Liraglutide attenuates autoimmune myocarditis by inhibiting NLRP3 and NF-κb pathways

Figures

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  Fig. 1. liraglutide attenuated the pathogenicity of myocarditis. (A) Heart weight/body weight (HW/BW) ratios for each group. (B) ELISA analysis of serum cTnT levels in the indicated groups. (C) Representative HE-stained images (left) and quantification of inflammation area (right) in ventricular sections. Scale bar = 200 μm–50 μm. (D) Representative Masson-stained images (left) and quantification of fibrotic area (right) in ventricular sections. Scale bar = 200 μm–50 μm. N = 8. *P < 0.05, **P < 0.01, ****P < 0.0001. Lira: liraglutide.

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  Fig. 2. Liraglutide improves ventricular ejection function in mice with myocarditis. (A) Representative echocardiography images. (B and C) Left ventricular ejection fraction (EF) and fractional shortening (FS) measured by echocardiography in each group. N = 8. *P < 0.05, **P < 0.01. Lira: liraglutide.

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  Fig. 3. Liraglutide reduces T cell and macrophage infiltration. Representative immunofluorescence images for CD3 (A) and F4/80 (B) and their quantification per high-power field (HPF, average of 3–6 different 200x visual fields) in the hearts of each group (n = 6); scale bar, 50 μm.N = 6. **P < 0.01, ***P < 0.001, ****P < 0.0001. Lira: liraglutide.

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  Fig. 4. Liraglutide attenuates the expression of inflammatory factors. RT-qPCR analysis for relative mRNA expression of Il1b, Il6, Tnf, Ifng, Il17a in the heart of each group. N = 8. *P < 0.05, **P < 0.01, ***P < 0.001, or ****P < 0.0001. Lira: liraglutide.

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  Fig. 5. ELISA verified that liraglutide reduced IL-1β, IL6, and TNF-α levels in serum. ELISA analysis of serum IL-1β, IL-6, TNF-α levels in the indicated groups. N = 6. *P < 0.05, ***P < 0.001, or ****P < 0.0001.

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  Fig. 6. RNA-Seq reveals the mechanism by which liraglutide regulates the pathogenesis of myocarditis. (A): Volcano plots showing differentially expressed genes between the EAM + vehicle group and the EAM + liraglutide group. Red dots indicate genes upregulated by liraglutide treatment, while blue dots indicate genes downregulated by liraglutide treatment. (B and C): GO (B) and KEGG (C) enrichment analysis of differentially expressed genes between the EAM + vehicle group and the EAM + liraglutide group. (D, E, and F) Heatmaps showing significantly differentially expressed genes between the EAM + vehicle group and the EAM + liraglutide group in the cytokine-cytokine receptor interaction pathway (D), the NOD-like receptor signaling pathway (E), and the NF-κB signaling pathway (F). N = 4 per group. The KEGG database is available at www.kegg.jp/kegg/kegg1.html.

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  Fig. 7. Liraglutide attenuates the expression of NF-κB and NLRP3 signaling pathway. (A) RT-qPCR analysis for relative mRNA expression of Socs3, Socs1, Csfrb2, Osm, Nlrp3 and Il18 in the heart of each group. (B) Western blot analysis for relative protein levels of NF-κB (p-p65/p65) and NLRP3, and normalize to GAPDH. N = 5. *P < 0.05, **P < 0.01, ***P < 0.001, or ****P < 0.0001. Lira: liraglutide.