Abstract 4352318: Interleukin-4 Exacerbates Doxorubicin-induced Cardiotoxicity by Promoting Cardiac Fibrosis and Apoptosis

Background: Doxorubicin (DOX) is a potent chemotherapeutic agent; however, its clinical use is limited by dose-dependent cardiotoxicity, leading to doxorubicin-induced cardiomyopathy (DIC). Although inflammation has been implicated in the progression of DIC, the underlying molecular mechanisms remain unclear. Hypothesis: We hypothesized that the cytokines upregulated in the heart and cardiomyocytes following DOX treatment contribute to the progression of DIC. We aim to identify the cytokine highly upregulated by DOX and elucidate its role in DOX-induced cardiotoxicity. Methods: A DIC model was induced in male BALB/c and IL-4 knockout (KO) mice by three DOX intravenous injections at 6 mg/kg each on days 0, 2, and 4. DIC hearts were physiologically and biochemically analyzed. Serum IgE levels were quantified using ELISA, and eosinophil infiltration was histologically assessed. Neonatal rat ventricular cardiomyocytes, cardiac fibroblasts, and EoL-1 cells (a human eosinophilic cell line) were used for in vitro experiments. Results: Among several cytokines (IL-1β, IL-4, IL-5, IL-6, IL-13, IFN-γ, and TNF-α), IL-4 was the most highly upregulated cytokine following DOX treatment (approximately 50-fold in cardiomyocytes and 6-fold in DIC mouse hearts). NOTCH1 signaling was activated in cardiomyocytes and hearts treated with DOX, and gene upregulation of Il4 was significantly suppressed by a NOTCH1 inhibitor. DIC mice showed reduced left ventricular ejection fraction (LVEF), increased apoptosis, myocardial fibrosis, and allergic responses, such as elevated serum IgE and eosinophilic infiltration in the myocardium. In contrast, the LVEF reduction observed in DIC was ameliorated (51±2% [wild type] vs. 60±2% [IL-4 KO], mean±SEM; p < 0.01), and myocardial apoptosis, fibrosis, IgE elevation, and eosinophil infiltration were significantly attenuated in IL-4 KO mice. Mechanistically, IL-4 directly promoted cardiac fibroblast proliferation through c-MYC activation. Furthermore, co-culture with EoL-1 cells or treatment with granules derived from EoL-1 cells promoted DOX-induced apoptosis in cultured cardiomyocytes. Conclusion: IL-4, upregulated in response to DOX via NOTCH1 signaling, exacerbates its cardiotoxicity by promoting myocyte apoptosis and interstitial fibrosis through eosinophilic infiltration and c-MYC activation, respectively. IL-4 may serve as a therapeutic target for attenuating DOX-induced cardiotoxicity and mitigating the limitation of anthracycline-based chemotherapy.