Combined SGLT2 inhibition and GLP-1 receptor agonism synergistically reduce rejection-associated oxidative stress in cardiac allografts of heart transplant recipients

Background Cardiac allograft rejection is characterized by immune activation, oxidative injury, endothelial dysfunction, and progressive adverse remodeling. While SGLT2 inhibitors (SGLT2i) and GLP-1 receptor agonists (GLP-1Ra) confer cardiometabolic protection in clinical trials, a potential protective role in cardiac allografts remains undefined. Purpose Characterize cardiac SGLT2 (SLC5A2) and GLP-1R (GLP1R) pattern of expression across different allograft rejection grades and evaluate ex vivo the effects of combined SGLT2i and GLP-1Ra. Methods The cohort comprised 100 heart transplant recipients (median age 58 years [53–66], 71% male) with substantial cardiometabolic burden, including hypertension (49%), dyslipidemia (56%), diabetes (29%), and median BMI 27.0 kg/m² [23.5–30.8]. Endomyocardial biopsies were classified as Grade 0, Quilty lesions, cellular rejection (CMR), or antibody-mediated rejection (pAMR) based on conventional histological assessment. Gene expression (RT-qPCR), reactive oxygen species (ROS) formation (dihydroethidium staining), and ex vivo pharmacological modulation with SGLT2i empagliflozin and GLP-1Ra semaglutide at different concentrations as well as a potential synergy were assessed. Results Compared with Grade 0 biopsies, samples classified as CMR and pAMR exhibited significantly higher expression of pro-inflammatory cytokines (IL1B, IL6, TNFA), pro-inflammatory macrophage marker (CD68, CD86), RAAS-related genes (AT1R, NR3C2), NADPH oxidase subunits (CYBA, CYBB), endothelial adhesion molecules (ICAM1, VCAM1), and remodeling and fibrosis mediators (MMP2, MMP9, TGFB1), SLC5A2 (SGLT2) and GLP1R (GLP-1R) alongside reduced NOS3 (eNOS) expression. Interestingly, SLC5A2 and GLP1R expression correlated positively and robustly with TNFA (r² = 0.9619 and 0.9330, respectively; both P < 0.0001) and CD86 (r² = 0.9243 and 0.9575, respectively; both P < 0.0001). Similarly, ROS levels were approximately 4-fold higher in CMR and 6-fold higher in pAMR compared with Grade 0 biopsies and showed strong positive correlations with TNFA, CD86, SLC5A2, and GLP1R (all P < 0.0001). On the other hand, in CMR and pAMR biopsies, ROS levels were significantly reduced by 100 nM of either empagliflozin (-49.3 %) or semaglutide (-46.4%), whereas each compound alone at 10 nM was without effect. Notably, combining both empagliflozin and semaglutide at 10 nM each produced synergistic suppression of oxidative stress beyond single-agent treatment reaching 68.5% reduction. Conclusion Cardiac allograft rejection is characterized by coordinated inflammatory, pro-oxidant, pro-fibrotic and pro-remodeling activation tightly associated with SGLT2 and GLP-1R expression. Combined SGLT2i and GLP-1Ra exert synergistic antioxidant effects, highlighting a potential adjunctive strategy to attenuate inflammatory injury in transplanted hearts.