Interleukin-5-induced Eosinophil Population Improves Cardiac Function After Myocardial Infarction

Background and aims:Interleukin(IL)-5 mediates the development of eosinophils(EOS)that are essential for tissue post-injury repairs.It is unknown whether IL-5 or EOS play any role in the heart repairs after myocardial infarction(MI).This study aims to test whether IL-5-induced EOS population promotes the healing and repair process post-MI and reveal the underlying mechanisms.Methods:MI was induced by permanent ligation of the left anterior descending coronary artery in C57BL/6 mice.Expression of IL-5 in heart was detected by western blot(WB)analysis and real-time polymerase chain reaction(RT-PCR).Fluorescence-activated cell sorting(FACS)and immunofluorescence was applied to investigate the cellular sources of IL-5.Recombinant mouse IL-5 or equal volume of phosphate-buffered saline(20 minutes,1 day and 2 days after MI surgery)was injected intraperitoneally in MI mice with or without TRFK5 pretreatment(EOS depletion).Cardiac function was analyzed at day 28 after MI.Early and late infarct size and angiogenesis in the border zone was determined by 2,3,5-triphenyltetrazolium chloride(TTC)staining,Sirius red staining and immunohistochemistry,respectively.FACS,hematoxylin-eosin staining and immunohistochemistry were used to evaluate the number of EOS in heart.EOS in peripheral blood was automatically enumerated by a hematology analyzer.CD206+macrophages were evaluated using immunofluorescent staining and FACS at day 7 after MI.RT-PCR examined the levels of CD206+macrophage-related gene expression.EOS and bone marrow derived macrophages(BMDMs)were co-cultured in a Transwell system to investigate the impact of EOS on macrophage polarization.FACS and RT-PCR were used to examine the proportion of CD206+ macrophages and the levels of CD206+macrophage-related gene expression in BMDMs,respectively.Protein levels of total signal transducer and activator of transcription 6(STAT6)and phosphorylated STAT6 in BMDMs or sorted cardiac macrophages at day 7 post-MI were determined by WB analysis.Results:WB and RT-PCR analysis revealed elevated expression of IL-5 in the heart at 5 days post-MI.CD45+cells(macrophages and type 2 innate lymphoid cells),but not CD45-cells(endothelial cells and fibroblasts)in heart were the main source of IL-5.External supply of recombinant mouse IL-5 reduced the early and late infarct size,increased angiogenesis in the border zone and improved the cardiac function after MI.A significant expansion of EOS was detected in both the peripheral blood and infarcted myocardium after IL-5 administration.Pharmacological depletion of EOS by TRFK5 pretreatment muted the beneficial effects of IL-5 in MI mice.Mechanistic studies also demonstrated that IL-5 increased the accumulation of CD206+macrophages,as well as the expression of CD206+macrophage-related genes in infarcted myocardium at 7 days post-MI.TRFK5 pretreatament inhibited effects of IL-5 on CD206+macrophage polarization.In vitro co-culture experiments showed that EOS shifted BMDMs towards the CD206+phenotypes.This activity of EOS was abolished by IL-4 neutralizing antibody,but not IL-10 or IL-13 neutralization.WB analyses demonstrated that EOS promoted the BMDM downstream STAT6 phosphorylation.Consistently,STAT6 phosphorylation was promoted in sorted cardiac macrophages after IL-5 treatment.Conclusion:IL-5 facilitates the recovery of cardiac dysfunction post-MI by promoting EOS accumulation and subsequent CD206+macrophage polarization via the IL-4/STAT6 axis.