Neohesperidin Alleviates Ferroptosis in Doxorubicin-Induced Cardiotoxicity via Mettl3-m6A-Nrf2 Signaling.

Doxorubicin (DOX) has antitumor efficacy, but its clinical application is restricted by multiple side effects, especially cardiotoxicity. Neohesperidin, a flavonoid in citrus fruits, has various health benefits. The aim of this study was to explore the cardioprotective effects and mechanisms of neohesperidin against DOX-induced cardiotoxicity (DIC). The study used DOX-treated mice to evaluate cardiac function and cardiomyocyte injury. In addition, primary cardiomyocytes treated with DOX were used to measure cell viability and death. FIN56, a ferroptosis inhibitor, was used to test the relationship between neohesperidin's beneficial effect and ferroptosis. Mechanistic investigations were carried out to explore the role of N6-methyladenosine (m6A) levels, nuclear factor erythroid-2-related factor 2 (Nrf2) expression, and Methyltransferase-like 3 (Mettl3) in the cardioprotective effect of neohesperidin. Overexpression and knockdown of Mettl3 were also performed to confirm its role in the cardioprotective effect. Neohesperidin attenuated cardiac function and cardiomyocyte injury in DOX-treated mice. It alleviated DOX-induced oxidative stress by reducing reactive oxygen species and malondialdehyde levels and increasing superoxide dismutase and glutathione levels, restoring ATP production. In DOX-treated primary cardiomyocytes, neohesperidin increased cell viability and decreased cell death. FIN56 abolished the beneficial effect of neohesperidin. Mechanistically, neohesperidin decreased m6A levels, leading to increased Nrf2 expression. Nrf2 activation was related to the inhibition of Mettl3, an m6A "writer," which protected cardiomyocytes against DOX-induced ferroptosis. Overexpressing Mettl3 reversed the cardioprotective effects of neohesperidin and diminished its benefits on ferroptosis. These findings suggest that neohesperidin exerts a cardioprotective effect against DIC by inhibiting ferroptosis. This effect is achieved by regulating the Mettl3-m6A-Nrf2 signaling pathway, indicating the therapeutic potential of neohesperidin.