Late Sodium Current in Atrial Cardiomyocytes Contributes to the Induced and Spontaneous Atrial Fibrillation in Rabbit Hearts

Figure 1. Cardio-metabolic vulnerability associated with PI3Kα inhibition in HR+/HER2− PIK3CA-mutant breast cancer. In patients receiving alpelisib combined with endocrine therapy, inhibition of insulin–PI3K signaling induces insulin resistance and hyperglycemia, which may amplify oxidative stress, inflammatory activation, and myocardial susceptibility to injury. This clinical scenario creates a cardio-oncologic vulnerability state that may predispose to cardiovascular adverse events and underscores the need for integrated cardiometabolic protection strategies.

Figure 2. Effects of alpelisib, fulvestrant, and dapagliflozin on cardiomyocyte viability, mitochondrial membrane potential, and apoptotic signaling under hyperglycemic and normoglycemic conditions. Human iPSC-derived cardiomyocytes were treated for 24 h with alpelisib (100 nM), fulvestrant (100 nM), or their combination, in the absence or presence of dapagliflozin (1 µM), under hyperglycemic (25 mM glucose) or normoglycemic (5.5 mM glucose) conditions. (A) Cell viability was assessed by MTS assay and expressed as percentage relative to that of untreated control cells. (B) Mitochondrial membrane potential (ΔΨm) was measured using the TMRM method and expressed as relative changes in fluorescence. (C) Caspase-3 activity, a marker of execution-phase apoptosis, was quantified by luminescent assay and normalized to total protein content. Drug treatments were associated with reduced viability, mitochondrial depolarization, and increased apoptotic signaling, with more pronounced effects under hyperglycemic conditions. Dapagliflozin attenuated these alterations across conditions. Data are presented as mean ± SEM with individual data points representing independent biological replicates (n = 6).…

Figure 3. Alpelisib and fulvestrant induce cardiomyocyte injury, attenuated by dapagliflozin co-treatment. Human iPSC-derived cardiomyocytes were treated for 24 h with alpelisib (100 nM), fulvestrant (100 nM), or their combination, in the absence or presence of dapagliflozin (1 µM) under hyperglycemic conditions. (A) Heart-type fatty acid binding protein (H-FABP) and (B) cardiac troponin I levels were quantified in culture supernatants as markers of cardiomyocyte injury and normalized to total protein content where applicable. Drug treatments were associated with increased biomarker release, while dapagliflozin reduced these responses. Data are presented as mean ± SEM with individual data points representing independent biological replicates (n = 6). Statistical analysis was performed using two-way ANOVA followed by Tukey’s multiple-comparison test. * p < 0.05, *** p < 0.001, **** p < 0.0001; ns, not significant.

Figure 4. Dapagliflozin and NHE1 inhibition differentially modulate oxidative stress and electrophysiological alterations in human cardiomyocytes exposed to PI3Kα inhibition under hyperglycemic conditions. Human iPSC-derived cardiomyocytes were treated for 24 h with alpelisib (100 nM), fulvestrant (100 nM), or their combination, in the absence or presence of dapagliflozin (1 µM) or a selective NHE1 inhibitor under hyperglycemic conditions. (A) Lipid peroxidation was assessed by measuring malondialdehyde (MDA) levels in cardiomyocyte lysates. (B) 4-hydroxynonenal (4-HNE) levels were quantified as an additional marker of oxidative lipid damage. (C) Late sodium current (InaL), expressed as a percentage of peak sodium current, was measured to evaluate electrophysiological alterations. Drug treatments were associated with increased oxidative stress and InaL. Both dapagliflozin and NHE1 inhibition reduced these alterations, with dapagliflozin showing a more pronounced effect. Data are presented as mean ± SEM with individual data points representing independent biological replicates (n = 6). Statistical analysis was performed using two-way ANOVA followed by Tukey’s multiple-comparison test. * p <…

Figure 5. PI3Kα inhibition and estrogen receptor blockade activate inflammasome-related signaling in cardiomyocytes, attenuated by dapagliflozin. Human iPSC-derived cardiomyocytes were treated for 24 h with alpelisib (100 nM), fulvestrant (100 nM), or their combination, in the absence or presence of dapagliflozin (1 µM) under hyperglycemic conditions. Intracellular levels of (A) NLRP3 and (B) MyD88 were quantified in cardiomyocyte lysates and normalized to total protein content. Drug treatments were associated with increased expression of inflammasome-related signaling components, while dapagliflozin reduced these responses. Data are presented as mean ± SEM with individual data points representing independent biological replicates (n = 6). Statistical analysis was performed using two-way ANOVA followed by Tukey’s multiple-comparison test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Figure 6. Dapagliflozin suppresses PI3Kα inhibitor-associated inflammatory cytokine signaling in human cardiomyocytes under hyperglycemic stress. Human iPSC-derived cardiomyocytes were treated for 24 h with alpelisib (100 nM), fulvestrant (100 nM), or their combination, in the absence or presence of dapagliflozin (1 µM) under hyperglycemic conditions. Intracellular levels of (A) IL-1β and (B) IL-18, and secreted levels of (C) IL-6, (D) TNF-α, and (E) CCL2 were quantified and normalized to total protein content. Alpelisib and combined treatment were associated with increased inflammatory signaling, while dapagliflozin reduced cytokine levels across conditions. Data are presented as mean ± SEM with individual data points representing independent biological replicates (n = 6). Statistical analysis was performed using two-way ANOVA followed by Tukey’s multiple-comparison test. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ns, not significant.

Figure 7. Experimental design and conceptual overview of hyperglycemia-associated cardiomyocyte stress induced by PI3Kα inhibition and estrogen receptor blockade, and its modulation by dapagliflozin. Human iPSC-derived cardiomyocytes (hiPSC-CMs) were cultured under hyperglycemic conditions (25 mM glucose) and exposed for 24 h to the PI3Kα-selective inhibitor alpelisib (100 nM) and/or the estrogen receptor antagonist fulvestrant (100 nM), in the absence or presence of the SGLT2 inhibitor dapagliflozin (1 µM). Combined PI3Kα inhibition and estrogen receptor blockade under hyperglycemia were associated with a cardiomyocyte injury phenotype characterized by increased late sodium current (INaL), cardiac troponin release, mitochondrial membrane depolarization (ΔΨm loss), oxidative stress and lipid peroxidation, and inflammasome-associated inflammatory signaling. Dapagliflozin attenuated these alterations and was associated with preservation of mitochondrial integrity and redox homeostasis. The schematic provides a conceptual representation of interconnected pathways rather than a strictly linear mechanistic cascade and summarizes the experimental model and the principal biological endpoints…