Abstract P3077: The Role Of Neutrophils In Initiating Type I Interferon Response In Myocardial Ischemia/reperfusion

Abstract

Background: Inflammation succeeding myocardial ischemia/reperfusion (MI/R) plays a significant role in damaging cardiomyocytes and influencing infarct size, with neutrophils being the most rapid and most numerous cells in the post-MI/R heart. While neutrophils play a role in wound healing, they can cause further damage through the generation of reactive oxygen species, proteolytic enzymes, and neutrophil extracellular traps (NETs). We thus hypothesize that double stranded DNA stemming from these NETs activates cGAS-STING signaling in a murine model of MI/R, leading to the production of type I interferons by neutrophils, which worsens reperfusion injury. Methods: Single-cell RNA sequencing was performed on neutrophils isolated from the hearts of mice 24 hours post-MI/R. Additionally, mice underwent MI/R and heart tissues were harvested 6- and 24- hours after reperfusion for ELISA and qPCR analysis. Results: Utilizing single-cell RNA sequencing to better characterize the role of neutrophils and NETs in MI/R, we identified a significant population of neutrophils with a strong type I interferon (IFN) signature recruited to the area of infarct 24 hours post MI/R in a murine model. To further investigate the IFN response seen in this population of neutrophils, we preformed protein and gene expression analysis on heart tissue samples isolated from mice at timepoints in which neutrophils would be present in the area of infarct. Analysis by qPCR shows an increased fold change in gene expression of IFNα (7.22 ± 6.05, n = 3; 3.83 ± 2.22, n = 4) and IFNβ (6.74 ± 5.39, n = 3; 3.64 ± 2.25, n = 4) at 6- and 24- hours post MI/R respectively, compared to sham mice. Furthermore, analysis by ELISA shows a significant increase in IFNβ protein levels in the heart after 6- hours (0.41pg/mg ± 0.04, n =8) compared to sham mice (0.16 pg/mg ± 0.06, n = 7; p ≤ 0.01, Student’s t-test). Conclusions: These data show a rapid increase in type I IFN in the hearts of mice post-MI/R, which implicates neutrophils may be participating in the IFN signaling pathway. Identifying neutrophils as an early source of IFN can provide a potentially new therapeutic target in treating MI/R.