presence of inhibitors of Gα i activity (Pertussis toxin), ROCK1 (Y-27632), myosin light chain kinase (ML7), and protein kinase D (CID755673). Inhibition of Gα i activity prior to ICL1-9 stimulation led to a decreased contractile response. Consistent with RhoA activation by ICL1-9, ROCK1 inhibition was able to attenuate ICL1-9-mediated contractility, as was inhibition of MLCK. Interestingly, we observed that inhibition of PKD also attenuated ICL1-9-mediated contractility. These data suggest that ICL1-9 acts proximally to engage a β2AR/Gα i /βarr signaling axis, which may distally increase the activation of kinases including PKD, MLCK, and ROCK to alter the regulation of sarcomeric proteins.