Dapagliflozin protects against dilated cardiomyopathy progression by targeting NLRP3 inflammasome activation

Abstract

Dilated cardiomyopathy (DCM) is the major cause of heart failure and has a poor prognosis. The accumulating evidence points to an essential role of the inflammatory component in the process of DCM. Inhibitors of sodium-glucose cotransporter 2 (SGLT2) are widely used to treat heart failure patients due to their cardiac benefits. However, their role in DCM remains unclear. We used the doxorubicin (Dox)-induced DCM model for our study. The SGLT2 inhibitor dapagliflozin (Dapa) improved cardiac function in mice treated with doxorubicin and attenuated the activation of the nucleotide-binding oligomerization domain-like receptor family protein 3 (NLRP3) inflammasome pathway and the expression of inflammatory factors. In addition, dapagliflozin suppresses NLRP3 activation by decreasing p38-dependent toll-like receptor 4 (TLR4) expression. In our study, dagliflozin improves cardiac function in DCM by inhibiting the activity of the NLRP3 inflammasome.

Keywords: Dilated cardiomyopathy; Inflammation; NLRP3 inflammasome; SGLT2 inhibitors; TLR4.

Fig. 1

Dapagliflozin treatment significantly improved cardiac functions in doxorubicin-induced DCM mice. A Murine cardiac functions are analyzed by echocardiography. B The left ventricular ejection fraction (LVEF), the left ventricular fractional shortening (LVFS), the left ventricular internal dimensions (LVIDd), and the left ventricular internal dimensions (LVIDs) were measured at the end of the experiment (n = 10 per group). C Representative images of the staining of HE in heart sections from mice treated with doxorubicin or dapagliflozin (scale bars, 2 mm)

Fig. 2

Dapagliflozin inhibited the activation of the NLRP3 inflammasome in doxorubicin-induced DCM mice. A Relative NLRP3, ASC, Caspase-1, and AIM2 mRNA expression level were detected by real-time PCR in control, Dox, and Dox + Dapa mice. (n = 5 per group). B Representative immunofluorescence images of NLRP3 in control, Dox, and Dox + Dapa mice. cTNT (red), NLRP3 (green), and DAPI (blue) (scale bars, 100 μm). C–D Representative western blot of NLRP3 inflammasome pathway and densitometric analysis in control, Dox, and Dox + Dapa mice (n = 5).​ *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001

Fig. 3

Dapagliflozin inhibited the expression of the inflammatory factors interleukin (IL)-6, L-1β, and IL-18 in cardiac tissues. A Representative immunohistochemical images of IL-6, IL-1β, and IL-18 and densitometric analysis in control, Dox, and Dox + Dapa mice (scale bars, 50 μm). B Representative western blot of IL-6, pro-IL-1β, cleaved-IL-1β, IL-18, and densitometric analysis in control, Dox, and Dox + Dapa mice (n = 5). **P < 0.01, ***P < 0.001, ****P < 0.0001

Fig. 4

In vitro, dapagliflozin suppressed the Dox-induced inflammatory response. A Representative immunofluorescence images of NLRP3 inflammasome in control, Dox, and Dox + Dapa cells (scale bars, 100 μm). B Representative western blot of NLRP3 inflammasome pathway and densitometric analysis in control, Dox, and Dox + Dapa cells.​ C Representative western blots of inflammatory factors IL-6, pro-IL-1β, cleaved-IL-1β, IL-18, and densitometric analysis in control, Dox, and Dox + Dapa cells. D ELISA analysis of inflammatory factors (IL-6, IL-1β, and TNF-α) in cell supernatant of control, Dox, and Dox + Dapa cells. (n = 5 per group). ***P < 0.001, ****P < 0.0001

Fig. 5

Dapagliflozin reduced the expression of NLRP3 inflammasome through the inhibition of TLR4 by p38. A–B Representative western blots (A) and densitometric analysis (B)of TLR4, P-p38, and P-ERK in control, Dox, Dox + Dapa, and Dox + SB cells. ***P < 0.001, ****P < 0.0001