Sodium–Glucose Co-Transporter 2 Inhibition With Empagliflozin Improves Cardiac Function After Cardiac Arrest in Rats by Enhancing Mitochondrial Energy Metabolism

Figures

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  FIGURE 1. Experimental procedures in VF-induced cardiac arrest model. VF ventricular fibrillation, CA cardiac arrest, CPR cardiopulmonary resuscitation, ROSC return of spontaneous circulation, EMP empagliflozin.

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  FIGURE 2. Empagliflozin (EMP) improved cardiac function after cardiac arrest in rats. (A) Effect of EMP on heart rate within 1 h after return of spontaneous circulation (ROSC) (n = 7–10). (B) Effect of EMP on mean artery pressure (MAP) within 1 h after ROSC (n = 7–10). (C) EMP increased left ventricular ejection fraction at 24 h after ROSC (n = 6). (D) EMP increased cardiac output at 24 h after ROSC (n = 6). (E) EMP improved survival of the rats after cardiac arrest within 24 h (n = 10). Data are presented as mean ± SD, *p < 0.05, **p < 0.01.

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  FIGURE 3. Empagliflozin (EMP) decreased myocardial injury after cardiac arrest in rats. (A) Representative photographs of Masson staining of heart sections obtained 24 h after return of spontaneous circulation (ROSC). Masson staining showed that EMP decreased myocardial fibrosis (stained blue) after ROSC. Scale bar = 50 μm. (B) Quantification of myocardial fibrotic area from Masson stained section. (C) EMP decreased serum cardiac troponin I (cTnI) level at 24 h after ROSC. Data are presented as mean ± SD, n = 6, *p < 0.05, **p < 0.01.

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  FIGURE 4. Empagliflozin (EMP) decreased myocardial oxidative stress after cardiac arrest in rats. (A) Representative photographs of 4-hydroxynonenal (4-HNE) and 8-hydroxy-2′-deoxyguanosine (8-OHdG) immunostaining of heart sections obtained 24 h after return of spontaneous circulation (ROSC). Scale bar = 50 μm. (B) Quantification of 4-HNE staining levels from stained heart sections. (C) Quantification of 8-OHdG staining levels from stained heart sections. (D) The relative reactive oxygen species (ROS) levels in heart tissues obtained 24 h ROSC. Data are presented as mean ± SD, n = 6, *p < 0.05, **p < 0.01.

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  FIGURE 5. Effect of empagliflozin (EMP) on structural integrity of the myocardial mitochondria after cardiac arrest in rats. (A) Representative heart transmission electron microscopy (TEM) images of heart mitochondria at 24 h after return of spontaneous circulation (ROSC). Arrow indicates intermitochondrial junctions (IMJs); Arrowhead indicates lipid droplet. Scale bar = 1 μm. (B) Quantification of heart mitochondrial area by TEM after ROSC. n = 5 and at least 60 mitochondria were counted in each group. (C) Quantification of heart mitochondrial lipid droplet content by TEM after ROSC. n = 5 and 30 photographs were counted in each group. (D) Quantification of heart IMJs by TEM after ROSC. n = 5 and 25 photographs were counted in each group. Data are presented as mean ± SD, *p < 0.05, **p < 0.01.

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  FIGURE 6. Empagliflozin (EMP) increased myocardial energy levels and mitochondrial activity after cardiac arrest in rats. (A) EMP increased myocardial ATP levels at 6 and 24 h after return of spontaneous circulation (ROSC). (B) EMP increased the myocardial mitochondrial complex Ⅰ activity at 6 and 24 h after ROSC. (C) EMP increased the myocardial mitochondrial respiratory control ratio (RCR) at 6 and 24 h after ROSC. Data are presented as mean ± SD, n = 6, *p < 0.05, **p < 0.01.

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  FIGURE 7. Empagliflozin (EMP) increased myocardial ketone metabolism after cardiac arrest in rats. (A) EMP decreased serum glucose levels at 6 and 24 h after return of spontaneous circulation (ROSC). (B) EMP decreased serum lactate levels at 6 and 24 h after ROSC. (C) EMP increased the serum β-hydroxybutyrate (BHB) levels at 6 and 24 h after ROSC. (D) EMP increased the myocardial BHB levels at 6 and 24 h after ROSC. (E) EMP increased the myocardial NAD+/NADH levels at 6 and 24 h after ROSC. (F) EMP increased the heart β-hydroxybutyrate dehydrogenase (BDH1) protein expression at 24 h after ROSC. GAPDH was used as the loading control. Data are presented as mean ± SD, n = 3 or 6, *p < 0.05, **p < 0.01.