JPET: Empagliflozin

Empagliflozin, an SGLT2 Inhibitor, Reduced the
Mortality Rate after Acute Myocardial Infarction
with Modification of Cardiac Metabolomes and
Antioxidants in Diabetic Rats.
Saturday, 18 April 2020 1
JournalClub
HirotoOshimaet.al.
Department of Cardiovascular, Renal & Metabolic Medicine,
Sapporo Medical University & School of Medicine, Japan.
The Journal of Pharmacology & Experimental
Therapeutics.
✩ Volume 368 ✩ Number 3
✩ March 2019 ✩ Pg. 524-534
The American Society for Pharmacology &
ExperimentalTherapeutics.

Introduction
EMPAGLIFLOZIN: SGLT2 Inhibitor, FDA approved in 2014.
Mechanism of Action:

How do SGLT2 inhibitors prevent HF and death?
 Ferrannini & Mudaliar et al. (2016) stated Myocardial Fuel Hypothesis.
 SGLT2 inhibitors increase blood OHB levels.
 HF leads to ↑ reactive oxygen species in myocardium.
 Ketone bodies are more energy efficient than FFAs.
 Ketone oxidation is ↑ in failing heart & ↓ ketone utilization by
cardiomyocyte-specific knockout of SCOT, resulted in adverse cardiac
remodeling after pressure overload.
 OHB shown to suppress oxidative stress by inhibition of class I
histone deacetylases.
 Benefits of chronic elevation of OHB is uncertain.
 Association of  ketone body with AE has been reported.
 Mizuno et al. (2017), Level of ketone uptake was positively
correlated with plasma BNP level.
 Obokata et al. (2017),  serum OHB level was associated with
CVS events in pts. with renal failure.

Materials & Methods
Protocol 1: Post-MI Survival Rate Analysis
 Used: Male OLETF and LETO rats.
 Age: 25-30 weeks.
 Rats were pretreated for 2 weeks with:
 empagliflozin (10 mg/kg per day) or vehicle
 Administered S.C. via osmotic minipumps.
Protocol 2: Post-MI Survival Rate Analysis
 Examine effects of exogenous OHB on mortality.
 S.C. administered OLETF for 7 days before MI
 OHB (8.0 mmol/kg per day) or
 vehicle (dimethyl sulfoxide and polyethylene glycol)
 After 12 hours of fasting, MI was induced as in protocol 1.
Otsuka Long-Evans Tokushima Fatty Rats were selected as:
1. OLETF rats spontaneously develop DM.
2. Ventricular dysfunction is similar to that in humans.
3. Mortality rate after MI is significantly higher.

Fig. 1. Experimental protocols for survival study & cardiac tissue sampling in Empagliflozin (A) & OHB (B) experiments.

Results
Metabolic and Hemodynamic Profiles:
Table 1. Metabolic, hemodynamic, and echocardiographic parameters before induction of MI
N = 17–27; *P<0.05 vs. LETO; †P<0.05 vs. LETO + EMPA; ‡P<0.05 vs. OLETF.
Table 2. Metabolic parameters at 12 hours after MI (before tissue
sampling) N = 11–16; *P<0.05 vs. LETO; †P<0.05 vs. OLETF.
Table 3. Metabolic and hemodynamic
parameters before MI in ketone infusion Study;
N = 13–15, *P<0.05 vs. vehicle.

Survival Rate after MI: Myocardial Levels of OHB, ATP & BNP: (Fig. 2D & 2E)
Fig. 2. Effects of empagliflozin on survival rate, myocardial ATP, and BNP after MI.

Expression of Genes that Regulate Glucose Metabolism, -Oxidation & Ketone Oxidation:
Fig. 3. Expression of molecules regulating energy substrate metabolism.

Metabolome Analyses:
Fig. 4. Summary of changes in metabolomics profile.
Metabolites of energy production pathway [glycolysis, ketone, lipid oxidation, TCA cycle] & pentose phosphate pathway.

Antioxidant Proteins and Reactive Oxygen Species Production:
Fig. 5. Effects of empagliflozin on lipid peroxidation and
antioxidant proteins.
Fig. 6. Effects of OHB on lipid peroxidation and
antioxidant proteins.

Discussion
 Rx for 2 weeks significantly improved the survival rate after acute MI.
 ↑ mortality is due to HF but not lethal arrhythmia.
 Congestion of Lung but not cardiac rupture.
 BNP at 12 hours after MI was slightly suppressed.
 Thus, suppression of HF is most likely explanation.
 Unlike DM, beneficial effect on post-MI mortality was observed without
a significant change in:
 body weight
 blood pressure or
 heart rate in OLETF rats.
 Changes in patterns of myocardial metabolites are consistent with
notion that empagliflozin:
 ↑ glucose oxidation
 ↑ ketone oxidation
 ↓ fatty acid oxidation leading to maintenance of ATP.

 Empagliflozin causes suppression of oxidative stress by:
 ↑ expression of antioxidant stress proteins
 ↓ biomarkers of oxidant stress
 βOHB partially mimicked the protective effects on:
 Mortality
 ATP level and
 Oxidative stress after MI in OLETF rats.
 Empagliflozin ↑ expression of antioxidant enzymes by:
 ↑ SOD2 level
 Restored catalase level
 Significantly ↑ Sirt3 protein level.
 Infusion of OHB upregulates Sirt3 expression:
 mechanism of which is unclear.
 Empagliflozin effects are distinct from PPAR activators & Insulin, like
 ↑ utilization of ketones
 suppression of myocardial oxidant stress

Limitations
1.Metabolome analysis does not allow direct measurement of the flux in
metabolic pathways. Hence, assessment of cardiac energy metabolism
made is interpretive.
2.Infusion of exogenous βOHB mimicked effect of empagliflozin on Sirt3
and antioxidant proteins, but its effect on myocardial metabolomes was
not analysed.
3.Whether the cardioprotective effect is dose dependent and/or is
glycaemic control dependent remained unclear.

Conclusion
 Empagliflozin significantly attenuated DM-induced increase in acute
mortality after MI in Type 2 DM model.
 Protective effects of an SGLT2 inhibitor were:
 Associated with
 Improved energy metabolism.
 Upregulation of antioxidative proteins.
 Independent of
 Hemodynamic changes.
 Haematopoiesis.
 Changes in cardiac metabolism & antioxidants and their role in
protection afforded warrants further investigation.

Thank YouPrepared by:
Dr. Bhargav Darji
2nd Year Resident
Pharmacology Dept.
GMC, Surat.
Fig. 7. Possible mechanisms of cardiac protection by SGLT2 inhibitors.

JPET: Empagliflozin

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