Dapagliflozin Attenuates Na⁺/H⁺ Exchanger-1 in Cardiofibroblasts via AMPK Activation

Purpose: We assessed whether the SGLT-2 inhibitor dapagliflozin (Dapa) attenuates the upregulation of the cardiac Na⁺/H⁺ exchanger (NHE-1) in vitro in mouse cardiofibroblasts stimulated with lipopolysaccharides (LPS) and whether this effect is dependent on adenosine monophosphate kinase (AMPK) activation. Methods: Mouse cardiofibroblasts were exposed for 16 h to Dapa (0.4 μM), AMPK activator (A769662 (10 μM)), AMPK inhibitor (compound C (CC) (10 μM)), an SGLT-1 and SGLT-2 inhibitor (phlorizin (PZ) (100 μM)), Dapa+CC, or Dapa+PZ, and then stimulated with LPS (10 ng/ml) for 3 h. NHE-1 mRNA levels were assessed by rt-PCR and total AMPK, phosphorylated-AMPK (P-AMPK), NHE-1, and heat shock protein-70 (Hsp70) protein levels in the whole cell lysate by immunoblotting. In addition, NHE-1 protein levels attached to Hsp70 were assessed by immunoprecipitation. Results: Exposure to LPS significantly reduced P-AMPK levels in the cardiofibroblasts. A769662 and Dapa equally increased P-AMPK. The effect was blocked by CC. Phlorizin had no effect on P-AMPK. LPS exposure significantly increased NHE-1 mRNA levels. Both Dapa and A769662 equally attenuated this increase. The effect of Dapa was blocked with CC. Interestingly, none of the compounds significantly affected NHE-1 and Hsp70 protein levels in the whole cell lysate. However, LPS significantly increased the concentration of NHE-1 attached to Hsp70. Both Dapa and A69662 attenuated this association and CC blocked the effect of Dapa. Again, phlorizin had no effect and did not alter the effect of Dapa. Conclusions: Dapa increases P-AMPK in cardiofibroblasts exposed to LPS. Dapa attenuated the increase in NHE-1 mRNA and the association between NHE-1 and Hsp70. This effect was dependent on AMPK.