Quercetin improves myocardial ischemia-reperfusion injury by regulating macrophage M2 polarization through Bcl-2/Beclin-1 complex

Abstract

Background: Myocardial ischemia-reperfusion injury (MIRI) is a common pathological phenomenon during the treatment of acute myocardial infarction. Recent studies suggest that macrophage polarization plays a crucial role in MIRI progression. However, whether quercetin mitigates MIRI by modulating macrophage polarization and the underlying molecular mechanisms remain unclear.

Methods: The protective effects of quercetin against MIRI were assessed using TTC-Evans blue staining, echocardiography, and myocardial enzyme assays. Histological changes, including myocardial fibrosis, were evaluated via HE and Masson staining. Western blot, qPCR, and immunofluorescence were performed to analyze macrophage M1/M2 polarization. Additionally, co-immunoprecipitation (Co-IP) assays were conducted to determine whether quercetin regulates M2 polarization through autophagy modulation.

Results: Quercetin significantly reduced infarct size, improved cardiac function, and alleviated inflammation and myocardial fibrosis in a dose-dependent manner. Western blot and immunofluorescence analyses showed that quercetin downregulated M1 markers while upregulating M2 markers and enhancing IL-10 secretion. In vitro experiments further confirmed that quercetin promoted M2 macrophage polarization under H/R conditions, thereby attenuating cardiomyocyte injury through a paracrine mechanism. Mechanistically, quercetin facilitated autophagic flux by reducing the binding affinity between Bcl-2 and Beclin-1, leading to enhanced M2 macrophage polarization, an effect partially reversed by 3-MA.

Conclusion: This study provides the first evidence that quercetin exerts cardioprotective effects in MIRI by promoting M2 macrophage polarization. Furthermore, it elucidates a novel molecular mechanism in which quercetin regulates autophagy to drive M2 polarization, offering experimental support for quercetin as a potential therapeutic strategy for MIRI.

Keywords: Autophagy; Macrophage polarization; Myocardial ischemia–reperfusion injury; Quercetin.

Fig. 1

Quercetin alleviates cardiac dysfunction in MIRI rats. A Serum biochemical analysis of CK-MB and LDH levels. B ELISA analysis of inflammatory cytokines IL-1β, TNF-α, and IL-10. C Evans Blue and TTC double staining used to assess myocardial infarct size in each group (n = 5). The area at risk (AAR) is shown in red, while infarcted regions appear white. Bar graph represents infarct size as a percentage of AAR. D Echocardiography was used to evaluate cardiac function by measuring left ventricular ejection fraction (LVEF) and fractional shortening (LVFS) in each group. Each group included n = 5 rats. Data are expressed as mean ± SD. No mortality occurred during the experiment. Sham: Sham-operated control group with no LAD ligation. MIRI: Myocardial ischemia–reperfusion injury group. MIRI + Que-L: MIRI with low-dose quercetin (50 mg/kg) treatment. MIRI + Que-H: MIRI with high-dose quercetin (100 mg/kg) treatment. MIRI + Lip-Clo: MIRI with liposomal clodronate for macrophage depletion. MIRI + Lip-Clo + Que: MIRI with combined liposomal clodronate and quercetin treatment. *p < 0.05; **p < 0.01

Fig. 2

Quercetin attenuates cardiac inflammation and fibrosis in MIRI rats. A H&E staining showing morphological changes in myocardial tissue across groups. B Masson’s trichrome staining evaluating collagen deposition and fibrosis. Tissue sections were 5 μm thick; images are representative of 3 fields per rat (N = 5 per group). Scale bar = 20 μm; magnification =  × 40. Data are presented as mean ± SD.

Fig. 3

Quercetin promotes M2 macrophage polarization in MIRI rats. A Western blot and qPCR results for M1 marker iNOS and M2 marker Arg-1 in each group. B Immunofluorescence staining indicating CD86 and CD206 expression levels. N = 5 rats per group. Scale bar = 20 μm. Data shown as mean ± SD; *p < 0.05; **p < 0.01

Fig. 4

Quercetin induces M2 polarization of macrophages in vitro. A Flow cytometry analysis of M1/M2 polarization. B Western blot detection of iNOS and Arg-1 expression across groups. C Immunofluorescence staining showing changes in CD86 and CD206 expression. N = 3 independent replicates per group. Scale bar = 20 μm. Representative images and dot plots shown. Data are mean ± SD. Control: Untreated baseline group. LPS/IFN-γ: M1-polarizing stimuli (LPS and IFN-γ). IL-4: M2-polarizing stimulus (IL-4). M1: Untreated M1-polarized macrophage group. H/R: Hypoxia/reoxygenation injury group. H/R + Que-L: H/R plus low-dose quercetin. H/R + Que-H: H/R plus high-dose quercetin

Fig. 5

Quercetin mitigates H/R-induced myocardial injury by promoting M2 macrophage polarization. A Flow cytometry analysis of cardiomyocyte apoptosis across treatment groups. B–E qPCR analysis of TNF-α, LDH, IL-1β, and IL-10 expression. N = 3 independent experiments per group. Data are expressed as mean ± SD. Control: Normoxic control group. H/R: Hypoxia/reoxygenation injury group. H/R + Que-L: H/R with low-dose quercetin. H/R + Que-H: H/R with high-dose quercetin. H/R + M-Que-L: H/R plus macrophage-conditioned medium from low-dose quercetin treatment. H/R + M-Que-H: H/R plus macrophage-conditioned medium from high-dose quercetin treatment. *p < 0.05; **p < 0.01

Fig. 6

Quercetin enhances autophagy and promotes M2 polarization by disrupting Bcl-2/Beclin-1 interaction. A Western blot analysis of autophagy-related proteins and macrophage polarization markers. B Co-immunoprecipitation results demonstrating that quercetin reduces Bcl-2 binding to Beclin-1. N = 3 independent experiments. Data shown as mean ± SD. M1 macrophages: Control group of M1-polarized macrophages. H/R Hypoxia/reoxygenation group. Que Quercetin treatment group. 3-MA Autophagy inhibitor (3-methyladenine) treatment group