Dapagliflozin Impairs the Suppression of Endogenous Glucose Production in Type 2 Diabetes Following Oral Glucose

SGLT2 inhibitors (SGLT2i) reduce plasma glucose by producing glucosuria (1–3), but their glucose-lowering efficacy is partially offset by an increase in endogenous glucose production (EGP) (2–4), which accounts for ∼50% the amount of glucose excreted in urine (2,5). In response to acute (3) and chronic (4) SGLT2i administration, the fasting plasma glucagon concentration increases and the plasma insulin concentration decreases. However, these hormonal changes cannot explain the rise in basal EGP, which persists when plasma glucagon and insulin are clamped at fasting levels (6). Further, the increase in basal EGP is not abolished when SGLT2i is administered with a glucagon-like peptide 1 receptor antagonist (GLP-1 RA) or dipeptidyl peptidase 4 inhibitor, which increase plasma insulin and decrease plasma glucagon (7,8). Recently, we have shown that the SGLT2i-induced rise in EGP is mediated, in part, by activation of renal nerves, which transmit a signal to the liver to stimulate hepatic glucose output (9).

All previous studies examining the effect of SGLT2i on EGP have been carried out under fasting conditions. No previous study has examined the effect of SGLT2i on EGP suppression following a meal, nor have the tissues responsible for the disposition of the glucose load under these conditions been determined. These are of particular importance since in patients with type 2 diabetes and in individuals with prediabetes, EGP suppression following oral glucose is impaired (10,11). In the current study, we have used a double-tracer technique (10,12,13) to test the hypothesis that the SGLT-2i dapagliflozin (DAPA) impairs the suppression of EGP following an oral glucose load. In addition, we sought to determine the tissues responsible for glucose disposal after the ingestion of glucose. The data presented in this article represent analysis and interpretation of a fraction of the entire study reported in the clinical trial referenced (ClinicalTrials.gov NCT03331289).

Forty-eight subjects with type 2 diabetes participated in two protocols. Prior to entering into the study, patients were treated with diet alone (n = 3), a stable (>3 months) dose of metformin (n = 26), or metformin plus sulfonylurea (n = 19). Except for diabetes, all subjects were in good general health based on medical history, physical examination, screening blood tests, urinalysis, and electrocardiogram. Baseline clinical, anthropometric, and laboratory data are presented in Table 1. Weight was stable (±1.5 kg) in all subjects for at least 3 months prior to study, and no subject participated in any excessively heavy exercise program. Subjects taking drugs known to affect glucose metabolism (other than metformin and sulfonylurea) were excluded. The study was approved by the University of Texas Health San Antonio Institutional Review Board, and informed written consent was obtained from all participants.

Subjects with type 2 diabetes were randomly assigned 1:1:1:1 to receive a single dose of: 1) placebo (PCB); 2) exenatide (EXE), 5 µg s.c.; 3) DAPA, 10 mg; 4) DAPA 10 mg plus EXE 5 µg, and EGP was measured with [3-³H]-glucose.

Subjects received a double-tracer ([1-¹⁴C]-glucose and [3-³H]-glucose) oral glucose tolerance test (OGTT) to measure EGP and glucose kinetics. Prior to OGTT, subjects received a single dose of PCB, EXE, DAPA, or EXE plus DAPA as described previously. All studies were performed at the Clinical Research Center of Texas Diabetes Institute at 0700 h following a 10-h overnight fast.

During protocol I, subjects received 8-h prime (40 µCi × fasting plasma glucose [FPG]/100) continuous (0.40 µCi/min) infusion of [3-³H]-glucose (PerkinElmer, Boston, MA) via antecubital vein catheter to measure EGP. After a 3-h tracer equilibration period (at time zero), subjects received in random order: 1) oral PCB; 2) EXE, 5 µg s.c.; 3) DAPA, 10 mg; and 4) DAPA, 10 mg plus EXE 5 µg. Arterialized venous blood samples (“hot box” technique) were drawn at −30, −20, −10, −5, 0, and every 15–20 min thereafter; for 5 h, plasma glucose, insulin, C-peptide, glucagon concentrations, and tritiated glucose-specific activity were measured. Urine was collected before and post–drug administration for measurement of urinary glucose excretion rate (UGE). Plasma insulin was determined with IRMA assay (Bioz, Louvain, Belgium) and C-peptide and glucagon with radioimmunoassay (EMD Millipore, Billerica, MA). The antibody used in the glucagon assay is specific for pancreatic glucagon with a cross-reactivity to oxyntomodulin (“gut glucagon”) <0.1%.

During protocol II, all subjects had a 5-h double-tracer OGTT (10,12). A 7.5-h prime continuous infusion of [3-³H]-glucose was administered as in protocol I. After 3 h, subjects received: oral PCB; EXE 5 µg s.c.; DAPA 10 mg; or DAPA 10 mg plus EXE. At 3.5 h (time zero), subjects ingested 75 g of glucose containing 100 µCi of [1-¹⁴C]-glucose (PerkinElmer). Urine was collected prior to and post–drug administration for measurement of UGE. Between protocols I and II, subjects were asked to maintain their stable treatment with diet/exercise and either metformin or metformin/sulfonylurea.

During protocol I, the primary end point was change in EGP from baseline (−30 to 0 min) to last hour (240–300-min time period) following drug versus PCB. Under steady-state postabsorptive conditions, the baseline rate of EGP equals [3-³H]-glucose infusion rate divided by steady-state plasma tritiated glucose-specific activity (10,12). After drug administration, nonsteady conditions for [3-³H]-glucose–specific activity prevailed, and rates of total body glucose appearance (R_a = EGP) and disappearance (R_d) were calculated using Steele’s equation (14). During protocol II, to calculate the rate of oral glucose appearance (RaO), [1-¹⁴C]-glucose plasma radioactivity was divided by the specific activity of the glucose drink to obtain the plasma oral glucose concentration that would be attained in the systemic circulation if the sole source of glucose was the oral load and by subtracting the plasma endogenous glucose concentration. The calculated endogenous glucose concentration and [3-³H]-glucose data were then used to compute EGP in peripheral plasma. RaO was obtained as the difference between total and endogenous R_a rates (10,12). Splanchnic (represents hepatic plus gastrointestinal tissues) glucose uptake was calculated as the difference between ingested glucose (75 g) and RaO.

Changes from baseline in EGP following drug administration were compared with changes from baseline in the PCB study using paired t test. Changes from baseline to mean values for all glucose kinetics parameters post–glucose ingestion were also compared using a paired t test. Differences in EGP and all glucose kinetic parameters between results after each drug were compared among the four treatment groups using ANOVA. Post hoc testing was done with Bonferroni correction. Similar comparisons were made for changes in plasma insulin and glucagon concentrations. Values are presented as mean ± SEM, unless otherwise noted. A P value <0.05 was considered statistically significant.

The data sets generated during and/or analyzed during the current study are available from the corresponding author upon reasonable request.

Following PCB ingestion, fasting plasma glucose decreased from 177 ± 3 to 143 ± 2 mg/dL, while after DAPA, the decrease in FPG (160 ± 6 to 111 ± 3 mg/dL) at 300 min was greater than PCB (P < 0.05) (Fig. 1A). At 300 min, the decrease in plasma glucose with EXE (164 ± 4 to 101 ± 2 mg/dL) and with DAPA/EXE (164 ± 4 to 107 ± 2 mg/dL) was similar to DAPA and greater than PCB (P < 0.05) (Fig. 1A). However, the decrement in plasma glucose AUC was significantly greater (P < 0.01) with both EXE and EXE/DAPA compared with DAPA alone and PCB.

Following PCB, EGP showed a steady decrease from 2.16 ± 0.15 to a mean value of 1.57 ± 0.08 mg/kg/min, while EGP did not change significantly (2.04 ± 0.17 vs. 1.94 ± 0.18 mg/kg/min) following DAPA (P < 0.05) (Fig. 1B). After EXE, EGP decreased from 2.13 ± 0.16 to 1.58 ± 0.03 mg/kg/min (P < 0.05), whereas after DAPA/EXE, there was an initial decrease followed by an increase in EGP during the last hour (2.13 ± 0.10 vs. 2.09 ± 0.03 mg/kg/min; P = NS). As a result, the mild decrease in EGP was similar in DAPA and DAPA/EXE, a change that was significantly (P < 0.05) attenuated compared with both PCB and EXE.

During the entire 300-min period, plasma insulin concentration did not change significantly and was similar in PCB (12 ± 1 vs. 10 ± 1 µU/mL) and DAPA (14 ± 1 vs. 11 ± 1 µU/mL) groups. In both the EXE (14 ± 1 vs. 16 ± 1 µU/mL) and DAPA/EXE (13 ± 1 vs. 16 ± 1 µU/mL) groups, plasma insulin increased similarly (both P < 0.01 vs. PCB and DAPA) (Fig. 1C). Similarly, baseline plasma C-peptide did not change in either PCB (3.0 ± 0.1 vs. 2.9 ± 0.1 ng/ml) or DAPA (3.5 ± 0.1 vs. 3.2 ± 0.1 ng/mL) groups. After EXE (3.5 ± 0.2 vs. 4.1 ± 0.2 ng/mL) and DAPA/EXE (3.2 ± 0.1 vs. 3.5 ± 0.1 ng/mL), plasma C-peptide increased, and changes were significantly greater than with PCB and DAPA (P < 0.05). Plasma glucagon decreased slightly with PCB (51 ± 4 vs. 46 ± 2 pg/mL), a decline that was attenuated (P < 0.05) in DAPA (53 ± 3 vs. 50 ± 2 pg/mL). Plasma glucagon decreased significantly (P < 0.05) with both EXE (49 ± 2 to 37 ± 2 pg/mL) and DAPA/EXE (53 ± 3 to 42 ± 2 pg/mL) (Fig. 1D).

During double-tracer OGTT, subjects received a single dose of medication 30 min prior to glucose ingestion. Following PCB, PG increased from 169 ± 3 to a mean of 255 ± 4 mg/dL during the 300-min post-OGTT period. In the DAPA group, the increase in plasma glucose was attenuated (146 ± 4 vs. 200 ± 6 mg/dL) versus PCB (P < 0.05). In the EXE group, the rise in PG was significantly (P < 0.001) less than with PCB (152 ± 4 vs. 178 ± 4 mg/dL), while in the DAPA/EXE group, baseline PG actually decreased after glucose ingestion (178 ± 5 to 160 ± 4 mg/dL) (P < 0.001 vs. DAPA and PCB and P < 0.05 vs. EXE) (Fig. 2A).

Following PCB ingestion, there was a progressive decline in EGP (Fig. 2B). In the DAPA group, EGP remained unchanged during the first 2 h (Δ = 0.13 ± 0.05 mg/kg/min) and decreased modestly during the last 3 h (Δ = −0.99 ± 0.20) (Fig. 2B). In the EXE group, EGP decreased steadily during the first 2 h (Δ= −0.92 ± 0.15 mg/kg/min) and reached a nadir during the last 3 h (Δ= −1.48 ± 0.08). In the study when DAPA was administered with EXE, the decrease in EGP over the 300-min time period was significantly impaired versus EXE alone (−0.80 ± 0.06 vs. −1.20 ± 0.09; P < 0.01).

Following oral glucose, plasma insulin increased similarly in the PCB (13.3 ± 0.8 to 32.3 ± 2.0 µU/mL) and DAPA (13.6 ± 0.7 to 27.2 ± 1.4 µU/mL) groups (Fig. 2C). In the EXE group, the rise in plasma insulin (14.1 ± 0.9 to 42.4 ± 2.2 µU/mL) was greater (P < 0.05) versus both PCB and DAPA. Administration of DAPA with EXE significantly attenuated the rise in plasma insulin (14.2 ± 0.9 to 30.1 ± 2.6 µU/mL) compared with EXE (Fig. 2C). Plasma C-peptide showed a similar pattern to those of plasma insulin (PCB, 3.3 ± 0.1 to 6.2 ± 0.3; DAPA, 3.2 ± 0.1 to 6.2 ± 0.2; EXE, 3.6 ± 0.2 to 8.7 ± 0.4; and DAPA/EXE, 3.2 ± 0.1 to 6.0 ± 0.2 ng/mL) (P < 0.05, EXE vs. DAPA/EXE).

In the PCB group, baseline plasma glucagon decreased slightly from 52 ± 2 to 45 ± 1 pg/mL after oral glucose and did not change in DAPA (53 ± 3 vs. 55 ± 2 pg/mL) (P < 0.05). The decrease in plasma glucagon with EXE (44 ± 3 to 32 ± 2 pg/mL) was attenuated in the DAPA/EXE group (44 ± 3 to 39 ± 2 pg/mL) (P < 0.05 vs. DAPA/EXE) (Fig. 2D). The insulin-to-glucagon (INS/GCN) ratio in PCB increased from 0.26 ± 0.03 (baseline) to a mean of 0.71 ± 0.06 μU/mL per pg/mL (Δ = 0.45) during the 300-min post–oral glucose, whereas with DAPA, the increase in INS/GCN ratio was blunted (0.26 ± 0.02 to 0.50 ± 0.04 μU/mL per pg/mL [Δ = 0.24]; P < 0.05). In the EXE group, baseline INS/GCN ratio increased significantly (0.32 ± 0.03 to 1.31 ± 0.08 [Δ = 0.99]) during 300-min post–oral glucose challenge (P < 0.001 vs. all), while with DAPA/EXE, the rise in INS/GCN ratio was attenuated (0.32 ± 0.03 to 0.78 ± 0.08 [Δ = 0.46]) (P < 0.01 vs. EXE). Similar trends were observed with the C-peptide/GCN ratio during the 300-min post–oral glucose in all groups (PCB, 0.06 ± 0.02 to 0.14 ± 0.03; DAPA, 0.06 ± 0.02 to. 0.11 ± 0.03; EXE, 0.08 ± 0.03 to 0.27 ± 0.04; DAPA/EXE, 0.07 ± 0.02 to 0.15 ± 0.04).

Total R_a in the systemic circulation in the PCB group increased from 2.30 ± 0.05 (baseline) to a mean of 3.98 ± 0.13 mg/kg/min during the 300 min following glucose ingestion (Table 2). During the same time period with DAPA, total R_a increased similarly from 2.20 ± 0.04 to 4.16 ± 0.17 mg/kg/min. In the EXE (2.53 ± 0.08 to 3.43 ± 0.09 mg/kg/min) and DAPA/EXE (2.48 ± 0.05 to 3.48 ± 0.09) groups, the increase in total R_a was significantly less compared with both PCB and DAPA (P < 0.05). The mean rate of RaO in the systemic circulation during 300 min after glucose ingestion was 2.53 ± 0.09 mg/kg/min with PCB and 2.45 ± 0.13 mg/kg/min with DAPA. RaO was significantly reduced in the EXE (2.07 ± 0.10 mg/kg/min) and DAPA/EXE (1.84 ± 0.12 mg/kg/min) groups compared with both PCB and DAPA (P < 0.01).

Following oral glucose, the baseline rate of total R_d increased in the PCB group from 2.30 ± 0.05 to a mean of 3.72 ± 0.15 mg/kg/min and in the DAPA group from 2.20 ± 0.04 to 3.95 ± 0.14 mg/kg/min (P < 0.05 vs. PCB). The increase in total R_d was similarly attenuated in both the EXE (2.53 ± 0.08 vs. 3.26 ± 0.10 mg/kg/min) and DAPA/EXE (2.48 ± 0.05 vs. 3.41 ± 0.09 mg/kg/min) groups (P < 0.05 vs. DAPA and PCB). After glucose ingestion, UGE in PCB increased from 0.07 ± 0.03 to a mean of 0.53 ± 0.13 mg/kg/min and in DAPA from 0.02 ± 0.01 to 1.31 ± 0.25 mg/kg/min (P < 0.001 vs. PCB). In EXE, UGE increased slightly (0.03 ± 0.01 to 0.11 ± 0.04), while in DAPA/EXE, UGE increased from 0.08 ± 0.04 to 1.13 ± 0.17 mg/kg/min (P < 0001 vs. EXE and P < 0.05 vs. DAPA). The rate of tissue R_d was calculated by subtracting UGE rate from total R_d. In PCB after glucose ingestion, baseline tissue R_d increased from 2.23 ± 0.13 to a mean of 3.20 ± 0.22 mg/kg/min and in EXE from 2.49 ± 0.17 to 3.15 ± 0.14 mg/kg/min. In DAPA, the increase in tissue R_d was attenuated (2.18 ± 0.09 to 2.63 ± 0.13) (P < 0.05 vs. PCB and EXE), and in DAPA/EXE, it actually decreased (2.40 ± 0.10 to 2.28 ± 0.16 mg/kg/min; P < 0.01 vs. all) (Table 2).

Total metabolic clearance rate of glucose (MCR_G) in the PCB group increased from 1.35 ± 0.07 (baseline) to 1.46 ± 0.11 mL/kg/min during the 300 min after glucose intake and from 1.51 ± 0.08 to 1.97 ± 0.09 mL/kg/min in the DAPA group (P < 0.05) (Table 2). In EXE, total MCR_G increased from 1.67 ± 0.07 to 1.83 ± 0.08 mL/kg/min, and in DAPA/EXE, it increased further from 1.40 ± 0.07 to 2.13 ± 0.12 mL/kg/min (P < 0.05). Tissue MCR_G in the PCB group did not change significantly after the oral glucose load (1.31 ± 0.07 to 1.26 ± 0.11 mL/kg/min) and decreased slightly in the DAPA group (1.50 ± 0.07 to 1.31 ± 0.06 mL/kg/min; P = NS). In EXE, tissue MCR_G increased from 1.65 ± 0.08 to 1.77 ± 0.07 mL/kg/min and in DAPA/EXE from 1.35 ± 0.06 to 1.42 ± 0.06 mL/kg/min (both P < 0.05 vs. PCB and DAPA).

In PCB, 105.1 ± 3.4 g of glucose appeared in peripheral circulation over the 300-min period following ingestion of 75 g of glucose. Of these, 66.8 ± 2.4 g were derived from the oral glucose load and 38.3 ± 1.6 g from EGP. During the same time period, 14.0 ± 3.4 g was excreted in the urine; this represented ∼13.3% of the total glucose that appeared in peripheral circulation. In DAPA, 111.1 ± 4.5 g of glucose appeared in peripheral circulation over the 300-min time period; of this, 65.4 ± 3.5 g was derived from the oral glucose load (P = NS vs. PCB) and 45.7 ± 1.9 g from EGP (P < 0.05 vs. PCB). After DAPA, 35.0 ± 4.8 g was excreted in the urine, representing ∼31.5% of total R_a in the peripheral circulation. In EXE, 88.5 ± 2.3 g of glucose appeared in peripheral circulation, of which 53.4 ± 2.6 g was derived from the oral glucose load and 35.1 ± 1.5 g from EGP. During the same period, UGE was only 2.8 ± 1.0 g, which represents ∼3.2% of the total R_a in peripheral circulation. In DAPA/EXE, 94.0 ± 2.4 g of glucose appeared in peripheral circulation, of which 49.7 ± 3.2 g was from oral glucose load and 44.3 ± 1.9 g from EGP. UGE during the same period was 30.5 ± 4.6 g, representing ∼32.4% of total R_a in peripheral circulation.

In the current study, we confirm our previous observations and those of others (2–6) that SGLT2i impair the normal time-related decline in EGP following overnight fast, as well as the suppression of EGP by EXE in patients with type 2 diabetes (Fig. 1B). No prior study has examined the effect of SGLT2i on EGP suppression following an oral glucose load. The present results provide three novel observations. First, following glucose ingestion in individuals with type 2 diabetes, SGLT2i with DAPA impairs EGP suppression in the presence of hyperinsulinemia and hyperglycemia (Fig. 2A–C). Second, the suppressive effect of EXE on EGP following glucose ingestion also is significantly impaired by SGLT2i with DAPA (Fig. 2B). In fact, the calculated difference between EGP suppressed following DAPA and PCB over the 300 min post–oral glucose (45.7 ± 1.9 minus 38.3 ± 1.6 g) amounts to ∼7.40 g, which is remarkably similar (∼9.20 g) to the calculated difference between EXE and DAPA/EXE (44.3 ± 1.9 − 35.1 ± 1.5 g). Third, the results provide the first comprehensive description of the effect of SGLT2i administration alone and in combination with GLP-1 RA on glucose kinetics following oral glucose load.

In an earlier study (6), we demonstrated that, when administered under fasting conditions, the SGLT2i-induced rise in EGP could not be explained by changes in plasma insulin or glucagon concentrations. In the current study, impaired EGP suppression during the OGTT observed with DAPA versus PCB also cannot be entirely explained by changes in insulin, since the elevation in plasma insulin was similar (except for a small difference between the 120- and 240-min time period) in DAPA and PCB studies (Fig. 2C). However, since the increment in plasma insulin in response to EXE was significantly reduced when DAPA was added, it is conceivable that it may have contributed to sustaining higher EGP. Nonetheless, plasma insulin was still increased, and, yet, EGP was not adequately suppressed. Thus, consistent with our previous studies (6), in this study, we show that following glucose ingestion, impaired EGP suppression after DAPA administration is still present, despite elevations in plasma insulin. Likewise, when DAPA was given alone and in combination with EXE, the decrease in plasma glucagon was prevented. As a result, the INS/GCN ratio increased with PCB (Δ = 0.45) and was attenuated by ∼50% (Δ = 0.24) with DAPA alone. The greater elevation in INS/GCN ratio in the EXE group (Δ = 0.99) also was reduced by ∼50% when DAPA was added (Δ = 0.46). The calculated C-peptide-to-glucagon ratio yielded similar results. Therefore, even though the INS/GCN ratio was lower in those patients who received DAPA, the increase in plasma insulin levels after oral intake of glucose in these same patients was not sufficient to suppress EGP. It should be mentioned that different elevations in plasma glucose after the oral load may have also contributed to suppress EGP more or less efficiently. These novel findings reinforce the concept that EGP stimulation in response to SGLT2i-induced glycosuria is not primarily regulated by hormonal changes, even during an oral glucose load. Moreover, our observations strongly suggest that additional factors must be involved and that a role for activation of the autonomic sympathetic nervous system triggered by the kidney cannot be excluded (9,15).

Despite the increase in EGP, the plasma glucose response following glucose ingestion with DAPA versus PCB and with DAPA/EXE versus EXE was significantly reduced (Fig. 2A). The improvement in glucose tolerance was in large part due to the marked increase in UGE (Table 2). Of the total rate of RaO (RaO + EGP) in the systemic circulation, 14.0 g (13.3%) was excreted following PCB versus 35 g (31.5%) after DAPA. Following EXE alone, only 2.8 g (3.2% of the total R_a) was excreted in urine compared with 30.5 g (32.4% of the total R_a). It should be emphasized that the calculation of the total amount of oral glucose entering peripheral blood assumes that there is no residual glucose retained in the gastrointestinal tract after 5 h. In fact, the administration of EXE, either alone or in combination with DAPA, was followed by a decrease in the RaO, which certainly contributed to the greater decline observed in plasma glucose when compared with PCB and to DAPA alone (Table 2). These data are consistent with the known inhibitory action of GLP-1 RAs on gastric emptying (16). It is noteworthy that EXE alone and DAPA/EXE similarly reduced the rise in plasma glucose concentration following glucose ingestion (Fig. 2A), even though the suppression of EGP was significantly impaired by concomitant DAPA administration (Fig. 2B). The latter was entirely accounted for by a greater decrease in RaO with DAPA/EXE versus EXE. The mechanisms responsible for the greater decline in RaO with combined DAPA/EXE are unclear, but could be accounted for by an inhibitory effect of DAPA on gut glucose absorption (17,18). The fact that RaO was modestly, but not significantly, decreased with DAPA alone also raises the question as to whether these agents inhibit gut glucose absorption and/or enhance splanchnic, including hepatic uptake of glucose, although an effect of DAPA on intestinal SGLT1 has not yet been demonstrated in vivo.

Tissue R_d (total R_d − UGE) increased following glucose ingestion with PCB in concert with the increase in plasma insulin and glucose concentrations (Table 2). When glucose was ingested with DAPA, tissue R_d increased, but the increase was less than with PCB, most likely because of the smaller increases in plasma glucose and insulin concentrations (Fig. 2A and C). Following EXE, the rise in tissue R_d was similar to that with PCB (Table 2). This was accounted for by a marked reduction in plasma glucose concentration countered by a marked rise in plasma insulin concentration (Fig. 2A and C). When DAPA was administered with EXE, tissue R_d actually declined (Table 2), resulting from a marked reduction in the rise in plasma glucose concentration combined with a modest reduction in the rise in plasma insulin concentration. When glucose was ingested with PCB, somewhat surprisingly, tissue MCR remained unchanged despite the increase in plasma insulin concentration. Following DAPA, the MCR declined compared with PCB, in conjunction with the decrease in plasma insulin concentration. Following EXE alone or in combination with DAPA, MCR increased in concert with the increase in plasma insulin concentration. These changes in glucose kinetics emphasize the complex changes in RaO, EGP, tissue R_d, UGE, and plasma insulin concentration that determine plasma glucose response to oral glucose administered after a single dose of DAPA, EXE injection, and with both DAPA and EXE.

The current study has some limitations. First, we were not able to discern the extent to which lower rates of RaO in the systemic circulation were secondary to glucose retained in stomach (e.g., slowed gastric emptying) and/or to increased glucose uptake by the splanchnic tissues, including the liver. A second limitation relates to the fact that, by study design, our observations are limited to acute events, following a single exposure to oral DAPA or injectable EXE, prior to glucose ingestion. Therefore, our data cannot provide insight into long-term effects of these agents on insulin and glucose kinetics (19,20).

In summary, we demonstrate for the first time that SGLT2i-induced stimulation of EGP is observed during an oral glucose challenge. This also was documented when EXE (which increased plasma insulin and reduced plasma glucagon) was combined with DAPA. After a single dose of DAPA, the increase in UGE was the single most important factor responsible for the attenuated plasma glucose rise following glucose ingestion. The greatest reduction in plasma glucose was observed when DAPA was administered with EXE, resulting in glycosuria, increased rate of tissue glucose disposal, and decreased RaO.

Acknowledgments. The authors thank Lorrie Albarado and Andrea Hansis-Duarte (UT Health San Antonio) for administrative assistance and for preparation of this manuscript.

Funding. This study was supported by AstraZeneca Pharmaceuticals, the Texas Diabetes Institute and University Health System, and National Institute of Diabetes and Digestive and Kidney Diseases grant RO1-DK107680-05.

Duality of Interest. C.S.-H. is a member of the speakers bureau of Novo Nordisk and member of the advisory board of Bayer. A.G. has received honorarium from Novo Nordisk and is consultant for Boehringer Ingelheim, Eli Lilly and Company, Gilead Sciences, Inc., Inventiva Pharma, and Pfizer. R.A.D. is a member of the advisory boards of AstraZeneca, Janssen, Lexicon Pharmaceuticals, Boehringer Ingelheim-Lilly Alliance, and Novo Nordisk; is a member of the speakers bureau of Novo Nordisk and AstraZeneca; and has received grant support from AstraZeneca and Janssen. E.C. has grant support from AstraZeneca. No other potential conflicts of interest relevant to this article were reported.

Author Contributions. M.A., C.A., C.S.-H., O.L., J.A., A.G., C.T., and E.C. conducted the studies and analyzed and interpreted the data. R.A.D. and E.C. designed the study, reviewed, analyzed, and interpreted the data, and wrote the manuscript. R.A.D. is the guarantor of this work and, as such, had full access to all of the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis.