Comparison of the effects of metformin and empagliflozin on cardiac function in heart failure with preserved ejection fraction mice

Figures

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  Figure 1. Construction of hFpEF mouse model. (A) Experimental design. C57BL/6J mice were maintained on different dietary regimens (filled triangle) and followed up to the eighth week (empty red triangles). (B) The representative LV M-mode echocardiographic tracings. (C) Percent LV ejection fraction (LVEF%). (E) The representative pulsed-wave Doppler (top) and tissue Doppler (bottom) tracings. (D) Ratio between mitral E wave and A wave(E/A). (F) Ratio between mitral E wave and E’ wave (E/E’). (G) Lung tissue was stained with H&E, where blue arrows indicate massive red blood cell infiltration (Control group: n = 5; Experimental group: n = 4).

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  Figure 2. Role of EMPA and MET in body weight and blood pressure in hFpEF mice. (A) Experimental design. C57BL/6J mice were maintained on dietary regimens to the eighth week, and then EMPA and EMT were given intragastric administration respectively for continuous 4 weeks, followed up to the twelveth week (empty red triangles). The mice only given a same volume of 0.5% CMC-Na as vehicle group. (B) Body weight changes in HFpEF mice following EMPA (10 mg/kg/day) or MET (200 mg/kg/day) treatment vs. Vehicle. (C) Blood pressure changes (SBP and DBP) in HFpEF mice following EMPA or MET treatment vs. Vehicle (Vehicle group: n = 7; EMPA group: n = 7; EMT group: n = 8).

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  Figure 3. Role of EMPA and MET in glucose metabolism in hFpEF mice. (A) Blood glucose changes and Area under curve (AUC) in glucose tolerance test (GTT) between EMPA group and Vehicle group. (B) Blood glucose changes and AUC in Insulin tolerance testing (ITT) between EMPA group and Vehicle group. Vehicle group: n = 7; EMPA group: n = 7; MET group: n = 8.

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  Figure 4. Role of EMPA and MET in cardiac function and exercise tolerance in hFpEF mice. (A) The representative LV M-mode echocardiographic tracings. (B) Percent LV ejection fraction (LVEF%). (C) The representative pulsed-wave Doppler (top) and tissue Doppler (bottom) tracings. (D) Ratio of E/A. (E) Ratio of E/E’. (F) Lung tissue was stained with H&E. (G) The whole lung was weighed as wet after using gauze to remove residual blood and fluid from the lung surface. Subsequently, the tissues were subjected to a drying process in an oven at 80°C for 48 h until a constant weight was achieved, after which the dry weight was measured to determine the W/D ratio. (H) Exercise exhaustion test in each group. (I) The levels of plasma NT-proBNP. Vehicle group: n = 7; EMPA group: n = 7; MET group: n = 8.

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  Figure 5. Role of EMPA and MET in cardiac structure in hFpEF mice. (A) The representative gross view of heart from each group. (B) The heart weight (HW) and tibial length (TL) were quantified, and the ratio of HW to TL was subsequently computed. (C) The representative images of H&E and Masson staining of Heart. (D) The proportion of collagen fiber area was calculated. (E) The representative images of WGA staining of Heart. (F) The cross sectional area (CSA) of cardiomyocyte analyzed by Image J Pro software. Vehicle group: n = 7; EMPA group: n = 7; MET group: n = 8.

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  Figure 6. Pharmacological mechanism of EMPA and MET in the treatment of heart failure explored through network pharmacology analysis. (A) Venn diagram of the intersection targets between EMPA, MET, and heart failure with preserved ejection fraction. (B) Protein-protein interaction network of the intersection targets analyzed using the STRING database. (C) Five key core genes of EMPA and MET acting on heart failure, screened using the CytoHubba plugin and MCC algorithm in Cytoscape software. (D) GO analysis of the intersection targets performed using R packages (clusterProfiler, enrichplot, and ggplot2).

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  Figure 7. Expression analysis of Key genes and proteins in heart failure models treated with EMPA and MET. (A–E) Relative mRNA expression levels of Hsp90aa1 (A), Il1b (B), Map2k1 (C), Nfkb1 (D), and Prkaca (E) in heart tissue from vehicle, EMPA, and MET treated groups. Data are presented as the mean ± SEM. (F) Representative Western blot images showing the protein expression levels of HSP90, TGFβ, and β-actin in heart tissue from vehicle, EMPA, and MET treated groups. (G–H) Quantification of Western blot results. Protein levels of HSP90 (G) and TGFβ (H) were normalized to β-actin. Data are presented as the mean ± SEM (Vehicle group: n = 7; EMPA group: n = 7; MET group: n = 8).