Abstract

The relationship between disturbances in glucose homeostasis and heart failure (HF) progression is bidirectional. However, the mechanisms by which HF intrinsically impairs glucose homeostasis remain unknown. The present study tested the hypothesis that the bioavailability of intact glucagon-like peptide-1 (GLP-1) is affected in HF, possibly contributing to disturbed glucose homeostasis. Serum concentrations of total and intact GLP-1 and insulin were measured after an overnight fast and 15 min after the ingestion of a mixed breakfast meal in 49 non-diabetic patients with severe HF and 40 healthy control subjects. Similarly, fasting and postprandial serum concentrations of these hormones were determined in sham-operated rats, and rats with HF treated with an inhibitor of the GLP-1-degrading enzyme dipeptidyl peptidase-4 (DPP4), vildagliptin, or vehicle for 4 weeks. We found that HF patients displayed a much lower increase in postprandial intact and total GLP-1 levels than controls. The increase in postprandial intact GLP-1 in HF patients correlated negatively with serum brain natriuretic peptide levels and DPP4 activity and positively with the glomerular filtration rate. Likewise, the postprandial increases in both intact and total GLP-1 were blunted in HF rats and were restored by DPP4 inhibition. Additionally, vehicle-treated HF rats displayed glucose intolerance and hyperinsulinemia, whereas normal glucose homeostasis was observed in vildagliptin-treated HF rats. We conclude that the postprandial increase in GLP-1 is blunted in non-diabetic HF. Impaired GLP-1 bioavailability after meal intake correlates with poor prognostic factors and may contribute to the establishment of a vicious cycle between glucose disturbance and HF development and progression.

Introduction

Heart failure (HF) is a complex heterogeneous syndrome that involves the activation of several neurohormonal, metabolic, and immune responses. Although the etiology of the syndrome can be diverse, diseases such as hypertension, myocardial infarction, and diabetes are important risk factors [1]. Almost 50 years ago, the Framingham Heart Study demonstrated that the risk of developing HF is 2-fold higher for diabetic men and 5-fold higher for diabetic women compared with non-diabetic subjects [2]. In fact, diabetes is a strong predictor of mortality in HF patients [3]. Studies have also suggested that disturbances in glucose homeostasis, such as glucose intolerance and insulin resistance, correlate with poor prognosis in cardiovascular diseases, including coronary artery disease [4], acute myocardial infarction [5], idiopathic dilated cardiomyopathy [6], and HF [7–9].

Interestingly, the relationship linking disturbances in glucose homeostasis and HF progression seems to be bidirectional [8,10]. Indeed, Tenenbaum and colleagues [8] observed that non-diabetic patients with advanced HF (NYHA class III) have a significantly increased risk of developing diabetes during a 6- to 9-year follow-up than that of those with lower functional classes of HF. Furthermore, in a cohort of 50,874 patients, HF severity predicted the risk of developing diabetes after myocardial infarction [11]. However, the mechanism of impaired glucose homeostasis intrinsically caused by advanced HF is largely unknown.

Glucagon-like peptide-1 (GLP-1) is a gut-derived incretin hormone that plays a determinant role in blood glucose homeostasis by virtue of its ability to enhance insulin secretion, suppress glucagon release, and reduce gastric emptying and food intake [12]. The basal rate of GLP-1 secretion by enteroendocrine L-cells during the fasting state is low. However, its circulating concentration rises rapidly, by approximately 10 min, after the intake of nutrients, including carbohydrates, fats, and proteins [13,14]. Intact GLP-1(7-36) amide is rapidly metabolized by the ubiquitously expressed serine protease enzyme dipeptidyl-peptidase-4 (DPP4), which cleaves the two N-terminal amino acids of the molecule, resulting in the formation of the metabolite GLP-1(9-36) amide [15]. Given the essential role of GLP-1 in glucose homeostasis, strategies that potentiate the incretin pathway (DPP4 inhibitors and GLP1 receptor agonists that are resistant to DPP4 inactivation) have been used for the treatment of Type 2 diabetes.

Emerging evidence from both preclinical and clinical studies has raised the possibility that DPP4 may also be involved in the pathophysiology of HF [16]. Indeed, our group and others have found that DPP4 activity is increased in non-diabetic HF patients and rats [17–19]. Moreover, patients with acute HF that exhibit elevated levels of circulating DPP4 (highest quartile) display a 3-fold higher risk of death due to HF within 6 months [19]. Despite the knowledge of the importance of DPP4, little is known about circulating intact GLP-1 levels in HF. In light of the above, we hypothesize that intact GLP-1 bioavailability may be affected in HF, thereby contributing to disturbed glucose homeostasis in HF patients with normal fasting glycemia.

Methods

Study subjects

All patients signed an informed consent form. The study was approved by the Ethics Committee of the Heart Institute of the University of São Paulo, São Paulo, Brazil, and was carried out following the ethical standards of the Helsinki Declaration of the World Medical Association. The study included 49 non-diabetic patients with HF in functional class III or IV of the New York Heart Association from an ongoing inception cohort from the General Outpatient Clinic of the Heart Institute, University of São Paulo Medical School (Table 1). The ascertainment period was from 2014 to 2018. The diagnosis of heart failure was made according to previously published criteria [20]. Patients with a known diagnosis of diabetes mellitus (DM) and a left ventricular (LV) ejection fraction ≥ 40% were excluded from the study. Even patients with no history of diabetes but with fasting blood glucose > 110 mg/dl and glycated hemoglobin A1c levels higher than 5.8% at the time of enrolment were excluded. Forty healthy individuals were used as appropriate controls. For all subjects included in the study, venous blood samples were collected during the following two periods: (1) after a fasting period of 8 h (t = 0 min) and (2) 15 min after the ingestion of a mixed meal containing 69 g carbohydrates, 10 g protein, and 7 g fat, yielding 381 kcal (t = 15 min). Blood samples were collected into chilled Vacutainer plastic serum tubes (BD Biosciences, San Jose, CA, U.S.A.) containing 10 mM of the DPP4 inhibitor P32/98 (Abcam, Burlingame, CA, U.S.A.). The tubes were centrifuged at 3000 rpm at 4°C for 15 min, and the serum was stored at −80°C. Serum was used to determine glucose and insulin levels, DPP4 activity, total brain natriuretic peptide (BNP) concentration, total and intact GLP-1, and creatinine. The estimated glomerular filtration rate (eGFR) was calculated using the MDRD (modification of diet in renal disease) equation.

Table 1
Clinical characteristics of the studied population
Control subjectsHF patientsP value
Number of individuals 40 49  
Age, years (SD) 49.8 (12.5) 52.6 (11.5) 0.2749# 
Gender    
Male, N (%) 28 (70.0) 39 (79.6)  
Weight, kg (SD) 73.0 (14.0) 71.1 (10.4) 0.4649# 
Height, m (SD) 1.67 (0.10) 1.66 (0.07) 0.5947& 
Body mass index, kg/m2 (SD) 25.5 (3.4) 26.0 (3.3) 0.6993& 
Blood pressure, mmHg (SD)    
  Systolic 115 (9) 118 (23) 0.4052# 
  Diastolic 72 (10) 74 (15) 0.4722# 
Heart rate, beats/min (SD) 70 (10) 76 (14) 0.0253# 
LV ejection fraction, % (SD) 68 (5) 27 (5) <0.001# 
Systemic arterial hypertension, N (%) 0 (0) 34 (69.4)  
Serum BNP, pg/ml (SD) 47 (6) 1987 (1495) <0.001* 
Fasting serum glucose, mg/dl (SD) 93 (5) 92 (8) 0.4738# 
Postprandial serum glucose, mg/dl (SD) 112 (2) 140 (27) <0.001* 
Hb1Ac, % (SD) 5.0 (0.3) 5.1 (0.4) 0.1941# 
Serum creatinine, mg/dl (SD) 0.8 (0.1) 1.1 (0.3) <0.001* 
Creatinine clearance (MDRD), ml/min/1.73 m2 (SD) 117 (7) 81 (25) <0.001& 
Control subjectsHF patientsP value
Number of individuals 40 49  
Age, years (SD) 49.8 (12.5) 52.6 (11.5) 0.2749# 
Gender    
Male, N (%) 28 (70.0) 39 (79.6)  
Weight, kg (SD) 73.0 (14.0) 71.1 (10.4) 0.4649# 
Height, m (SD) 1.67 (0.10) 1.66 (0.07) 0.5947& 
Body mass index, kg/m2 (SD) 25.5 (3.4) 26.0 (3.3) 0.6993& 
Blood pressure, mmHg (SD)    
  Systolic 115 (9) 118 (23) 0.4052# 
  Diastolic 72 (10) 74 (15) 0.4722# 
Heart rate, beats/min (SD) 70 (10) 76 (14) 0.0253# 
LV ejection fraction, % (SD) 68 (5) 27 (5) <0.001# 
Systemic arterial hypertension, N (%) 0 (0) 34 (69.4)  
Serum BNP, pg/ml (SD) 47 (6) 1987 (1495) <0.001* 
Fasting serum glucose, mg/dl (SD) 93 (5) 92 (8) 0.4738# 
Postprandial serum glucose, mg/dl (SD) 112 (2) 140 (27) <0.001* 
Hb1Ac, % (SD) 5.0 (0.3) 5.1 (0.4) 0.1941# 
Serum creatinine, mg/dl (SD) 0.8 (0.1) 1.1 (0.3) <0.001* 
Creatinine clearance (MDRD), ml/min/1.73 m2 (SD) 117 (7) 81 (25) <0.001& 

Abbreviations: LV, left ventricle; SD, standard deviation. Comparisons were performed by unpaired Student’s t-test (#), t-test with Welch’s correction for different variances (&) or the Mann–Whitney test for non-Gaussian distributions (*).

Animal protocols, surgical procedures, and drug treatment

All animal procedures were approved by the Institutional Animal Care and Use Committee of the University of São Paulo Medical School and were performed following the ethical principles of the Brazilian College of Animal Experimentation. Experiments were performed on male Wistar rats (8–10 weeks old, 200–250 g) purchased from the University of São Paulo (São Paulo, Brazil). The animals were housed at the Heart Institute animal facility under a constant temperature and a 12:12-h dark–light cycle and had free access to food and water. One day before surgery, blood and urine samples were collected from rats. Experimental HF was then induced in the rats through LV myocardial injury after radiofrequency catheter ablation, as described previously [21]. In brief, the animals were anesthetized with an intraperitoneal injection of ketamine and xylazine (50 and 10 mg/kg, respectively). Once the anesthesia was confirmed, rats were intubated under artificial ventilation (Hugo Basile - model 7025 - Biological Research Apparatus, Comélio, Itália), a left thoracotomy was performed, and a murine surgical retractor was inserted in the fourth intercostal space. The heart was restrained with an indifferent electrode, the catheter tip placed in the anterolateral left ventricle wall, and the lesions created with radiofrequency ablation apparatus (model TEB RF10; Tecnologia Eletrônica Brasileira Ltda, São Paulo, Brazil). Subsequently, the electrode and catheter were removed, and the chest immediately closed. Sham-operated rats underwent left thoracotomy and were mock-ablated. Echocardiographic evaluation of left ventricular systolic function, as well as serum levels of BNP, were used to characterize HF. HF was considered when the fractional area change (FAC) was lower than 40% and circulating levels of BNP were higher than 1.0 ng/ml. Six weeks after surgery, the radiofrequency-ablated rats that developed HF were randomly divided into two groups and treated for 4 weeks with vildagliptin (120 mg/kg/day, twice daily) or vehicle by oral gavage. Vehicle-treated sham rats were used as controls. At the end of the treatment period and after a 6-h fast, the rats were anesthetized by an intraperitoneal injection of pentobarbital (50 mg/kg), and the rats were subsequently euthanized by decapitation. The heart and lungs were excised and weighed. The heart was immediately frozen in liquid nitrogen and stored at −80°C for subsequent protein extraction. The expression levels of the GLUT4 glucose transporter in the heart plasma membrane were evaluated by immunoblotting, as previously described [22]. The animal study design is depicted in Supplementary Figure S1.

Echocardiography

Echocardiographic indices of systolic and diastolic function were performed 6 weeks after LV-radiofrequency ablation or sham surgery (pre-treatment) and after treatment with vehicle or vildagliptin (post-treatment). They were quantified by an expert echocardiographer blinded to the study conditions. Images were analyzed according to the method reported in [23].

Determination of the serum concentrations of intact and total GLP-1 and total BNP

To measure GLP-1 in the rats, blood was withdrawn from the retroorbital plexus after 6 h of fasting and after 5 min of oral glucose administration (2 g/kg). Experiments to determine the levels of GLP-1 in the rats were performed at two time points: (1) Baseline: before radiofrequency catheter ablation for HF induction or sham operation and (2) Post-treatment: after 4 weeks of treatment with vehicle or vildagliptin (Supplementary Figure S1). The blood was immediately transferred to chilled tubes (BD Biosciences, San Jose, CA, U.S.A.) containing the DPP4 inhibitor P32/98 (Abcam) and centrifuged at 3000 rpm at 4°C for 10 min. Serum samples were frozen and stored at −80°C. The serum concentration of total and active GLP-1 (7–36) was determined by enzyme-linked immunosorbent assay (ELISA) (Merck Millipore, Burlington, MA, U.S.A.) according to the manufacturer’s instructions. BNP levels were determined in serum isolated from humans and rats using the Human BNP ELISA kit and the Rat BNP 32 ELISA kit (Abcam), respectively, according to the manufacturer’s instructions.

Determination of the urinary concentration of cAMP

Rats were placed into metabolic cages (Techniplast) after oral glucose administration (2 g/kg) and blood withdrawal for GLP-1 measurements. Urine was collected for 4 h and analyzed for cAMP concentration. Experiments were performed at the baseline and post-treatment time points (Supplementary Figure S1). The urinary content of cAMP was determined by ELISA (Arbor Assays, Ann Arbor, MI) following the manufacturer’s instructions.

Determination of the serum concentrations of glucose, insulin, and DPP4 activity

Serum glucose was measured by the hexokinase method using a commercially available kit (Labtest, Lagoa Santa, MG, Brazil). The serum insulin concentration was measured using a Human Insulin ELISA kit or a Rat/Mouse Insulin ELISA kit purchased from Merck Millipore according to the manufacturer’s protocol. DPP4 activity in the serum was measured colorimetrically using glycyl-prolyl-para-nitroanilide as a chromogenic substrate, as previously reported [24]. Two days before killing, the rats were subjected to an oral glucose tolerance test (OGTT) (2 g/kg), as described previously [25]. The area under the curve (AUC) was determined for the quantification of the OGTT.

SDS-PAGE and immunoblotting

Harvested hearts from rats were homogenized in a Polymix PX-SR 50 E homogenizer (Kinematica, AG, Switzerland) in ice-cold phosphate-buffered saline (PBS) (2.8 mM sodium phosphate monobasic, 7.2 mM sodium phosphate dibasic, 150 mM NaCl, pH 7.4) containing phosphatase inhibitors (15 mM NaF and 50 mM sodium pyrophosphate) and Halt Protease Inhibitor Cocktail (Thermo Fisher Scientific, Rockford, IL). Equal amounts of heart proteins were resolved by SDS-PAGE and analyzed by immunoblotting, as described elsewhere [26]. The visualized bands were digitized using an ImageScanner (GE HealthCare) and quantified using the Scion Image Software package (Scion Corporation, Frederick, MD).

Statistical analysis

The data are expressed as the mean ± SEM unless otherwise specified. For each data set, the Levene test was performed to examine the homogeneity of variances. If the Levene test revealed unequal variance, Welch ANOVA was used. If the variance was equal, then Student’s t-test or one-way ANOVA followed by Bonferroni post hoc tests was performed. The relationship between the postprandial increase in intact GLP-1 and HF parameters in patients was assessed by the Spearman correlation test. The difference was considered to be statistically significant if P was less than 0.05.

Results

Postprandial increase in GLP-1 is impaired in non-diabetic patients with HF

The characteristics of the study population are shown in Table 1 and Supplementary Table S1. The evaluation of the intact and total levels of GLP-1 in the serum of patients with HF and control individuals is presented in Figure 1. The fasting serum concentration of intact GLP-1 was similar between patients with HF and control subjects (Figure 1A). However, 15 min after meal ingestion, the serum concentration of intact GLP-1 was much lower in patients with HF than in controls (Figure 1A). Indeed, the percentage of postprandial increase in intact GLP-1 was practically negligible in patients with HF when compared with controls (5 ± 4 vs. 65 ± 4%, P < 0.0001) (Figure 1B).

Postprandial increase in intact and total GLP-1 is impaired in non-diabetic patients with HF

Figure 1
Postprandial increase in intact and total GLP-1 is impaired in non-diabetic patients with HF

Blood samples were collected after an overnight fast and 15 min after nutrient intake. (A) Intact GLP-1 fasting (t = 0) and postprandial (t = 15) levels and (B) the percentage of increase in serum levels of intact GLP-1. (C) Total GLP-1 fasting (t = 0) and postprandial (t = 15) levels and (D) the percentage of increase in serum levels of total GLP-1. The values are the means ± SEM; *P<0.05, ***P<0.001, and ****P<0.0001 versus control subjects.

Figure 1
Postprandial increase in intact and total GLP-1 is impaired in non-diabetic patients with HF

Blood samples were collected after an overnight fast and 15 min after nutrient intake. (A) Intact GLP-1 fasting (t = 0) and postprandial (t = 15) levels and (B) the percentage of increase in serum levels of intact GLP-1. (C) Total GLP-1 fasting (t = 0) and postprandial (t = 15) levels and (D) the percentage of increase in serum levels of total GLP-1. The values are the means ± SEM; *P<0.05, ***P<0.001, and ****P<0.0001 versus control subjects.

Surprisingly, the serum levels of total GLP-1 under fasting conditions were higher in patients with HF than in controls (Figure 1C). However, as shown in Figure 1D, the postprandial increase in total GLP-1 was much less pronounced in patients with HF than in control individuals (14 ± 4 vs. 53 ± 5%, P < 0.0001).

Postprandial increases in insulin and glucose levels are higher in non-diabetic patients with HF

Figure 2 shows the fasting and postprandial levels of insulin (left panel) and glucose (right panel) in patients with HF and healthy controls. No differences were found in the fasting concentrations of insulin and glucose between patients with HF and control subjects. Interestingly, the postprandial concentrations of both insulin (21.4 ± 1.3 vs. 10.8 ± 0.6 µU/ml; P < 0.01) and glucose (140 ± 3 vs. 118 ± 2 mg/dl; P < 0.001) were significantly augmented in the HF group.

Non-diabetic patients with HF display higher postprandial insulin and glucose levels than healthy controls

Figure 2
Non-diabetic patients with HF display higher postprandial insulin and glucose levels than healthy controls

Blood samples were collected after an overnight fast and 15 min after nutrient intake in 49 non-diabetic heart failure patients and 40 control subjects. (A) Fasting and postprandial blood insulin levels and (B) fasting and postprandial blood glucose levels. The values are the means ± SEM; *P<0.05, ***P<0.001, and ****P<0.0001 versus fasting; ##P<0.01 and ###P<0.001 versus postprandial control subjects.

Figure 2
Non-diabetic patients with HF display higher postprandial insulin and glucose levels than healthy controls

Blood samples were collected after an overnight fast and 15 min after nutrient intake in 49 non-diabetic heart failure patients and 40 control subjects. (A) Fasting and postprandial blood insulin levels and (B) fasting and postprandial blood glucose levels. The values are the means ± SEM; *P<0.05, ***P<0.001, and ****P<0.0001 versus fasting; ##P<0.01 and ###P<0.001 versus postprandial control subjects.

Postprandial increase in intact GLP-1 correlates with HF biomarkers in non-diabetic patients with HF

A significant negative correlation was found between the postprandial increase in intact GLP-1 and serum BNP levels in patients with severe HF (Figure 3A). Conversely, the postprandial rise in intact GLP-1 did not correlate with the LV ejection fraction in patients with HF (Figure 3B). Moreover, statistically significant correlations were observed between the postprandial increase in intact GLP-1 and GFR (Figure 3C). As expected, the lower the postprandial increase in intact GLP-1, the higher was the activity of its degrading enzyme DPP4 (Figure 3D).

Postprandial increase in intact GLP-1 levels is negatively correlated with circulating BNP and DPP4 levels

Figure 3
Postprandial increase in intact GLP-1 levels is negatively correlated with circulating BNP and DPP4 levels

The Pearson’s correlation coefficient (r) between serum GLP-1 levels and (A) brain natriuretic peptide (BNP) levels, (B) left ventricle ejection fraction (LVEF), (C) estimated glomerular filtration rate (eGFR) and (D) circulating dipeptidyl peptidase-4 (DPP4) activity in patients with HF was determined.

Figure 3
Postprandial increase in intact GLP-1 levels is negatively correlated with circulating BNP and DPP4 levels

The Pearson’s correlation coefficient (r) between serum GLP-1 levels and (A) brain natriuretic peptide (BNP) levels, (B) left ventricle ejection fraction (LVEF), (C) estimated glomerular filtration rate (eGFR) and (D) circulating dipeptidyl peptidase-4 (DPP4) activity in patients with HF was determined.

Postprandial increase in GLP-1 is blunted in HF rats, and it is restored by DPP4 inhibition

The biometric and Doppler echocardiographic characteristics of sham-operated and HF rats treated or not with the DPP4 inhibitor vildagliptin are shown in Supplementary Tables S2 and S3, respectively. Ten weeks after cardiac injury by radiofrequency ablation, rats exhibited typical markers of cardiac remodeling and dysfunction (Supplementary Tables S2 and S3), consistent with the literature [21,23]. Additionally, HF rats displayed higher serum DPP4 activity and levels than sham rats (Supplementary Table S2). Moreover, in line with a previous study [23], the treatment of rats with established HF with the DPP4 inhibitor vildagliptin conferred improvements in cardiac function, pulmonary congestion, and BNP levels (Supplementary Tables S2 and S3).

To evaluate the GLP-1 response to oral glucose in sham, HF and HF rats treated with vildagliptin, the concentrations of intact and total GLP-1 were evaluated under fasting conditions and 5 min after glucose ingestion (2 g/kg) (Figure 4). A series of experiments were performed at baseline, i.e. before heart failure induction by LV radiofrequency ablation or sham operation (Figure 4A–D). At this time point, the fasting and postprandial concentrations of intact and total GLP-1 were similar among the three groups (Figure 4A,C). Accordingly, at baseline, the percentage of postprandial increase in both intact and total GLP-1 did not vary among the experimental groups (Figure 4B,D).

Postprandial increase in intact and total GLP-1 is impaired in heart failure rats and restored by the DPP4 inhibitor vildagliptin

Figure 4
Postprandial increase in intact and total GLP-1 is impaired in heart failure rats and restored by the DPP4 inhibitor vildagliptin

Blood samples were collected after an overnight fast and 5 min after an oral glucose load of 2 g/kg from sham, HF rats, and HF rats treated with vildagliptin (HF + DPP4i). Experiments were conducted at baseline, i.e., before LV radiofrequency ablation for HF induction or sham operation (A–D) and at post-treatment (E–H). (A and E) Intact GLP-1 fasting (t = 0) and postprandial (t = 5) levels. (B and F) The percentage of increase in serum levels of intact GLP-1 after an oral glucose load. (C and G) Total GLP-1 fasting (t = 0) and postprandial (t = 5) levels. (D and H) The percentage of increase in serum levels of total GLP-1 after an oral glucose load. The values are the means ± SEM; **P<0.01 and ****P<0.0001 versus sham; #P<0.05, ###P<0.001 and ####P<0.0001 versus HF.

Figure 4
Postprandial increase in intact and total GLP-1 is impaired in heart failure rats and restored by the DPP4 inhibitor vildagliptin

Blood samples were collected after an overnight fast and 5 min after an oral glucose load of 2 g/kg from sham, HF rats, and HF rats treated with vildagliptin (HF + DPP4i). Experiments were conducted at baseline, i.e., before LV radiofrequency ablation for HF induction or sham operation (A–D) and at post-treatment (E–H). (A and E) Intact GLP-1 fasting (t = 0) and postprandial (t = 5) levels. (B and F) The percentage of increase in serum levels of intact GLP-1 after an oral glucose load. (C and G) Total GLP-1 fasting (t = 0) and postprandial (t = 5) levels. (D and H) The percentage of increase in serum levels of total GLP-1 after an oral glucose load. The values are the means ± SEM; **P<0.01 and ****P<0.0001 versus sham; #P<0.05, ###P<0.001 and ####P<0.0001 versus HF.

Post-treatment, the fasting serum concentration of intact GLP-1 was similar between HF and sham rats (Figure 4E). Figure 4E also shows that vildagliptin-treated HF rats displayed higher fasting levels of intact GLP-1 than those of both sham and vehicle-treated HF rats. In line with the patient data, the postprandial rise in intact GLP-1 was entirely blunted in HF rats (6 ± 4 vs. 62 ± 10%, P < 0.0001 vs. sham), and it was restored by vildagliptin treatment in HF rats (54 ± 6 vs. 62 ± 10%) (Figure 4E,F). The fasting levels of total GLP-1 were higher in HF than in sham rats (45 ± 2 vs. 34 ± 2 pM, P < 0.05) (Figure 4G). Additionally, no significant differences were observed in the fasting levels of total GLP-1 between vildagliptin-treated HF rats and sham rats (29 ± 3 vs. 34 ± 2 pM) (Figure 4G). Despite no differences in the concentration of total GLP-1 after oral glucose stimulation among the three groups of rats, the percentage of glucose-dependent total GLP-1 secretion was markedly reduced in HF rats compared with sham rats (9 ± 2 vs. 62 ± 10%, P < 0.0001) (Figure 4H). Surprisingly, the inhibition of DPP4 by vildagliptin not only potentiated the increase in intact biologically active GLP-1, but also restored the response of total GLP-1 secretion after oral glucose intake (91 ± 12 vs. 9 ± 2%, P < 0.0001 vs. vehicle-treated HF rats) (Figure 4H).

To corroborate the finding that postprandial increases in intact GLP-1 are blunted in severe HF in rats, we assessed whether the production of cAMP, a downstream signaling pathway activated by the binding of intact GLP-1 to its receptor [27–29], would be affected in these rats. Urine for cAMP determination was collected from the three groups of rats was for 4 h, starting immediately after oral glucose administration (2 g/kg) and blood withdrawal for GLP-1 measurements. As seen in Figure 5, changes in the serum levels of GLP-1 were accompanied by changes in urinary cAMP excretion. More specifically, at baseline, when no differences were found in the postprandial increase in serum intact GLP-1 (Figure 4A,B) among the three groups, urinary cAMP excretion remained unchanged (Figure 5A). Conversely, at post-treatment, vehicle-treated HF rats exhibited decreased urinary cAMP excretion compared with sham rats (50 ± 4 vs. 74 ± 6 nmol/kg/4 h, P < 0.01). As depicted in Figure 5B, vildagliptin treatment restored urinary cAMP excretion in HF rats to levels similar to those of the sham (72 ± 6 vs. 74 ± 6 nmol/kg/4 h). Collectively, the results of Figures 4 and 5 demonstrate that the postprandial bioavailability of GLP-1 is impaired in HF rats.

Urinary cAMP excretion is diminished in HF rats and is restored by treatment with the DPP4 inhibitor vildagliptin

Figure 5
Urinary cAMP excretion is diminished in HF rats and is restored by treatment with the DPP4 inhibitor vildagliptin

The three groups of rats were placed into metabolic cages after oral glucose administration (2 g/kg) and blood withdrawal for GLP-1 measurements for 4 h. (A) Urinary cAMP excretion was measured at baseline, i.e., before LV radiofrequency ablation for HF induction or sham operation. (B) Post-treatment urinary cAMP excretion from sham, HF rats, and HF rats treated with vildagliptin for 4 weeks (HF + DPP4i). The values are the mean ± SEM. The number of rats per experimental group is indicated by the bars; **P<0.01 vs. sham and ##P<0.01 versus HF.

Figure 5
Urinary cAMP excretion is diminished in HF rats and is restored by treatment with the DPP4 inhibitor vildagliptin

The three groups of rats were placed into metabolic cages after oral glucose administration (2 g/kg) and blood withdrawal for GLP-1 measurements for 4 h. (A) Urinary cAMP excretion was measured at baseline, i.e., before LV radiofrequency ablation for HF induction or sham operation. (B) Post-treatment urinary cAMP excretion from sham, HF rats, and HF rats treated with vildagliptin for 4 weeks (HF + DPP4i). The values are the mean ± SEM. The number of rats per experimental group is indicated by the bars; **P<0.01 vs. sham and ##P<0.01 versus HF.

Glucose homeostasis is disturbed in rats with established HF, and it is normalized by the inhibition of DPP4

The levels of serum glucose were similar among the three groups of rats (Figure 6A). However, the OGTT demonstrated a significant increase in the area under the curve (AUC) of the HF group (Figure 6B), suggesting the existence of some degree of glucose intolerance in HF rats despite normal fasting glucose levels. Besides, basal serum insulin levels were increased in HF rats compared with sham and vildagliptin-treated HF rats (Figure 6C). Moreover, vildagliptin-treated HF rats display an OGTT AUC and insulinemia similar to those of sham rats (Figure 6B,C). As shown in Figure 6D, GLUT4 expression in the heart was reduced in the HF group compared with the sham group, while the long-term inhibition of DPP4 by vildagliptin up-regulated GLUT4 expression in this tissue. These results suggest that both systemic and tissue glucose homeostasis seem to be affected in HF.

Glucose homeostasis in rats with HF

Figure 6
Glucose homeostasis in rats with HF

(A) Fasting serum glucose levels. (B) Time-dependent blood glucose response and the area under the curve (AUC). (C) Fasting insulin serum concentration. (D) GLUT4 protein expression in the hearts of sham, vildagliptin-treated, and untreated HF rats. The values are presented as the means ± SEM. The number of rats per experimental group is indicated by the bars; *P<0.05, **P<0.01, and ***P < 0.001 versus sham #P<0.05 and ###P<0.001 versus HF.

Figure 6
Glucose homeostasis in rats with HF

(A) Fasting serum glucose levels. (B) Time-dependent blood glucose response and the area under the curve (AUC). (C) Fasting insulin serum concentration. (D) GLUT4 protein expression in the hearts of sham, vildagliptin-treated, and untreated HF rats. The values are presented as the means ± SEM. The number of rats per experimental group is indicated by the bars; *P<0.05, **P<0.01, and ***P < 0.001 versus sham #P<0.05 and ###P<0.001 versus HF.

Discussion

The association between glucose abnormalities and HF has been recognized for a long time. However, whether and how HF affects glucose homeostasis, or vice-versa is largely unknown. The present study sheds light upon the mechanisms by which HF patients exhibit glucose abnormalities despite normal fasting glucose levels and how these disturbances may contribute to HF progression. More specifically, we demonstrate that non-diabetic HF patients and rats exhibit lower circulating postprandial levels of intact GLP-1 and higher insulin and glucose concentrations than those of controls. Moreover, our data suggest that the GLP-1 secretion mechanism after an oral nutrient load is also affected in HF, suggesting that GLP-1-producing cells might play an important role in the glucose abnormalities that are observed in HF patients. Finally, the increased expression and activity of DPP4 may play a role in this process since the inhibition of DPP4 by vildagliptin restores the bioavailability and secretion of GLP-1, glucose intolerance and cardiac function in HF rats.

After nutrient intake, one of the main mechanisms of blood glucose control and insulin secretion is the release of incretin hormones by the gut. An important incretin hormone responsible for blood glucose control is GLP-1. Several studies have shown that diabetic patients have an impaired GLP-1 response [30–32]. Herein, we show for the first time that non-diabetic HF patients and rats exhibit lower intact GLP-1 bioavailability as well as lower GLP-1 secretion after oral nutrient stimulation. Additionally, we demonstrate that non-diabetic HF patients have an increase in fasting levels of total GLP-1 despite similar levels of intact GLP-1 in comparison with those of control subjects. This finding suggests that, due to an increase in GLP-1 inactivation by DPP4, GLP-1-producing cells sustain normal levels of this hormone during fasting by releasing more GLP-1. In this regard, we observed a significant negative correlation between the postprandial increase in circulating intact GLP-1 and DPP4 activity in non-diabetic HF patients. Furthermore, after oral nutrient stimulation, the rise in intact GLP-1, and the total levels of this peptide were reduced in HF animals and patients. These findings suggest that GLP-1-producing cells in HF subjects are less sensitive to the nutrient-dependent GLP-1 release in response to oral glucose intake. Interestingly, DPP4 inhibition by vildagliptin abolished all disturbances in glucose homeostasis that were evaluated in HF rats.

Clinical and experimental studies have shown that the higher the activity of circulating DPP4, the poorer the cardiovascular outcomes in HF [17–19], suggesting that DPP4 might be involved in the pathophysiology of this syndrome. Therefore, in theory, one may postulate that DPP4 inhibition confers cardioprotection. Indeed, the results presented herein (Supplementary Tables S2 and S3), combined with those from other preclinical studies [16–18], and with a pilot study that demonstrates that DPP4 inhibition by sitagliptin improves the myocardial response to dobutamine and mitigates postischemic stunning in patients with coronary artery disease [33], support this hypothesis. On the other hand, all four large trials that have evaluated the cardiovascular outcomes of gliptins in diabetic patients, named SAVOR-TIMI 53 [34], EXAMINE [35], TECOS [36], and CARMELINA [37], failed to show any cardiovascular benefit when compared with controls. In fact, SAVOR-TIMI showed a 27% increase in the relative risk of hospitalization for HF in patients assigned to the saxagliptin group [34]. The neutral cardiovascular effects of the majority of gliptins might be explained at least in part by the fact that DPP4 metabolizes a broad array of substrates, including both cardioprotective and cardiodepressant peptides. For instance, neuropeptide Y (NPY), a DPP4 substrate that exerts vasoconstrictor and cardiodepressant actions, is known to be elevated in the plasma of HF patients [38]. Therefore, the beneficial ability of DPP4 inhibition to increase GLP-1 might be outweighed by the enhanced bioavailability of peptides, including NPY, that contribute to impaired cardiovascular function in the context of HF. It is worth mentioning that, as opposed to those of DPP4 inhibitors, the beneficial cardiovascular effects of the GLP-1 receptor (GLP-1R) agonist liraglutide observed in animal models have been successfully translated to clinical practice. In the LEADER trial [39], Marso et al. found that the risk of death from cardiovascular causes, nonfatal myocardial infarction, and nonfatal stroke was lower among patients with Type 2 diabetes who were randomly assigned to liraglutide than among those who were assigned to placebo.

Over the past few years, some studies have suggested that disturbances in glucose homeostasis, such as glucose intolerance and insulin resistance, are widespread in HF patients without diabetes [8,40]. Others have suggested that fasting glucose levels might underestimate the actual number of HF patients with some degree of glucose intolerance and that the OGTT might be more accurate in revealing glucose disturbance in a high proportion of undiagnosed HF patients [41,42]. In line with this concept, we demonstrate in the present study that neither HF patients nor HF rats display increased levels of fasting glucose. On the other hand, 10 weeks after cardiac injury, experimental HF rats display a slight but significant degree of glucose intolerance compared with that of control animals after the OGTT. Likewise, HF patients show higher levels of glucose than those of healthy control subjects 15 min after nutrient intake.

The increase in postprandial insulin in HF patients as well as fasting hyperinsulinemia and the reduced expression of the major cardiac isoform of the insulin-stimulated glucose transporter, i.e., GLUT4, in the rat model of HF are not only a sign of insulin resistance but may also represent an aggravating mechanism for the progression of HF. Indeed, insulin may exert deleterious effects on cardiac remodeling and vasculature, in addition to potentiating renal tubular sodium reabsorption, thereby worsening the clinical course of HF. Notably, although both have hyperglycemia, the HF phenotype is worse in patients with Type 2 diabetes than in patients with Type 1 diabetes, possibly because only the former have characteristic hyperinsulinemia, and the same is true for rodent models [40]. Thus, rather than hyperglycemia, hyperinsulinemia may be the most critical glucose disturbance that contributes to HF progression.

Interestingly, we found that the postprandial increase in intact GLP-1 correlates with markers of HF, such as serum BNP levels and the glomerular filtration rate (GFR). It is well documented that GLP-1 has cardiovascular protective effects, including beneficial effects on atherosclerosis, hypertension, and myocardial infarction [43–45]. Furthermore, several clinical and experimental studies have shown that GLP-1 receptor agonists exert beneficial renal effects [46]. The natriuretic effects of GLP-1 involve the inhibition of sodium–hydrogen exchanger isoform 3 (NHE3) in the renal proximal tubule [29]. However, hemodynamic effects such as vasodilation and changes in GFR have also been reported [29,47]. In fact, increased NHE3 expression and activity appear to play a role in the pathogenesis of fluid retention, pulmonary congestion, and resistance to endogenous natriuretic peptides in HF [48], which could explain the benefit of the suppression of NHE3 arising from increased GLP-1 bioavailability [49]. Notably, acute treatment with the GLP-1 receptor antagonist exendin-9 significantly reduces urinary flow, urinary sodium excretion, tubular sodium reabsorption, and GFR in rats, indicating that GLP-1 is a physiologically relevant hormone for sodium balance maintenance [28]. Thus, lower postprandial GLP-1 bioavailability due to increased circulating DPP4 levels and decreased GLP-1 secretion after nutrient intake might contribute to glucose disorders, extracellular fluid volume abnormalities, and, therefore, the aggravation of HF.

Collectively, our data demonstrate that the postprandial increase in intact GLP-1 bioavailability and GLP-1 secretion is blunted in non-diabetic patients and rats with HF. The pronounced impairment in intact GLP-1 bioavailability after meal intake correlates with poor prognostic factors and may contribute to establishing a vicious cycle between glucose disturbance and HF development and progression. It remains to be determined whether the disturbance in glucose homeostasis and the worsening of cardiac function are concomitant outcomes in the natural history of HF or whether some cause–effect relationship exists.

Clinical perspectives

  • Type 2 diabetes (T2D) is a major risk factor for heart failure (HF), whereas HF severity predicts the risk of developing diabetes after myocardial infarction. The bioavailability of intact glucagon-like peptide-1 (GLP-1), an incretin hormone that plays a determinant role in blood glucose homeostasis and exerts cardioprotective effects, is reduced in Type 2 diabetes. We hypothesized that the reduced bioavailability of GLP-1 might also be affected in severe HF.

  • We provide evidence that non-diabetic patients with HF and rats exhibit lower intact GLP-1 bioavailability after oral nutrient stimulation. Moreover, we found that impaired GLP-1 bioavailability after meal intake correlates with poor prognostic factors and may contribute to the establishment of a vicious cycle between glucose disturbance and HF development and progression.

  • Disturbed glucose metabolism associated with blunted postprandial GLP-1 increase could be feasibly screened by oral glucose tolerance tests to improve the follow-up of HF syndrome.

Competing Interests

The authors declare that there are no competing interests associated with the manuscript.

Funding

This work was supported by the São Paulo State Research Foundation (FAPESP) [grant numbers 2013/10619-8 and 2016/22140-7 (to A.C.G.) and 2013/17368-0 (to J.E.K.)].

Author Contribution

Conceptualization: A.C.G.; Data curation: D.F.A.J., F.L.M., L.J., R.D., and R.O.C.; Formal analysis: D.F.A.J., F.L.M., L.J., R.D., and L.S.; Funding acquisition: A.C.G. and J.E.K. Investigation: D.F.A.J., F.L.M., L.J., R.D., and R.O.C.; Methodology: T.A.S., E.L.A., and P.J.F.T.; Project administration: A.C.G. Resources: A.C.G., P.J.F.T., A.C.P., and J.E.K. Supervision: A.C.G., P.J.F.T., L.H.W.G., A.C.P., and J.E.K. Validation: L.H.G. and A.C.G.; Visualization: D.F.A.J., L.S., L.H.W.G., and A.C.G.; Writing – original draft: D.F.A.J., T.A.S., L.S. Writing – review & editing: A.C.G. and L.H.W.G.

Ethics Approval

The ethics committee of the University of São Paulo School of Medicine approved the protocol for human (Approval No. SDC 2368/03/162 of January 23rd, 2014) and animal studies (Approval No. 050/17 of May 8th, 2017).

Abbreviations

     
  • AUC

    area under the curve

  •  
  • BNP

    brain natriuretic peptide

  •  
  • CARMELINA

    Cardiovascular and Renal Microvascular Outcome Study With Linagliptin in Patients With Type 2 Diabetes Mellitus

  •  
  • DPP4

    dipeptidyl peptidase 4

  •  
  • ELISA

    enzyme-linked immunosorbent assay

  •  
  • EXAMINE

    examination of cardiovascular outcomes with alogliptin versus standard of care in patients with type 2 diabetes mellitus and acute coronary syndrome

  •  
  • GFR

    glomerular filtration rate

  •  
  • GLP-1

    glucagon-like peptide-1

  •  
  • GLUT4

    glucose transporter type 4

  •  
  • HF

    heart failure

  •  
  • LEADER

    Liraglutide and Cardiovascular Outcomes in Type 2 Diabetes

  •  
  • NHE3

    sodium–hydrogen exchanger isoform 3

  •  
  • OGTT

    oral glucose tolerance test

  •  
  • SAVOR-TIMI 53

    saxagliptin and cardiovascular outcomes in patients with type 2 diabetes mellitus

  •  
  • TECOS

    Trial Evaluating Cardiovascular Outcomes with Sitagliptin

References

References
1.
Yancy
C.W.
,
Jessup
M.
,
Bozkurt
B.
,
Butler
J.
,
Casey
D.E.
,
Drazner
M.H.
et al.
(
2013
)
2013 ACCF/AHA guideline for the management of heart failure: a report of the American College of Cardiology Foundation/American Heart Association Task Force on Practice Guidelines
.
J. Am. Coll. Cardiol.
62
,
e147
e239
[PubMed]
2.
Kannel
W.B.
,
Hjortland
M.
and
Castelli
W.P.
(
1974
)
Role of diabetes in congestive heart failure: the Framingham study
.
Am. J. Cardiol.
34
,
29
34
[PubMed]
3.
McKee
P.A.
,
Castelli
W.P.
,
McNamara
P.M.
and
Kannel
W.B.
(
1971
)
The natural history of congestive heart failure: the Framingham study
.
N. Engl. J. Med.
285
,
1441
1446
[PubMed]
4.
Akiyama
E.
,
Sugiyama
S.
,
Matsubara
J.
,
Kurokawa
H.
,
Konishi
M.
,
Nozaki
T.
et al.
(
2015
)
Decreased plasma levels of active glucagon-like peptide-1 in coronary artery disease
.
J. Am. Coll. Cardiol.
65
,
754
755
[PubMed]
5.
Norhammar
A.
,
Tenerz
A.
,
Nilsson
G.
,
Hamsten
A.
,
Efendíc
S.
,
Rydén
L.
et al.
(
2002
)
Glucose metabolism in patients with acute myocardial infarction and no previous diagnosis of diabetes mellitus: a prospective study
.
Lancet
359
,
2140
2144
[PubMed]
6.
Witteles
R.M.
,
Tang
W.H.
,
Jamali
A.H.
,
Chu
J.W.
,
Reaven
G.M.
and
Fowler
M.B.
(
2004
)
Insulin resistance in idiopathic dilated cardiomyopathy: a possible etiologic link
.
J. Am. Coll. Cardiol.
44
,
78
81
[PubMed]
7.
Berry
C.
,
Brett
M.
,
Stevenson
K.
,
McMurray
J.J.
and
Norrie
J.
(
2008
)
Nature and prognostic importance of abnormal glucose tolerance and diabetes in acute heart failure
.
Heart
94
,
296
304
[PubMed]
8.
Tenenbaum
A.
,
Motro
M.
,
Fisman
E.Z.
,
Leor
J.
,
Freimark
D.
,
Boyko
V.
et al.
(
2003
)
Functional class in patients with heart failure is associated with the development of diabetes
.
Am. J. Med.
114
,
271
275
[PubMed]
9.
Sud
M.
,
Wang
X.
,
Austin
P.C.
,
Lipscombe
L.L.
,
Newton
G.E.
,
Tu
J.V.
et al.
(
2015
)
Presentation blood glucose and death, hospitalization, and future diabetes risk in patients with acute heart failure syndromes
.
Eur. Heart J.
36
,
924
931
[PubMed]
10.
Zareini
B.
,
Rørth
R.
,
Holt
A.
,
Mogensen
U.M.
,
Selmer
C.
,
Gislason
G.
et al.
(
2019
)
Heart failure and the prognostic impact and incidence of new-onset of diabetes mellitus: a nationwide cohort study
.
Cardiovasc. Diabetol.
18
,
79
[PubMed]
11.
Andersson
C.
,
Norgaard
M.L.
,
Hansen
P.R.
,
Fosbøl
E.L.
,
Schmiegelow
M.
,
Weeke
P.
et al.
(
2010
)
Heart failure severity, as determined by loop diuretic dosages, predicts the risk of developing diabetes after myocardial infarction: a nationwide cohort study
.
Eur. J. Heart Fail.
12
,
1333
1338
[PubMed]
12.
Drucker
D.J.
(
2005
)
Biologic actions and therapeutic potential of the proglucagon-derived peptides
.
Nat. Clin. Pract. Endocrinol. Metab.
1
,
22
31
[PubMed]
13.
Elliott
R.M.
,
Morgan
L.M.
,
Tredger
J.A.
,
Deacon
S.
,
Wright
J.
and
Marks
V.
(
1993
)
Glucagon-like peptide-1 (7-36)amide and glucose-dependent insulinotropic polypeptide secretion in response to nutrient ingestion in man: acute post-prandial and 24-h secretion patterns
.
J. Endocrinol.
138
,
159
166
[PubMed]
14.
Ahrén
B.
and
Holst
J.J.
(
2001
)
The cephalic insulin response to meal ingestion in humans is dependent on both cholinergic and noncholinergic mechanisms and is important for postprandial glycemia
.
Diabetes
50
,
1030
1038
[PubMed]
15.
Holst
J.J.
(
2007
)
The physiology of glucagon-like peptide 1
.
Physiol. Rev.
87
,
1409
1439
[PubMed]
16.
Salles
T.A.
,
dos Santos
L.
,
Barauna
V.G.
and
Girardi
A.C.C.
(
2015
)
Potential role of dipeptidyl peptidase IV in the pathophysiology of heart failure
.
Int. J. Mol. Sci.
16
,
4226
4249
[PubMed]
17.
Dos Santos
L.
,
Salles
T.A.
,
Arruda-Junior
D.F.
,
Campos
L.C.G.
,
Pereira
A.C.
,
Barreto
A.L.T.
et al.
(
2013
)
Circulating dipeptidyl peptidase IV activity correlates with cardiac dysfunction in human and experimental heart failure
.
Circulation: Heart Fail.
6
,
1029
1038
[PubMed]
18.
Shigeta
T.
,
Aoyama
M.
,
Bando
Y.K.
,
Monji
A.
,
Mitsui
T.
,
Takatsu
M.
et al.
(
2012
)
Dipeptidyl peptidase-4 modulates left ventricular dysfunction in chronic heart failure via angiogenesis-dependent and -independent actions
.
Circulation
126
,
1838
1851
[PubMed]
19.
Lourenço
P.
,
Friões
F.
,
Silva
N.
,
Guimarães
J.T.
and
Bettencourt
P.
(
2013
)
Dipeptidyl peptidase IV and mortality after an acute heart failure episode
.
J. Cardiovasc. Pharmacol.
62
,
138
142
[PubMed]
20.
Hunt
S.A.
,
Abraham
W.T.
,
Chin
M.H.
,
Feldman
A.M.
,
Francis
G.S.
,
Ganiats
T.G.
et al.
(
2005
)
ACC/AHA 2005 Guideline Update for the Diagnosis and Management of Chronic Heart Failure in the Adult: a report of the American College of Cardiology/American Heart Association Task Force on Practice Guidelines (Writing Committee to Update the 2001 Guidelines for the Evaluation and Management of Heart Failure): developed in collaboration with the American College of Chest Physicians and the International Society for Heart and Lung Transplantation: endorsed by the Heart Rhythm Society
.
Circulation
112
,
e154
e235
[PubMed]
21.
Antonio
E.L.
,
Dos Santos
A.A.
,
Araujo
S.R.
,
Bocalini
D.S.
,
Dos Santos
L.
,
Fenelon
G.
et al.
(
2009
)
Left ventricle radio-frequency ablation in the rat: a new model of heart failure due to myocardial infarction homogeneous in size and low in mortality
.
J. Card. Fail.
15
,
540
548
[PubMed]
22.
Giannocco
G.
,
Oliveira
K.C.
,
Crajoinas
R.O.
,
Venturini
G.
,
Salles
T.A.
,
Fonseca-Alaniz
M.H.
et al.
(
2013
)
Dipeptidyl peptidase IV inhibition upregulates GLUT4 translocation and expression in heart and skeletal muscle of spontaneously hypertensive rats
.
Eur. J. Pharmacol.
698
,
74
86
[PubMed]
23.
Arruda-Junior
D.F.
,
Martins
F.L.
,
Dariolli
R.
,
Jensen
L.
,
Antonio
E.L.
,
Dos Santos
L.
et al.
(
2016
)
Dipeptidyl Peptidase IV Inhibition Exerts Renoprotective Effects in Rats with Established Heart Failure
.
Front. Physiol.
7
,
293
[PubMed]
24.
Pacheco
B.P.M.
,
Crajoinas
R.O.
,
Couto
G.K.
,
Davel
A.P.C.
,
Lessa
L.M.
,
Rossoni
L.V.
et al.
(
2011
)
Dipeptidyl peptidase IV inhibition attenuates blood pressure rising in young spontaneously hypertensive rats
.
J. Hypertens.
29
,
520
528
[PubMed]
25.
de Almeida Salles
T.
,
Zogbi
C.
,
de Lima
T.M.
,
de Godoi Carneiro
C.
,
Garcez
A.T.
,
Barbeiro
H.V.
et al.
(
2016
)
The contributions of dipeptidyl peptidase IV to inflammation in heart failure
.
Am. J. Physiol. Heart Circ. Physiol.
310
,
H1760
H1772
[PubMed]
26.
Crajoinas
R.O.
,
Lessa
L.M.
,
Carraro-Lacroix
L.R.
,
Davel
A.P.
,
Pacheco
B.P.
,
Rossoni
L.V.
et al.
(
2010
)
Posttranslational mechanisms associated with reduced NHE3 activity in adult vs. young prehypertensive SHR
.
Am. J. Physiol. Renal. Physiol.
299
,
F872
F881
[PubMed]
27.
Thorens
B.
(
1992
)
Expression cloning of the pancreatic beta cell receptor for the gluco-incretin hormone glucagon-like peptide 1
.
Proc. Natl. Acad. Sci. U.S.A.
89
,
8641
8645
[PubMed]
28.
Farah
L.X.
,
Valentini
V.
,
Pessoa
T.D.
,
Malnic
G.
,
McDonough
A.A.
and
Girardi
A.C.
(
2016
)
The physiological role of glucagon-like peptide-1 in the regulation of renal function
.
Am. J. Physiol. Renal. Physiol.
310
,
F123
F127
[PubMed]
29.
Crajoinas
R.O.
,
Oricchio
F.T.
,
Pessoa
T.D.
,
Pacheco
B.P.M.
,
Lessa
L.M.A.
,
Malnic
G.
et al.
(
2011
)
Mechanisms mediating the diuretic and natriuretic actions of the incretin hormone glucagon-like peptide-1
.
Am. J. Physiol.-Renal Physiol.
301
,
F355
F363
[PubMed]
30.
Campbell
J.E.
and
Drucker
D.J.
(
2013
)
Pharmacology, physiology, and mechanisms of incretin hormone action
.
Cell Metab.
17
,
819
837
[PubMed]
31.
Perley
M.J.
and
Kipnis
D.M.
(
1967
)
Plasma insulin responses to oral and intravenous glucose: studies in normal and diabetic sujbjects
.
J. Clin. Invest.
46
,
1954
1962
[PubMed]
32.
Holst
J.J.
,
Knop
F.K.
,
Vilsbøll
T.
,
Krarup
T.
and
Madsbad
S.
(
2011
)
Loss of incretin effect is a specific, important, and early characteristic of type 2 diabetes
.
Diabetes Care
34
,
S251
S257
[PubMed]
33.
Read
P.A.
,
Khan
F.Z.
,
Heck
P.M.
,
Hoole
S.P.
and
Dutka
D.P.
(
2010
)
DPP-4 inhibition by sitagliptin improves the myocardial response to dobutamine stress and mitigates stunning in a pilot study of patients with coronary artery disease
.
Circ. Cardiovasc. Imaging
3
,
195
201
[PubMed]
34.
Scirica
B.M.
,
Bhatt
D.L.
,
Braunwald
E.
,
Steg
P.G.
,
Davidson
J.
,
Hirshberg
B.
et al.
(
2013
)
Saxagliptin and cardiovascular outcomes in patients with type 2 diabetes mellitus
.
N. Engl. J. Med.
369
,
1317
1326
[PubMed]
35.
White
W.B.
,
Bakris
G.L.
,
Bergenstal
R.M.
,
Cannon
C.P.
,
Cushman
W.C.
,
Fleck
P.
et al.
(
2011
)
EXamination of cArdiovascular outcoMes with alogliptIN versus standard of carE in patients with type 2 diabetes mellitus and acute coronary syndrome (EXAMINE): a cardiovascular safety study of the dipeptidyl peptidase 4 inhibitor alogliptin in patients with type 2 diabetes with acute coronary syndrome
.
Am. Heart J.
162
,
620.e1
626.e1
36.
Green
J.B.
,
Bethel
M.A.
,
Armstrong
P.W.
,
Buse
J.B.
,
Engel
S.S.
,
Garg
J.
et al.
(
2015
)
Effect of Sitagliptin on Cardiovascular Outcomes in Type 2 Diabetes
.
N. Engl. J. Med.
373
,
232
242
[PubMed]
37.
Rosenstock
J.
,
Perkovic
V.
,
Johansen
O.E.
,
Cooper
M.E.
,
Kahn
S.E.
,
Marx
N.
et al.
(
2019
)
Effect of Linagliptin vs Placebo on Major Cardiovascular Events in Adults With Type 2 Diabetes and High Cardiovascular and Renal Risk: The CARMELINA Randomized Clinical Trial
.
JAMA
321
,
69
79
[PubMed]
38.
Maisel
A.S.
,
Scott
N.A.
,
Motulsky
H.J.
,
Michel
M.C.
,
Boublik
J.H.
,
Rivier
J.E.
et al.
(
1989
)
Elevation of plasma neuropeptide Y levels in congestive heart failure
.
Am. J. Med.
86
,
43
48
[PubMed]
39.
Marso
S.P.
,
Daniels
G.H.
,
Brown-Frandsen
K.
,
Kristensen
P.
,
Mann
J.F.
,
Nauck
M.A.
et al.
(
2016
)
Liraglutide and Cardiovascular Outcomes in Type 2 Diabetes
.
N. Engl. J. Med.
375
,
311
322
[PubMed]
40.
Packer
M.
(
2018
)
Potentiation of Insulin Signaling Contributes to Heart Failure in Type 2 Diabetes: A Hypothesis Supported by Both Mechanistic Studies and Clinical Trials
.
JACC Basic Transl. Sci.
3
,
415
419
[PubMed]
41.
Egstrup
M.
,
Schou
M.
,
Gustafsson
I.
,
Kistorp
C.N.
,
Hildebrandt
P.R.
and
Tuxen
C.D.
(
2011
)
Oral glucose tolerance testing in an outpatient heart failure clinic reveals a high proportion of undiagnosed diabetic patients with an adverse prognosis
.
Eur. J. Heart Fail.
13
,
319
326
[PubMed]
42.
Stevens
A.L.
,
Hansen
D.
,
Vandoren
V.
,
Westerlaken
R.
,
Creemers
A.
,
Eijnde
B.O.
et al.
(
2014
)
Mandatory oral glucose tolerance tests identify more diabetics in stable patients with chronic heart failure: a prospective observational study
.
Diabetol. Metab. Syndr.
6
,
44
[PubMed]
43.
Rakipovski
G.
,
Rolin
B.
,
Nøhr
J.
,
Klewe
I.
,
Frederiksen
K.S.
,
Augustin
R.
et al.
(
2018
)
The GLP-1 Analogs Liraglutide and Semaglutide Reduce Atherosclerosis in ApoE
.
JACC Basic Transl. Sci.
3
,
844
857
[PubMed]
44.
Poornima
I.
,
Brown
S.B.
,
Bhashyam
S.
,
Parikh
P.
,
Bolukoglu
H.
and
Shannon
R.P.
(
2008
)
Chronic glucagon-like peptide-1 infusion sustains left ventricular systolic function and prolongs survival in the spontaneously hypertensive, heart failure-prone rat
.
Circ. Heart Fail.
1
,
153
160
[PubMed]
45.
Boyle
J.G.
,
Livingstone
R.
and
Petrie
J.R.
(
2018
)
Cardiovascular benefits of GLP-1 agonists in type 2 diabetes: a comparative review
.
Clin. Sci. (Lond.)
132
,
1699
1709
[PubMed]
46.
Muskiet
M.H.A.
,
Tonneijck
L.
,
Smits
M.M.
,
van Baar
M.J.B.
,
Kramer
M.H.H.
,
Hoorn
E.J.
et al.
(
2017
)
GLP-1 and the kidney: from physiology to pharmacology and outcomes in diabetes
.
Nat. Rev. Nephrol.
13
,
605
628
[PubMed]
47.
Savignano
F.A.
,
Crajoinas
R.O.
,
Pacheco
B.P.M.
,
Campos
L.C.G.
,
Shimizu
M.H.M.
,
Seguro
A.C.
et al.
(
2017
)
Attenuated diuresis and natriuresis in response to glucagon-like peptide-1 in hypertensive rats are associated with lower expression of the glucagon-like peptide-1 receptor in the renal vasculature
.
Eur. J. Pharmacol.
811
,
38
47
[PubMed]
48.
Inoue
B.H.
,
dos Santos
L.
,
Pessoa
T.D.
,
Antonio
E.L.
,
Pacheco
B.P.M.
,
Savignano
F.A.
et al.
(
2012
)
Increased NHE3 abundance and transport activity in renal proximal tubule of rats with heart failure
.
Am. J. Physiol.-Regul. Integr. Comp. Physiol.
302
,
R166
R174
49.
Packer
M.
(
2017
)
Activation and Inhibition of Sodium-Hydrogen Exchanger Is a Mechanism That Links the Pathophysiology and Treatment of Diabetes Mellitus With That of Heart Failure
.
Circulation
136
,
1548
1559
[PubMed]

Author notes

*

These authors contributed equally to this work.