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Background Neutrophils are thought to be short‐lived first responders to tissue injuries such as myocardial infarction (MI), but little is known about their diversification or dynamics. Methods and Results We permanently ligated the left anterior descending coronary arteries of mice and performed single‐cell RNA sequencing and analysis of >28 000 neutrophil transcriptomes isolated from the heart, peripheral blood, and bone marrow of mice on days 1 to 4 after MI or at steady‐state. Unsupervised clustering of cardiac neutrophils revealed 5 major subsets, 3 of which originated in the bone marrow, including a late‐emerging granulocyte expressing SiglecF, a marker classically used to define eosinophils. SiglecF HI neutrophils represented ≈25% of neutrophils on day 1 and grew to account for >50% of neutrophils by day 4 post‐MI. Validation studies using quantitative polymerase chain reaction of fluorescent‐activated cell sorter sorted Ly6G + SiglecF HI and Ly6G + SiglecF LO neutrophils confirmed the distinct nature of these populations. To confirm that the cells were neutrophils rather than eosinophils, we infarcted GATA‐deficient mice (∆dblGATA) and observed similar quantities of infiltrating Ly6G + SiglecF HI cells despite marked reductions of conventional eosinophils. In contrast to other neutrophil subsets, Ly6G + SiglecF HI neutrophils expressed high levels of Myc‐regulated genes, which are associated with longevity and are consistent with the persistence of this population on day 4 after MI. Conclusions Overall, our data provide a spatial and temporal atlas of neutrophil specialization in response to MI and reveal a dynamic proinflammatory cardiac Ly6G + SigF + (Myc + NFϰB + ) neutrophil that has been overlooked because of negative selection.

Cardiac injury triggers an acute immune response that drives tissue healing and remodeling via the activation of compensatory mechanisms. Over time, remodeling and inflammation become chronic and have adverse effects that lead to a depression of cardiac function and eventual heart failure. Cardiac inflammation is characterized by dynamic spatial and temporal crosstalk between the resident cells of the heart and recruitment of circulating leukocytes. Until recently, the cardiomyocyte has not been accepted as a direct contributor to cardiac inflammation. It has now emerged as a key initiator of the acute immune response via its ability to produce cytokines and may also synchronize leukocyte recruitment post-injury. This review will focus on the role of the cardiomyocyte in the acute immune response to ischemic and non-ischemic injury and the mechanisms by which it may influence the course of cardiac remodeling and failure.

Author(s): Anderson Enni, Jacqueline Bella | Advisor(s): Heller Brown, Joan | Abstract: We previously demonstrated a significant role for NLRP3 inflammasome activation in cardiomyocytes in cardiac inflammation, remodeling and contractile dysfunction in response to stressors such as Angiotensin II infusion and pressure overload. There is limited data regarding responses of cardiomyocytes to interleukin-1 β (IL-1β) and interleukin 18 (IL-18), which are generated as a result of NLRP3 activation during the cardiac inflammatory response. To determine the effects of IL-1β and IL-18 on cardiomyocytes, isolated neonatal rat ventricular myocytes (NRVMs) were treated for various times with 1ng/mL recombinant IL-1β or 10ng/mL recombinant IL-18. Western blotting was used to measure the activation state of protein kinases and downstream signaling pathways. Quantitative Polymerase Chain Reaction (qPCR) was used to assess the expression of pro-inflammatory cytokines and chemokines in cardiomyocytes. Proteins and genes showed enhanced phosphorylation and expression when treated with either cytokine, but the majority of NRVMs responded with greater significance to IL-1β stimulation. After treatment with IL-1β, Western blotting showed a several-fold increase in nuclear translocation of NF-κB p65 subunit and phosphorylation of mitogen-activated protein kinases (MAPKs) and P-STAT3, which regulate inflammatory functions. qPCR data showed significant increases in gene expression of pro-inflammatory chemokines and cytokines CXCL1, TGF-β and IL-1β by exogenous IL-1β. We have previously shown that the cardiomyocyte is a source of proinflammatory cytokines and chemokines, including IL-1β and IL-18. Our results suggest the possibility that IL-1β and IL-18 activate a pro-inflammatory positive feedback loop in cardiomyocytes which enhances and sustains cardiac inflammation, contributing to cardiac remodeling under stress conditions.