Empagliflozin attenuates adverse outcomes of acute ischemic heart failure by targeting NLRP3/IL1β mediated macrophage polarization and inflammatory response

Background The role of SGLT2 inhibitors in acute heart failure and associated mechanisms remain unclear. Objectives To elucidate the impact of empagliflozin on acute ischemic heart failure and its underlying mechanisms. Methods Left anterior descending coronary artery of male C57BL6/J mice was ligated permanently to induce myocardial infarction (MI). Empagliflozin (10 mg/kg) or isotype controls were treated by daily oral gavage 3 days before operation until sacrificed 3 weeks after MI. Echocardiography was performed to assess cardiac function. Myocardial hypertrophy and interstitial fibrosis were respectively analyzed by WGA and Masson-trichrome staining. Immunofluorescent co-staining of Mac3 and IL1β was employed to examine the percentage of IL1β positive macrophages in the infarct myocardium. The recruitment of immune cells into the infarct myocardium at acute phase post-MI (Day 0,3,7,14) was determined by flow cytometry. RNA-seq was performed on CCR2+ macrophages to compare the gene expression profiles between empagliflozin and isotype groups. Differentially expressed genes and their effects on related signaling pathways were evaluated by GO and KEGG enrichment analyses. In vitro, macrophages were treated with LPS (10ng/ml) and interferon-γ (2ng/ml) for pro-inflammatory activation, follow by treating ATP (5μM) for 30min for NLRP3 inflammasome activation. Pro-inflammatory gene and protein expression of were analyzed by qPCR and western blot. Plasma BNP and supernatant secreted IL-1β levels were detected by ELISA. Results Empagliflozin significantly improved cardiac remodeling and cardiac function in ischemic HF model, as demonstrated by significant reduction of plasma BNP levels, cardiac hypertrophy, and myocardial fibrosis, and recovery of cardiac function (EF, LVSV, and LVDV). FCM analysis showed that empagliflozin significantly inhibited the recruitment of Ly6Chi monocytes into the infarct myocardium and its subsequent polarization into CCR2+ macrophages at the early MI phase. Immunofluorescent double staining showed that the percentage of IL1b+ macrophages was also reduced by empagliflozin. Transcriptomic analysis revealed that empagliflozin up-regulate CCR2+ macrophage Cytokine-cytokine receptor interaction pathway and down-regulate lysosome pathway. Mechanistically, empagliflozin significantly attenuated the activation of the NLRP3 inflammasome, and reduced the expression of Il1β and its’ downstream inflammatory cytokines (Il6, Tnfα, etc.) and chemokines (Ccl2) in macrophages ex vivo. Conclusions Empagliflozin can attenuate the adverse outcomes of acute ischemic heart failure, in part by inhibiting the over-activation of Nlrp3 inflammasome, which in turn modulates macrophage polarization and promotes the inflammation resolution in early phase post injury. This process likely involves the interaction of cytokines and their corresponding receptors.