Innate Lymphoid Cells Promote Recovery of Ventricular Function After Myocardial Infarction

Figure 1. ILC2s Proliferate in PcAT After MI Flow cytometric analysis detects the expansion of ILC2 3 days post-MI (A) in heart (top) and pericardial adipose tissue accompanied with elevated ST2 surface expression (bottom), and increased proliferation (Ki67) (B). ILC2 proliferation in the PcAT was inhibited by injections of sST2, less pronounced in the heart (C). Representative images shown. Mann-Whitney U test. ILC2 = type 2 innate lymphoid cells; MI = myocardial infarction; PcAT = pericardial adipose tissue; sST2 = soluble ST2.

Figure 2. Diminished Cardiac Function in ILC2KO Mice After MI (A) Survival curve for knockout (KO: Rorafl/flCd127Cre/+, n = 5/14 ruptured) compared with wild type (WT: Rorafl/flCd127wt, n = 1/9 ruptured) mice after experimentally induced MI (P = 0.2). (B) A significant impact on cardiac function (% ejection fraction, EF) in KO mice at Day 14 post-MI (P = 0.0025) (representative images shown) and recovery was severely impacted up to 4 weeks post-surgery (P = 0.0006). (C) WT mice maintained more cardiac muscle compared with KO and had significantly smaller scars. (D) Further, Sirius red labeling demonstrates decreased collagen deposition in the scar tissue of KO mice. Mann-Whitney U test. Abbreviations as in Figure 1.

Figure 3. Alterations to the Myeloid Compartment Following MI Cells from heart (A), pericardial adipose tissue (PcAT) (B), and blood (C). Eosinophils, monocytes, macrophage, and Treg populations were characterized by flow cytometry at Day 5 post-MI. Mann-Whitney U test. Treg = T regulatory cell; other abbreviation as in Figure 1.

Figure 4. ILC2 Transcriptome After MI Highlight a Potentially Protective IL-2 Stimulus RNAseq analysis of ILC2s sorted from the PcAT on Day 3 post-MI. (A) Initial indications of alterations in signature ILC2 transcripts in MI mice as well as significant differences in expression of inflammatory and trafficking molecules. (B) Heatmap of in-depth pathway and predictive gene cluster analysis highlights changes in expression of novel genes induced by infarction, which may be central around modulation of cell proliferation and survival pathways (C). (D) Predictive upstream regulatory pathways identified an IL-2-driven survival fingerprint. IL = interleukin; RNAseq = RNA sequencing; other abbreviations as in Figure 1.

Figure 5. IL-2 Improves ILC2 Function During Acute MI in Rag2−/− Mice IL-2/Jes6-1 treatment enhances serum cytokine secretion of IL-5 at Day 3 post-MI (A). Intracellular detection of IL-5 (B, black histogram FM1, blue PBS, and red IL-2 treated) complements this. Splenic ILC2s (C) were expanded and expressed more IL-5 per cell. Splenic macrophages slew toward Arg1+ population (M2-like). PcAT ILC2s (C) were not expanded but expressed more IL-5 with biased macrophage population toward Arg1+ expression. Heart resident ILC2s (C) were not expanded but expressed more IL-5 per cell. Subsequent heart-infiltrating macrophages slew toward the Arg1+ population (M2-like). Mann-Whitney U test. Arg1+ = Arginase1+; PBS = phosphate-buffered saline; other abbreviations as in Figures 1 and 4.

Figure 6. IL-2 Treatment Improves the Recovery of Heart Function After MI (A)Rag2−/− mice (n = 9 per group) were subjected to LAD ligation and treated intraperitoneally 3 times a week with the IL-2/Jes6-1 complex or PBS vehicle alone (see Methods). IL-2 treatment substantially improves cardiac recovery after MI as shown by echocardiography (B) and the significant difference in % ejection fraction (%EF) between the 2 groups of mice by the primary endpoint at Day 28, despite no difference in infarct size (C) or collagen deposition (D). Mann-Whitney U test or 2-way analysis of variance where appropriate. Abbreviations as in Figures 1 and 4.

Figure 7. Low-Dose IL-2 Activates ILC2s in Patients With ACS PBMC culture with IL-2 (10 ng/mL) show rapid downregulation of ILC2 surface markers following IL-2, IL-33, and in combination (A). Secretion of IL-5 was increased (B). Patients with ACS enrolled in LILACS (C) were sampled at Visit 2 (Baseline) before receiving 5 daily consecutive IL-2 doses (1.5 U or 2.5 U) or placebo and follow-up before blood sampling on Visit 7 (Follow-up 1, Day 6) and Visit 8 (Follow-up 2, Day 13). (D) Flow cytometric analysis shows a dose-dependent decrease in canonical markers of peripheral blood ILC2 (CRTH2+CD161+ cells within Lin−CD127+ cells) over the course of the treatment, suggesting activation. Corresponding amounts of serum IL-5 (E) and blood eosinophil count (F) were both elevated at follow-up 1 and subsequently decreased after a further 6 days. Error bars show ± SEM. Multiple comparisons were made using the Kruskal-Wallis test followed by Mann-Whitney U test. ACS = acute coronary syndrome; PBMC = peripheral blood mononuclear cell; other abbreviations as in Figures 1 and 4.

Central Illustration. Type 2 Innate Lymphoid Cells Orchestrate a Cardioprotective Response After Myocardial Infarction In this report, we demonstrate that type 2 innate lymphoid cells (ILC2s) are resident in the heart and pericardium, expanding during experimental myocardial infarction. In mouse models deficient in ILC2s, cardiac recovery and repair are significantly retarded; the recovering scar contains less collagen, fewer T regulatory cells (Tregs) and more inflammatory monocyte/macrophage populations. Supplementing the mice with interleukin (IL)-2 increases ILC2 activation and promotes the secretion of cardioprotective cytokines (eg, IL-5) and ultimately improves recovery. We observe a similar phenotypic shift in ILC2s in patients with acute coronary syndromes after administration of low-dose IL-2, coupled with increased IL-5 secretion and associated eosinophilia.