EMPA Group Research Articles

Cardamom Essential Oil-Loaded Nanostructured Lipid Carrier Reduces Obesity and Mitigates Diabetes Mellitus in Rats Fed with High Sugar/High Fat Diet

Background: Obesity is a chronic disease and one of the characteristic features of metabolic syndrome that is associated with severe health problems such as diabetes, hypertension, and cardiovascular diseases. Objective: To assess the effectiveness of the Cardamom Essential Oil-Loaded Nanostructured Lipid Carrier) CEO-NLC) as a potential therapeutic agent to lower blood sugar and an anti-inflammatory, antioxidant, and weight-reducing agent. Methods: A total of 24 male Wistar rats, aged 8–10 weeks and weighing 200–410 g, were used in this study. The rats were divided into four groups: negative control (G1), diabetic control (G2), treatment group (CEO-NLC) (G3), and positive control (Empagliflozin; EMPA) (G4). Diabetes was induced in all groups by injection of Streptozotocin (50 mg/kg) except G1. Then, animals were treated consecutively for four weeks. On day 29, they were sacrificed, and blood/tissue samples were collected for serological and histopathological analysis, respectively. Results: Significant decrease in blood sugar levels on day 28 for both the CEO-NLC group and EMPA versus diabetic control group were seen (250 mg in CEO-NLC, and 300 mg in EMPA, vs 450 mg in the diabetic control group). CEO-NLC achieved a significant effect as a weight-losing agent (p = 0.004). Proinflammatory biomarkers and kidney and liver function tests were enhanced after CEO-NLC treatment. Additionally, substantial histopathological modifications in the CEO-NLC group were also seen compared to the diabetic group and EMPA group. Conclusions: CEO-NLC has potential activity in lowering blood sugar levels, aiding weight loss, and eventually, organ protection ability.

Empagliflozin reduces the adverse effects of diabetes mellitus on testicular tissue in type 2 diabetic Rats: A stereological and biochemical study

Empagliflozin as an antioxidant decreases blood glucose and insulin resistance in type 2 diabetes mellitus. Base on the empagliflozin antioxidant properties we decided to investigate the its effects on the testis histological changes through stereological techniques and biochemical evaluations in T2 diabetes mellitus rats. Rats were divided into: control, diabetes mellitus (DM, streptozotocin + nicotinamide) and diabetes mellitus + empagliflozin (DM + EMPA, 10 mg/kg/day) groups. 56 days after inducing diabetes mellitus testis histological changes and serum biochemical factors along with the level of Bax, Bcl2 and Nrf2 genes expression in the testicular tissue were assessed. A significant decrease in the mean total volume of testis and its components, the level of Bcl2 and Nrf2 gene expression (p < 0.001) along with a significant increase in the level of IL-6, TNF-α, MDA, Bax gene expression were observed in the DM group compared to the control group (p < 0.001). In the DM + EMPA group, the mean total volume of testis and its components, the level of Bcl2 gene expression (p< 0.01) and Nrf2 (p < 0.001) significantly increased whereas the mean level of IL-6 (p < 0.01), TNF-α (p < 0.001), MDA (p < 0.001), Bax (p < 0.001) gene expression significantly decreased compared to the DM group. Our results showed that empagliflozin, by improving the antioxidant defense system, can reduce testicular inflammation and apoptosis and partly prevent the adverse effects of diabetes mellitus on testicular tissue.

Effect of Semaglutide and Empagliflozin on Pulmonary Structure and Proteomics in Obese Mice.

This study utilized proteomics to investigate changes in protein expression associated with lung health in obese mice exposed to semaglutide and empagliflozin through a high-fat diet. Twenty-eight male C57BL/6JC mice were randomly assigned to two groups: a control diet group (n = 7) and a high-fat diet group (n = 21). The HFD group was further divided into three groups: HFD group (n = 7), Sema group (n = 7), and Empa group (n = 7). Post-treatment, mice underwent assessments including glucose tolerance, lipids, oxidative stress markers, body weight, lung weight, and structure. Proteomics identified differentially expressed proteins (DEPs) in lung tissue, and bioinformatics analyzed the biological processes and functions of these proteins. Semaglutide and empagliflozin significantly attenuated obesity-induced hyperglycemia, abnormal lipid metabolism, oxidative stress response, and can decrease alveolar wall thickness, enlarge alveolar lumen, and reduce collagen content in lung tissue. Both medications also attenuated lung elastic fibre cracking and disintegration. In the HFD/NCD group, there were 66 DEPs, comprising 30 proteins that were increased and 36 that were decreased. Twenty-three DEPs overlapped between Sema/HFD and Empa/HFD, with 11 up-regulated and 12 down-regulated simultaneously. After analysing DEPs in different groups, four proteins - LYVE1, BRAF, RGCC, and CHMP5 - were all downregulated in the HFD group and upregulated by semaglutide and empagliflozin treatment. This study demonstrates that obesity induced by a high-fat diet causes a reduction in the expression of LYVE1, BRAF, RGCC, and CHMP5 proteins, potentially affecting lung function and structure in mice. Significantly, the administration of semaglutide and empagliflozin elevates the levels of these proteins, potentially offering therapeutic benefits against lung injury caused by obesity. Merging semaglutide with empagliflozin may exert a more pronounced impact.

Empagliflozin restores the deregulated cardiac endothelial cell transcriptome in vivo upon myocardial ischemia reperfusion injury and reduces the infiltration of immune cells

Abstract Background/Introduction Empagliflozin (EMPA) is a sodium-glucose co-transporter 2 (SGLT-2) inhibitor with established cardioprotective effects in heart failure but its use in myocardial ischemia-reperfusion (IR) injury is still under investigation. We have shown that chronic administration of EMPA reduces infarct size in normoglycemic mice subjected to IR and that endothelial cells (ECs) are implicated in the protective mechanism. Purpose We aimed to determine EMPA’s distinct effect on three cell populations: cardiac ECs, fibroblasts (FBs) and cardiomyocytes (CMs) in vivo. Methods C57Bl6 male mice were randomized as follows: 1) Control-Sham, 2) Control-IR and 3) EMPA-IR. EMPA group received EMPA 10mg/kg/day for 6 weeks which is a clinically relevant dose and Control groups received vehicle (5% DMSO in normal saline). At the end of the administrations, mice were subjected to 30’I/2hR or sham operation and the ischemic heart was collected to isolate CMs. From the non-CMs populations the cardiac ECs and FBs were sorted. RNA was obtained from the distinct cell populations and 3’ RNA sequencing was applied to investigate the pathways affected by IR and EMPA treatment. An additional cohort of mice was employed to validate the results at mRNA level. In order to examine the effects of EMPA upon prolonged reperfusion, we repeated the experimental protocol, and the mice were sacrificed upon 7 days of R. Based on the RNA sequencing data, the infiltration of the heart with immune cells was examined. Results Comparison between IR and Sham groups revealed that IR alters the transcriptomic profile of ECs and FBs with 576 and 571 genes respectively being significantly deregulated (FDR&amp;lt;0.05). CMs transcriptome was not altered by IR. The comparison between Control-IR and EMPA-IR groups revealed that EMPA does not affect the FBs’ gene expression profile while in CMs, EMPA affects only 3 genes expression (FDR&amp;lt;0.05) related to mitochondrial metabolism. The ECs are mostly affected by EMPA treatment with 211 genes being significantly altered (FDR&amp;lt;0.05). Pathway enrichment analysis in ECs revealed that the deregulated pathways by IR related to extracellular matrix organization, matrix degradation and immune cell adhesion are restored by EMPA treatment. The expression levels of several significant genes of our dataset including Mmp-2, Timp-1, Icam, Vecam, etc was validated in the independent cohort of mice. On the 7th day of R, EMPA reduced the infiltration of neutrophils and inflammatory (Ly6C high) monocytes in the myocardium compared to the Control group indicating that EMPA may affect the remodeling process upon IR. Conclusions We identified ECs and FBs as cell target populations for therapeutic interventions at 2h of R. EMPA pre-treatment reverses the deregulated transcriptome related to extracellular matrix organization, matrix degradation and immune cell adhesion in ECs and eventually reduces the infiltration of immune cells in the ischemic myocardium.

Utility of residual SYNTAX score to predict clinical outcome after acute myocardial infarction

Abstract Aims Acute myocardial infarction (AMI) represents a huge health burden and a growing economical challenge, particularly with recurrent major adverse cardiovascular events (MACE). Large attention is critically mandated to identify population at higher risk for recurrent MACE to maximize secondary preventive measures. This study aimed to evaluate the utility of residual SYNTAX Score (RSS) after percutaneous coronary intervention (PCI) to the culprit vessel(s) in patients with AMI (STEMI or NSTEMI) to predict 6-months clinical outcomes. Methods and results A total of 492 eligible AMI patients were recruited through a period of 12 months. The RSS was systematically calculated after the PCI procedures. Median RSS was 4 (minimum 0–maximum 47.5). RSS showed excellent association to mid-term clinical outcomes. At cut-off value of RSS 3 had robust prediction for subsequent heart failure (HF), recurrent AMI, unplanned revascularization and the composite of MACE (P &amp;lt; 0.001). A significantly higher RSS was found in those who developed heart failure through the 6-months follow up period compared to those who did not (10 vs. 3, P &amp;lt; 0.001). Similar significant difference was found for recurrent ACS (11.5 vs. 3, P &amp;lt; 0.001), unplanned revascularization (15 vs. 3, P &amp;lt; 0.001) and occurrence of MACE (12 vs. 3, P &amp;lt; 0.001). Conclusions RSS is a sensitive predictor for 6-months adverse clinical outcomes after AMI. Calculating RSS at the end of PCI can guide intensification of secondary preventive measures. Ability of RSS- tailored management to impact clinical outcomes in high-risk patients is to be evaluated.Improvement, with subsequent amelioration of LV diastolic function. The Dapa group had a significantly greater improvement in LVEF at four months and six months compared to the Empa group. While the Empa group showed a significant improvement in HbA1C at six months compared to the Dapa group. Additional Content An author video to accompany this abstract is available on https://academic.oup.com/eurheartjsupp. Please click on the arrow next to ‘More Content’ and then click on ‘Author videos’.

Efficacy and Safety of Empagliflozine and Semaglutide (Once Weekly) in T2DM Patients in Shtip.

Objective: The efficacy and safety of the following new treatment agents were analyzed: once weekly semaglutide (OWSem) and the empagliflozine (Empa). This was done with patients with type 2 diabetes mellitus (T2DM) at the Clinical Hospital in Shtip, R.N. Macedonia. Material and methods: One-hundred-twenty-one diabetic patients were treated for the first time with OWSema or Empa and were retrospectively analyzed. Glycemic control, serum creatinine, decrease in weight, co-morbidities, and hospitalization during treatment were recorded. Results: Among the 61 patients treated with OWSema and 60 patients treated with Empa, there were not any statistically significant differences in age, sex, BMI, duration of diabetes, and a number of patients treated with insulin. Both agents (OWSema and Empa) achieved statistically significant HbA1c reduction after 6, 12, and 18 months (9.2; vs. 7.6; 6.7; 6.6, and 9.3; vs. 7.5; 7.2, 7.5%, respectively) treatment. There were not any differences in the value of creatinine between the visits in both groups. During the period of 2 years, 3 patients (5%) from the Empa group died, all with multiple comorbidities. One patient from Empa group was hospitalized because of acute pulmonary edema and two from the OWSema group because of TIA and acute coronary syndrome. The median decrease in weight was more pronounced in the OWSema group (6.0 vs. 4.0kg). Five patients stopped the treatment with Empa because of a simple urinary infection, and one stopped the OWSema because of GIT intolerance. Eight patients did not tolerate the dose of 1mg, and they therefore continued with 0.5mg of OWSema. Conclusion: Once weekly treatment with semaglutide and empagliflozine achieves a great reduction in HbA1c, and as such are safe for treatment of T2DM.

The Impact of an SGLT2 Inhibitor versus Ursodeoxycholic Acid on Liver Steatosis in Diabetic Patients.

Non-alcoholic fatty liver disease (NAFLD) is related to metabolic syndrome via insulin resistance, where preventing disease progression is crucial in the management process. The study included 240 NAFLD patients with type 2 diabetes who were randomly allocated into empagliflozin 25 mg (EMPA group), ursodeoxycholic acid 250 mg (UDCA group), or the control group (placebo). The study outcomes included: changes in liver fat content (LFC; %) (utilizing the Dixon-based MRI-PDFF approach), liver enzymes, lipid and glycemic profiles, FIB-4 index, and non-alcoholic fatty liver score (NFS). All endpoints were assessed at baseline and after 6 months. EMPA outperformed UDCA and placebo in decreasing LFC (-8.73% vs. -5.71% vs. -1.99%; p &lt; 0.0001). In post-treatment ultrasound images and MRI-PDFF calculations, more patients had normal fatty liver grade (no steatosis or LFC &lt; 6.5%) with EMPA compared to UDCA. EMPA and UDCA showed significant regression in the FIB-4 index (-0.34 vs. -0.55; p = 0.011) and NFS scores (-1.00 vs. -1.11; p = 0.392), respectively. UDCA achieved higher reductions in insulin resistance than EMPA (p = 0.03); however, only EMPA significantly increased beta-cell function (54.20; p = 0.03). When exploring the differences between the two drugs, EMPA was better in decreasing LFC (%), while UDCA achieved higher reductions in liver fibrosis scores. Both showed a similar safety profile in managing liver steatosis.

Abstract 15201: Empagliflozin Improves Baroreflex Sensitivity and Heart Function in Infarcted Rats

Introduction: Several experimental studies have shown improvement in cardiac function of infarcted animals (diabetic or not) when treated with empagliflozin. Likewise, several clinical studies have shown improved outcomes in patients with heart failure with reduced ejection fraction or preserved ejection fraction in patients using empagliflozin. Methods: Wistar rats were divided into 2 groups: AMI - acute myocardial infarction (n=8) and EMPA - acute myocardial infarction + Empagliflozin (n=9).The animals of both groups were anesthetized and submitted to thoracotomy and ligature of the left coronary artery. The animals in the EMPA group received 10 mg/kg of empagliflozin daily through oral gavage for 14 days. After this period, an echocardiographic examination was performed, followed by cannulation of the femoral vein and artery, to evaluate hemodynamic parameters and baroreflex activity. Data were compared using unpaired t test or Mann-Whitney test and considered significant when p&lt;0,05. Results: In the echocardiographic evaluation, an improvement in the systolic function indices was observed in the EMPA group in comparison to the AMI group: SF (27.42% ± 3.117 vs 19.17% ± 0.857), EF (50.69% ± 4.848 vs 37.95% ± 1.528), FAC (38.46% ± 2.382 vs 23.65 ± 1.415), as well as diastolic function indices: E'/A' ratio (1.571 ± 0.156 vs 0.574 ± 0.040) E/E' ratio: (16.24 ± 2.461 vs 28.27 ± 2.788).A reduction in mean arterial pressure was observed in the EMPA group (85.96 mmHg ± 1.437) in comparison to the AMI group (92.21 mmHg ± 2.345).In addition, there was an improvement in the baroreflex mediated tachy and bradycardic responses in the treated animals: tachycardic response (4.957 bpm/mmHg ± 0.125 vs 3.243 bpm/mmHg ± 0.157) and bradycardic response (1.532 bpm/mmHg ± 0.058 vs 0.710 bpm/ mmHg ± 0.017). Conclusions: Empagliflozin treatment in infarcted rats improved left ventricular systolic and diastolic function and improved baroreflex sensitivity.

Clinical Parameters Affecting the Therapeutic Efficacy of SGLT-2-Comparative Effectiveness and Safety of Dapagliflozin and Empagliflozin in Patients with Type 2 Diabetes.

(1) Background. We aimed to assess long-term efficacy and safety in inadequately controlled type 2 diabetes (T2DM) of two SGLT-2 inhibitors: empagliflozin (Empa) and dapagliflozin (Dapa), combined with metformin, other oral antidiabetics or insulin, according to the protocols in Romania. (2) Methods. The data of 100 patients treated for T2DM with associated dyslipidemia and/or cardiovascular diseases at the University Hospital and Consultmed Medical Center in Iasi were retrospectively reviewed (2017–2021). In total, 48 patients had received dapagliflozin (10 mg with oral antidiabetics or insulin) and 52 patients received empagliflozin (10 mg /25 mg with oral antidiabetics). (3) Results. In both groups, the lowering of BMI was significant: Dapa group (32.04 ± 4.49 vs. 31.40 ± 4.18 kg/m2; p = 0.006), and Empa group (34.16 ± 5.08 vs. 33.17 ± 4.99 kg/m2; p = 0.002). Blood sugar average levels decreased significantly (170 vs. 136 mg/dL; p = 0.001 for Dapa; 163 vs. 140 mg/dL; p = 0.002 for Empa) and also average levels of HbA1c (7.90% vs. 7.51%; p = 0,01 for Dapa; 7.72% vs. 7.35%; p = 0.004 for Empa). (4) Conclusions. Better results in all variables were observed in younger male patients with a shorter duration of diabetes and threshold BMI levels of 34.1, treated with SGLT2, and more significantly with Empa.

Empagliflozin significantly prevents QTc prolongation due to amitriptyline intoxication via intracellular calcium regulation

Abstract Background Empagliflozin is a SGLT-2 inhibitor used in the treatment of Type 2 diabetes and has positive effects on cardiovascular outcomes. Amitriptyline can be used in many clinical indications but leads to cardiotoxicity by causing QT prolongation. Aim Our aim in the present study is to observe the effect of the concomitant use of amitriptyline and empagliflozin together, which have an effect on sodium and calcium balance in myocytes, on QTc by using ECG. Materials and methods Twenty-four male Wistar-Albino rats were randomized into four groups. The control group received only serum physiologic (1ml) via orogastric gavage (OG). The EMPA group received empagliflozin (10 mg/kg) via OG. The AMT group received amitriptyline (100 mg/kg) via OG. The AMT+EMPA group (n: 6) received amitriptyline and empagliflozin at same dose. Under anesthesia; QT and QTC intervals were measured at baseline, first, and second hours. Results The differences in QT and QTc values between the AMT group and control group were observed from the 1nd hour (Table 1, Figure 1). It was detected that the measurements of the control group were within normal limits. In the control group, QT was 77.33±9.02 ms at the basal, 73.50±2.26 ms at the 1st hour, 78.17±6.18 ms at the 2nd hour. QTc calculation was 165.42±18.34±10.5 ms at the basal, 166.63±17.92 msat the 1st hour, 184.65±12.86 ms at the 2nd hour. ECG findings of the EMPA group were within normal limits and similar to the control group (Table 1). The durations of QT interval and QTc calculations were found to be statistically longer in the AMT group than the control group at the 1st and 2nd hour (p≤0.001). Empagliflozin significantly ameliorated AMT-induced QT and QTc prolongation. The durations of QT interval were significantly lower at first (p&amp;lt;0,001) and 2nd hours (p&amp;lt;0.01) in the AMT+EMPA group compared to the AMT group. Moreover, QTc calculation was significantly lower in the AMT+EMP group than the AMT group at 1st and 2nd hour (P&amp;lt;0.01) (Table 1). Electrocardiographic comparisons of all groups for one second within the second hour can be seen in Figure 2. Conclusion In the present study, we have detected that empagliflozin significantly ameliorates amitriptyline induced QT prolongation. This effect is probably due to the opposite effects of these two agents in the intracellular calcium balance. It's known that; toxic dose of amitriptyline increases the amount of Sarcoplasmic Ca and the calcium permeability of Ryanodine channels, decreases SERCA-mediated Ca-reuptake by decreasing the calcium binding capacity of calsequestrin, and these leads QTc prolongation. It has been shown that empagliflozin increases Ca reuptake by causing a significant increase in SERCA activity, and decreases Ca sparks by causing inhibition of Ryanodine activity. With more clinical trials, the routine use of empagliflozin may be suggested to prevent QTc prolongation in the diabetic patients receiving amitriptyline. Funding Acknowledgement Type of funding sources: None. Table 1. QT, QTc durations and HR for groupsFigure 1. ECG traces – a: CON, b: EMPA, c: AMT, d: AMT+EMP

Effects of empagliflozin in different phases of diabetes mellitus-related cardiomyopathy: a prospective observational study

BackgroundDiabetes mellitus-related cardiomyopathy (DMCMP), defined as left ventricular (LV) dysfunction caused by hyperglycemia in the absence of coronary artery disease, leads to heart failure (HF). Previous studies have shown that treatment with sodium-glucose co-transporter 2 inhibitor (SGLT2i) reduces the risk of exacerbation of HF. The beneficial effects of SGLT2i on HF depend not only on indirect actions such as osmotic diuresis but also on direct actions on the myocardium, leading to improvements in LV function. However, it remains unclear whether SGLT2i treatment is equally effective in any phase of DMCMP. The aim of this observational study was to compare the efficacy of SGLT2i treatment on LV dysfunction between early and advanced DMCMP.MethodsThirty-five symptomatic non-ischemic HF patients with LV ejection fraction > 40% and type 2 diabetes mellitus (T2DM) treated with empagliflozin (EMPA group) and 20 controls treated without SGLT2i were enrolled. According to the myocardial extracellular volume fraction (ECV), a reliable marker of cardiac fibrosis quantified by cardiac magnetic resonance, the EMPA group was further divided into early DMCMP (n = 16, ECV ≤ 30%) and advanced DMCMP (n = 19, ECV > 30%) groups and followed up prospectively. Echocardiography was performed at baseline and after 12 months. LV function assessed as LV global longitudinal strain (LVGLS) and the ratio of early diastolic mitral inflow velocity to early diastolic mitral annular velocity (E/e′) were compared.ResultsECV was strongly correlated with T2DM duration (r2 = 0.65, p < 0.001). At baseline, each group had a similar background. After 12 months, the EMPA group, especially the early DMCMP group, showed remarkable improvements in LVGLS (ΔLVGLS: 2.9 ± 3.0% (EMPA) vs. 0.6 ± 2.2% (controls), p = 0.005, and 4.6 ± 1.5% (early DMCMP) vs. 1.6 ± 3.3% (advanced DMCMP), p = 0.003) and E/e′ (ΔE/e′: − 1.5 ± 4.7 vs. − 0.3 ± 3.0, p = 0.253, and − 3.4 ± 5.5 vs. − 0.1 ± 3.5, p = 0.043).ConclusionsThe positive effects of empagliflozin on LV dysfunction were more remarkable in early than in advanced DMCMP. Early intervention of SGLT2i for DMCMP may be preferable.

Empagliflozin significantly attenuates sotalol-induced QTc prolongation in rats.

Sotalol is a class III antiarrhythmic drug commonly used in various arrhythmia treatments. However, due to its potent potassium channel inhibition, it can prolong the QT interval and lead to malignant arrhythmias. Empagliflozin is an inhibitor of sodium‑glucose cotransporter 2 (SGLT2) and has a positive effect on cardiovascular outcomes. Since the effect of empagliflozin on the activation of potassium channels is unknown, there is no recommendation regarding the coadministration of these drugs. The study aimed to evaluate the possible protective effects of empagliflozin on sotalol‑induced QT prolongation. We randomized 24 rats into 4 groups. The control group received only physiological saline, the EMPA group, empagliflozin; the SOT group, sotalol; and the EMPA+SOT group, empagliflozin and sotalol. PR and QT intervals and heart rates were measured under anesthesia at baseline and at 1, 2, and 3 hours in lead II. In the SOT group, the QT and QTc intervals as well as T‑wave duration were statistically longer, whereas heart rates were lower than in the control group (P <0.001 for all parameters). Empagliflozin ameliorated sotalol‑induced QT and QTc prolongation in the EMPA+SOT group. The QT interval, T‑wave duration, and QTc interval were shorter, and the heart rate was greater than in the SOT group (P <0.001, P = 0.002, P <0.001, and P <0.001, respectively). Empagliflozin significantly ameliorates sotalol‑induced QT prolongation and could be used safely with sotalol in clinical practice. Future clinical trials might recommend the routine use of empagliflozin to prevent QTc prolongation in diabetic patients receiving sotalol.

Empagliflozin Attenuates Non-Alcoholic Fatty Liver Disease (NAFLD) in High Fat Diet Fed ApoE(-/-) Mice by Activating Autophagy and Reducing ER Stress and Apoptosis.

Aims/hypothesis: SGLT-2 inhibitors (SGLT-2i) have been studied as potential treatments against NAFLD, showing varying beneficial effects. The molecular mechanisms mediating these effects have not been fully clarified. Herein, we investigated the impact of empagliflozin on NAFLD, focusing particularly on ER stress, autophagy and apoptosis. Methods: Five-week old ApoE(-/-) mice were switched from normal to a high-fat diet (HFD). After five weeks, mice were randomly allocated into a control group (HFD + vehicle) and Empa group (HFD + empagliflozin 10 mg/kg/day) for five weeks. At the end of treatment, histomorphometric analysis was performed in liver, mRNA levels of Fasn, Screbp-1, Scd-1, Ppar-γ, Pck-1, Mcp-1, Tnf-α, Il-6, F4/80, Atf4, Elf2α, Chop, Grp78, Grp94, Χbp1, Ire1α, Atf6, mTor, Lc3b, Beclin-1, P62, Bcl-2 and Bax were measured by qRT-PCR, and protein levels of p-EIF2α, EIF2a, CHOP, LC3II, P62, BECLIN-1 and cleaved CASPASE-8 were assessed by immunoblotting. Results: Empagliflozin-treated mice exhibited reduced fasting glucose, total cholesterol and triglyceride serum levels, as well as decreased NAFLD activity score, decreased expression of lipogenic enzymes (Fasn, Screbp-1c and Pck-1) and inflammatory molecules (Mcp-1 and F4/80), compared to the Control group. Empagliflozin significantly decreased the expression of ER stress molecules Grp78, Ire1α, Xbp1, Elf2α, Atf4, Atf6, Chop, P62(Sqstm1) and Grp94; whilst activating autophagy via increased AMPK phosphorylation, decreased mTOR and increased LC3B expression. Finally, empagliflozin increased the Bcl2/Bax ratio and inhibited CASPASE-8 cleavage, reducing liver cell apoptosis. Immunoblotting analysis confirmed the qPCR results. Conclusion: These novel findings indicate that empagliflozin treatment for five weeks attenuates NAFLD progression in ApoE(-/-) mice by promoting autophagy, reducing ER stress and inhibiting hepatic apoptosis.

Effect of empagliflozin as add-on therapy on transtubular potassium concentration gradient in patients with type 2 diabetes hospitalized for acute decompensated heart failure

Abstract Background The transtubular potassium concentration gradient (TTKG) has been reported to be a marker of renal aldosterone bioactivity, and has been shown to be a surrogate of arterial underfilling in patients with acute decompensated heart failure (ADHF). Moreover, high TTKG at discharge has been shown to be associated with poor prognosis in ADHF patients. Empagliflozin, one of the sodium glucose cotransporter 2 inhibitors, has been shown to reduce the risk of cardiovascular mortality in patients with type 2 diabetes mellitus (T2D) and cardiovascular disease. However, little is known about the effect of empagliflozin as add-on therapy on TTKG in T2D patients with ADHF. Purpose We sought to elucidate the effect of empagliflozin as add-on therapy on TTKG in T2D patients with ADHF. Methods We enrolled 58 consecutive T2D patients admitted for ADHF. On admission, enrolled patients were randomly assigned in a 1:1 ratio to either empagliflozin add-on therapy (EMPA(+)) or conventional glucose-lowering therapy (EMPA(−)). All patients in EMPA(+) group received empagliflozin (10 mg/day) throughout the study period. Left ventricular ejection fraction (LVEF) was measured at baseline using echocardiography. Body weight and vital signs, such as blood pressure and heart rate, were measured, and blood and urine samples were collected at baseline and 1, 2, 3 and 7 days after randomization. The TTKG was measured using the first morning urine samples collected on each day. TTKG was calculated according to the following equation: TTKG = (Ku/Ks)×(plasma osmolality/urine osmolality), where Ku is urine potassium concentration and Ks is serum potassium concentration, as previously reported. Results Thirty patients were assigned to the EMPA(+) group, and 28 patients were assigned to the EMPA(−) group. There were no significant baseline differences in LVEF, plasma B-type natriuretic peptide (BNP) level, body mass index, or serum creatinine level between the EMPA(+) and EMPA(−) groups. TTKG did not significantly differ between the two groups at baseline. However, seven days after randomization, plasma BNP level was significantly lower in the EMPA(+) group than in the EMPA(−) group (median 227 [IQR 114–381] pg/mL vs 362 [227–554] pg/mL, p=0.0294). Furthermore, TTKG of the EMPA(+) group was significantly lower at 2, 3 and 7 days after randomization (Figure). Conclusions This study demonstrated that empagliflozin as add-on therapy can lower TTKG in T2D patients with ADHF. Figure 1 Funding Acknowledgement Type of funding source: None

2054-P: Combination of Dipeptidyl Peptidase-4 (DPP-4) Inhibitor and Sodium–Glucose Cotransporter 2 (SGLT2) Inhibitor Substantially Protects Pancreatic ß-Cells Especially in Early Phase of Diabetes Rather than Advanced Phase

In diabetes treatment, it is important to protect pancreatic β-cells in addition to correct hyperglycemia. Although DPP-4 inhibitor and SGLT2 inhibitor are widely used nowadays, there is no report about β-cell protective effect by combination of both inhibitors. In this study, we examined the effects of DPP-4 inhibitor and/or SGLT2 inhibitor on β-cells and compared their effects between in early and advanced phase of diabetes. We used 7-week-old BKS. Cg-+ Leprdb/Jcl (db/db) mice as early phase and 16-week-old mice as advanced phase, and treated them for 2 weeks with oral administration of linagliptin 3 mg/kg (Lina group), empagliflozin 30mg/kg (Empa group), linagliptin + empagliflozin (L+E group), and 0.5% carboxymethylcellulose (Cont gourp). After 2-week treatment, blood glucose levels in Empa and L+E group were significantly lower than Cont group. OGTT after 1-week treatment showed that blood glucose levels were significantly lower in Empa and L+E group in both phases, and slightly lower in Lina group only in early phase. After 2-week treatment, islets size in L+E group was significantly larger than Cont group only in early phase. In GSIS using isolated islets, insulin levels in Lina, Empa and L+E group were significantly higher than Cont group in both phases. Furthermore, mRNA expression levels of ins1, ins2 and their transcription factors mafa, pdx-1, neurod and nkx6.1 were all higher in Lina and L+E group in early phase. Contrarily, mRNA expression levels of caspase3, 8 were lower in Lina and L+E group, and jnk and c-jun levels were lower in Empa and L+E group in early phase. Intriguingly, such changes were not observed in advanced phase. It seemed that DPP-4 inhibitor and SGLT2 inhibitor exerted beneficial effects on β-cells through some different mechanism. In conclusion, combination of DPP-4 inhibitor and SGLT2 inhibitor protects β-cells in early phase of diabetes. Disclosure Y. Fushimi: None. A. Obata: None. J. Sanada: None. Y. Nogami: None. T. Ikeda: None. Y. Obata: None. M. Shimoda: None. S. Nakanishi: None. T. Mune: None. K. Kaku: Advisory Panel; Self; Nippon Boehringer Ingelheim Co. Ltd., Novo Nordisk Inc. Consultant; Self; Sanwa Kagaku Kenkyusho. Research Support; Self; Taisho Pharmaceutical Co., Ltd. Speaker’s Bureau; Self; Astellas Pharma Inc., AstraZeneca, Daiichi Sankyo, Eli Lilly Japan K.K., Kowa Company, Ltd., Merck Sharp &amp; Dohme Corp., Mitsubishi Tanabe Pharma Corporation, Ono Pharmaceutical Co., Ltd., Takeda Pharmaceutical Company Limited. H. Kaneto: Research Support; Self; Astellas Pharma Inc., Boehringer Ingelheim Pharmaceuticals, Inc., Daiichi Sankyo, Eli Lilly Japan K.K., Kissei Pharmaceutical Co., Ltd., Mitsubishi Tanabe Pharma Corporation, Novo Nordisk Inc., Ono Pharmaceutical Co., Ltd., Sanofi, Sumitomo Dainippon Pharma Co., Ltd., Taisho Pharmaceutical Co., Ltd., Takeda Pharmaceutical Company Limited. Speaker’s Bureau; Self; Astellas Pharma Inc., AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Daiichi Sankyo, Eli Lilly Japan K.K., Mitsubishi Tanabe Pharma Corporation, Novartis Pharmaceuticals Corporation, Novo Nordisk Inc., Ono Pharmaceutical Co., Ltd., Sanofi, Sumitomo Dainippon Pharma Co., Ltd., Takeda Pharmaceutical Company Limited.

LB005KIDNEY IMPLICATIONS OF THE INITIAL EGFR RESPONSE TO SGLT2 INHIBITION WITH EMPAGLIFLOZIN: THE ‘EGFR DIP’ IN EMPA-REG OUTCOME

Abstract Background and Aims Empagliflozin (EMPA) reduces cardiovascular and renal risk in patients with type 2 diabetes (T2D) and established cardiovascular disease (CVD). EMPA induces an initial ‘dip’ in estimated glomerular filtration rate (eGFR). Although considered to be of haemodynamic origin and largely reversible, this needs to be better understood. We investigated whether the initial eGFR dip after EMPA initiation was influenced by baseline characteristics and/or might have an impact on the EMPA-induced risk reduction in kidney outcomes. Method In the EMPA-REG OUTCOME trial, patients with T2D and established CVD were treated (1:1:1) with EMPA 10 mg, 25 mg or placebo (PBO), in addition to standard of care. In this post hoc analysis, 6,668 participants who received at least one dose of study drug and had an available eGFR value at both baseline and Week 4 were categorised by initial percentage eGFR change from baseline. A multivariate logistic regression model was used to identify which baseline characteristics are predictive of an initial eGFR dip &amp;gt;10% in EMPA-treated participants versus PBO. Across these predictive baseline factors, we investigated the occurrence of incident or worsening nephropathy, hard kidney outcomes (defined as doubling of serum creatinine with eGFR [MDRD] ≤45 ml/min/1.73 m2 or initiation of renal replacement therapy or death from renal disease), and kidney safety (narrow standardized MedDRA query acute renal failure). The impact of an eGFR dip &amp;gt;10% on the risk reduction with EMPA for incident or worsening nephropathy was assessed using Cox regression analysis adjusting for such eGFR dip. Results In the EMPA-REG OUTCOME trial cohort, an initial eGFR dip of &amp;gt;10% from baseline at Week 4 occurred in more than twice as many participants on EMPA (28.3%) compared to PBO (13.4%). However, a more pronounced eGFR dip of &amp;gt;30% was uncommon, occurring in only 1.4% and 0.9%, respectively. Within the EMPA group, participants with an eGFR dip &amp;gt;10% were significantly older, had longer diabetes duration and showed a higher KDIGO (Kidney Disease: Improving Global Outcomes) risk category. Diuretic use and/or higher KDIGO risk category at baseline were predictive of an initial eGFR dip of &amp;gt;10% in EMPA vs. PBO. The average odds ratio [OR; 95% CI] for an eGFR dip &amp;gt;10% with EMPA was 2.7 [2.3–3.0]. In subgroups with a dipping odds ratio below vs. above that average, beneficial treatment effects with EMPA on incident or worsening nephropathy and the hard kidney outcome were consistent (panel A). Also, an eGFR dip &amp;gt;10% did not affect risk reduction for the primary kidney outcome (panel B). Acute renal failure rates were generally lower or similar in EMPA vs. PBO, regardless of baseline predictive factors for an eGFR dip. Conclusion T2D patients with more advanced kidney disease and/or on diuretic therapy at baseline were more likely to have an initial eGFR dip &amp;gt;10% with EMPA. However, EMPA treatment appeared to be safe and was associated with improved kidney outcomes, regardless of these baseline predictive factors or an initial eGFR dip &amp;gt;10%.

29-OR: Comparison of Empagliflozin and Sitagliptin on Ectopic Fat Accumulation and Tissue-Specific Insulin Sensitivity

Background: Ectopic fat accumulation is the major pathogenesis of insulin resistance, type 2 diabetes mellitus (T2DM), and cardiovascular diseases. To date, no prospective study compared the effects of sodium glucose cotransporter 2 inhibitors (SGLT2i) and dipeptidyl peptidase-4 inhibitors on ectopic fat accumulation and tissue-specific insulin sensitivity. Methods: This is the prospective, randomized, open-label, and blind-endpoint study in 44 Japanese patients with T2DM. Patients (HbA1c: 7.2±0.9%, known duration: 3.9±4.0 years, BMI: 29.1±4.8 kg/m2) were randomized to empagliflozin (Empa) add-on group or sitagliptin (Sita) add-on group, and treated for 12 weeks. The primary endpoint was the change of pericardial fat content estimated by magnetic resonance imaging. The secondary endpoints were the changes in the amount of intrahepatic, intra- and extra-myocellular lipid content estimated by proton magnetic resonance spectroscopy, and tissue-specific insulin sensitivity assessed by hyperinsulinemic-euglycemic clamp tests using stable isotope. Results: There was no difference in pericardial fat content between the groups. Interestingly, intrahepatic lipid content (IHL) was reduced significantly from baseline in Empa group compared to Sita (-23.0±26.3 vs. -3.9±19.8%, respectively, p&amp;lt;0.05). However, the change in clamped % suppression of endogenous glucose production was comparable between the groups. On the other hand, rate of glucose disappearance was significantly increased in Empa group compared to Sita (+18.5±55.2 vs. -8.6±20.2%, p&amp;lt;0.05), while the amount of intra- and extra-myocellular lipid content remained similar between the groups. Conclusion: This is the first study that compared Empa and Sita on ectopic fat content and tissue-specific insulin sensitivity. Empa significantly reduced IHL and increased insulin sensitivity in muscle compared to Sita. SGLT2i might be preferable in treatment of T2DM with hepatic steatosis. Disclosure S. Hiruma: None. F. Shigiyama: None. S. Hisatake: None. S. Mizumura: None. N. Shiraga: None. M. Hori: None. T. Ikeda: None. T. Hirose: Research Support; Self; Boehringer Ingelheim Pharmaceuticals, Inc., Daiichi Sankyo, Mitsubishi Tanabe Pharma Corporation, Ono Pharmaceutical Co., Ltd., Takeda Pharmaceutical Company Limited. Speaker’s Bureau; Self; AstraZeneca, Lilly Diabetes, Novo Nordisk Inc., Sanofi, Sumitomo Dainippon Pharma Co., Ltd. N. Kumashiro: Research Support; Self; Boehringer Ingelheim Pharmaceuticals, Inc. Speaker’s Bureau; Self; Novo Nordisk Inc., Ono Pharmaceutical Co., Ltd., Takeda Pharmaceutical Company Limited. Funding Nippon Boehringer Ingelheim Co., Ltd. (1245-0162)

Empagliflozin prevents doxorubicin-induced myocardial dysfunction

BackgroundEmpagliflozin showed efficacy in controlling glycaemia, leading to reductions in HbA1c levels, weight loss and blood pressure, compared to standard treatment. Moreover, the EMPA-REG OUTCOME trial demonstrated a 14% reduction of major adverse cardiovascular events (MACE), a 38% reduction in cardiovascular (CV) death and a 35% reduction in the hospitalization rate for heart failure (HF). These beneficial effect on HF were apparently independent from glucose control. However, no mechanistic in vivo studies are available to explain these results, yet. We aimed to determine the effect of empagliflozin on left ventricular (LV) function in a mouse model of doxorubicin-induced cardiomyopathy (DOX-HF).MethodsMale C57Bl/6 mice were randomly assigned to the following groups: controls (CTRL, n = 7), doxorubicin (DOX, n = 14), DOX plus empagliflozin (DOX + EMPA, n = 14), or DOX plus furosemide (DOX + FURO group, n = 7). DOX was injected intraperitoneally. LV function was evaluated at baseline and after 6 weeks of treatment using high-resolution echocardiography with 2D speckle tracking (Vevo 2100). Histological assessment was obtained using Haematoxylin and Eosin and Masson’s Goldner staining.ResultsA significant decrease in both systolic and diastolic LV function was observed after 6 weeks of treatment with doxorubicin. EF dropped by 32% (p = 0.002), while the LS was reduced by 42% (p < 0.001) and the CS by 50% (p < 0.001). However, LV function was significantly better in the DOX + EMPA group, both in terms of EF (61.30 ± 11% vs. 49.24 ± 8%, p = 0.007), LS (− 17.52 ± 3% vs. − 13.93 ± 5%, p = 0.04) and CS (− 25.75 ± 6% vs. − 15.91 ± 6%, p < 0.001). Those results were not duplicated in the DOX + FURO group. Hearts from the DOX + EMPA group showed a 50% lower degree of myocardial fibrosis, compared to DOX mice (p = 0.03). No significant differences were found between the DOX + FURO and the DOX group (p = 0.103).ConclusionEmpagliflozin attenuates the cardiotoxic effects exerted by doxorubicin on LV function and remodelling in nondiabetic mice, independently of glycaemic control. These findings support the design of clinical studies to assess their relevance in a clinical setting.

4330Effect of empagliflozin as add-on therapy on decongestion and renal function in diabetic patients hospitalized for acute decompensated heart failure: a prospective randomized controlled study

Abstract Background The mainstay of treatment of acute decompensated heart failure (ADHF) is decongestion by diuretic therapy. Empagliflozin has been shown to reduce the risk of hospitalization for heart failure in patients (pts) with type 2 diabetes mellitus (T2D) and cardiovascular disease. This may be explained by natriuresis and osmotic diuresis caused by empagliflozin, leading to plasma volume (PV) contraction and decongestion. However, little is known about the therapeutic effect of empagliflozin on decongestion and its association with renal function in T2D pts with ADHF. Purpose We sought to elucidate the effect of empagliflozin as add-on therapy on plasma B-type natriuretic peptide (BNP) level, hemoconcentration, PV contraction and renal function in T2D pts with ADHF. Methods We enrolled 38 consecutive T2D pts admitted for ADHF. On admission, enrolled pts were randomly assigned in a 1:1 ratio to either empagliflozin add-on therapy (EMPA(+)) or conventional glucose-lowering therapy (EMPA(−)). All pts in EMPA(+) group received empagliflozin (10mg/day) throughout the study period. Left ventricular ejection fraction (LVEF) was measured at baseline using echocardiography. Body weight and vital signs, such as blood pressure and heart rate, were measured, and blood and urine samples were collected at baseline and 1, 2, 3 and 7 days after randomization. Hemoconcentration was defined as a ≥3% absolute increase in hematocrit (Hct). Percent change in PV between admission and subsequent timepoints (%ΔPV) was calculated using the Strauss formula as follows: %ΔPV = ([(Hb1/Hb2) × ((100 − Hct2)/(100 − Hct1))] − 1) × 100 (%), where 1 = baseline values and 2 = subsequent values. Worsening renal function (WRF) was defined as an increase in serum creatinine ≥0.3 mg/dL above baseline within 7 days of randomization. Results Twenty pts were assigned to the EMPA(+) group, and 18 pts were assigned to the EMPA(−) group. There were no significant baseline differences in LVEF, plasma BNP level, Hct or serum creatinine level between the EMPA(+) and EMPA(−) groups. Seven days after randomization, plasma BNP level was significantly lower in the EMPA(+) group than in the EMPA(−) group (median 213 [IQR 116–360] pg/mL vs 362 [226–776] pg/mL, p=0.0437) and hemoconcentration was more frequently observed in the EMPA(+) group than in the EMPA(−) group (53% vs 12%, p=0.0105). The decrease in %ΔPV was larger in the EMPA(+) group than in the EMPA(−) group 2 days (−8.74±9.92% vs 1.14±14.71%, p=0.0228), 3 days (−11.28±10.65% vs −0.02±14.70%, p=0.0121) and 7 days after randomization (−10.62±14.89% vs 0.97±13.72%, p=0.0211). The incidence of WRF did not significantly differ between the EMPA(+) and EMPA(−) groups (15% vs 22%). Conclusions This study demonstrated that empagliflozin as add-on therapy can achieve effective decongestion without an increased risk of WRF in T2D pts with ADHF.

P5413Effect of empagliflozin as add-on therapy on serum uric acid level in patients with type 2 diabetes hospitalized for acute decompensated heart failure: a prospective randomized controlled study

Abstract Background Elevated serum uric acid (UA) level has been shown to be associated with reduced survival among patients (pts) with heart failure. Sodium glucose cotransporter 2 (SGLT2) inhibitors have been reported to lower serum uric acid level in pts with type 2 diabetes mellitus (T2D). Empagliflozin, one of the SGLT2 inhibitors, has been shown to reduce the risk of cardiovascular mortality in T2D pts with cardiovascular disease, and involvement of UA lowering effect by empagliflozin in the reduction of cardiovascular mortality has been suggested. However, little is known about the effect of empagliflozin as add-on therapy on serum UA level in T2D pts with acute decompensated heart failure (ADHF). Purpose We sought to elucidate the effect of empagliflozin as add-on therapy on serum UA level in T2D pts with ADHF. Methods We enrolled 38 consecutive T2D pts admitted for ADHF. On admission, enrolled pts were randomly assigned in a 1:1 ratio to either empagliflozin add-on therapy (EMPA(+)) or conventional glucose-lowering therapy (EMPA(−)). All pts in EMPA(+) group received empagliflozin (10 mg/day) throughout the study period. Left ventricular ejection fraction (LVEF) was measured at baseline using echocardiography. Body weight and vital signs, such as blood pressure and heart rate, were measured, and blood and urine samples were collected at baseline and 1, 2, 3 and 7 days after randomization. Renal handling of UA was evaluated by fractional excretion of UA (FEUA). Results Twenty pts were assigned to the EMPA(+) group, and 18 pts were assigned to the EMPA(−) group. There were no significant baseline differences in LVEF, plasma brain natriuretic peptide level, body mass index, or serum creatinine level between the EMPA(+) and EMPA(−) groups. In addition, prevalence rate of hyperuricemia, serum UA level, and FEUA did not significantly differ between the two groups at baseline. However, there was significant difference in the change in serum UA level from baseline at 2, 3 and 7 days after randomization between the two groups (Figure A). As a result, serum UA level was significantly lower in the EMPA(+) group than in the EMPA(−) group at 7 days after randomization (6.2±1.8 mg/dL vs 7.8±1.8 mg/dL, p=0.0127). Moreover, FEUN of the EMPA(+) group was significantly higher at 1, 2 and 7 days after randomization (Figure B), which suggested that serum UA level was lowered in the EMPA(+) group by increased urinary excretion of UA. Figure 1 Conclusions This study demonstrated that empagliflozin as add-on therapy can lower serum UA level in T2D pts with ADHF through the effect on the urinary excretion rate of UA.