Class effects of SGLT2 inhibitors in mouse cardiomyocytes and hearts: inhibition of Na+/H+ exchanger, lowering of cytosolic Na+ and vasodilation

Abstract

Aims/hypothesis: Sodium-glucose cotransporter 2 (SGLT2) inhibitors (SGLT2i) constitute a novel class of glucose-lowering (type 2) kidney-targeted agents. We recently reported that the SGLT2i empagliflozin (EMPA) reduced cardiac cytosolic Na⁺ ([Na⁺]_c) and cytosolic Ca²⁺ ([Ca²⁺]_c) concentrations through inhibition of Na⁺/H⁺ exchanger (NHE). Here, we examine (1) whether the SGLT2i dapagliflozin (DAPA) and canagliflozin (CANA) also inhibit NHE and reduce [Na⁺]_c; (2) a structural model for the interaction of SGLT2i to NHE; (3) to what extent SGLT2i affect the haemodynamic and metabolic performance of isolated hearts of healthy mice.

Methods: Cardiac NHE activity and [Na⁺]_c in mouse cardiomyocytes were measured in the presence of clinically relevant concentrations of EMPA (1 μmol/l), DAPA (1 μmol/l), CANA (3 μmol/l) or vehicle. NHE docking simulation studies were applied to explore potential binding sites for SGTL2i. Constant-flow Langendorff-perfused mouse hearts were subjected to SGLT2i for 30 min, and cardiovascular function, O₂ consumption and energetics (phosphocreatine (PCr)/ATP) were determined.

Results: EMPA, DAPA and CANA inhibited NHE activity (measured through low pH recovery after NH₄⁺ pulse: EMPA 6.69 ± 0.09, DAPA 6.77 ± 0.12 and CANA 6.80 ± 0.18 vs vehicle 7.09 ± 0.09; p < 0.001 for all three comparisons) and reduced [Na⁺]_c (in mmol/l: EMPA 10.0 ± 0.5, DAPA 10.7 ± 0.7 and CANA 11.0 ± 0.9 vs vehicle 12.7 ± 0.7; p < 0.001). Docking studies provided high binding affinity of all three SGLT2i with the extracellular Na⁺-binding site of NHE. EMPA and CANA, but not DAPA, induced coronary vasodilation of the intact heart. PCr/ATP remained unaffected.

Conclusions/interpretation: EMPA, DAPA and CANA directly inhibit cardiac NHE flux and reduce [Na⁺]_c, possibly by binding with the Na⁺-binding site of NHE-1. Furthermore, EMPA and CANA affect the healthy heart by inducing vasodilation. The [Na⁺]_c-lowering class effect of SGLT2i is a potential approach to combat elevated [Na⁺]_c that is known to occur in heart failure and diabetes.

Keywords: Cardiac; Diabetes; Heart failure; Na+/H+ exchanger; SGLT2i; Sodium; Vasodilation.

Fig. 1

EMPA, CANA and DAPA inhibit NHE activity, reduce [Na⁺]_c and bind to the Na⁺-binding site of NHE-1. (a) SGLT2i inhibit NHE activity in cardiomyocytes. Recovery of the pH is a measure of NHE activity and was only seen in vehicle (black line). All three drugs blocked pH recovery, and thus NHE activation, significantly, compared with vehicle (***p < 0.001). (b) [Na⁺]_c was reduced in all three SGLT2i (***p < 0.001). Data are presented as mean ± SD and are derived from eight cells from four mouse hearts for each condition. (c) In silico analysis of SGLT2i binding to a homology model of NHE. All three SGLT2i efficiently bind to the Na⁺-binding pocket of NHE. Calculated binding affinities of SGLT2i are much higher than the negative control, glucose. EMPA is shown in blue, DAPA is shown in red and CANA is shown in green

Fig. 2

EMPA and CANA induced coronary vasodilation in the healthy intact heart. Hearts were perfused for 30 min with vehicle (0.02% DMSO; n = 15, black bars), 1 μmol/l EMPA (n = 9, white bars), 1 μmol/l DAPA (n = 10, light grey bars) or 3 μmol/l CANA (n = 10, dark grey bars). (a) EMPA and CANA significantly reduced perfusion pressure after 30 min (*p < 0.05). (b) SGLT2i administration did not change rate pressure product (RPP) and (c) PCr/ATP (vehicle n = 13, EMPA n = 8, DAPA n = 10, CANA n = 10). Values at t = 30 min are normalised to values at t = 0 min from the same condition. (d) No changes were observed for O₂ consumption for all three SGLT2i (vehicle n = 12, EMPA n = 8, DAPA n = 8, CANA n = 10). The dashed line in each graph represents the mean of vehicle condition. Data for perfusion pressure (a) and RPP (b) are presented as mean ± SD; data of PCr/ATP (c) and O₂ consumption (d) are presented as median, interquartile range