FOXO1-NMNAT3 axis dysregulation promotes doxorubicin cardiotoxicity: NAD⁺ replenishment as a redox-targeted antioxidant therapy

Doxorubicin (DOX) induces dose-dependent cardiotoxicity, primarily through oxidative stress and metabolic dysregulation. Although NAD+ deficiency has been implicated in cardiovascular pathology, its role in DOX-induced cardiotoxicity (DIC) remains poorly understood. This study investigated NAD+ metabolism dysregulation as a redox-sensitive mechanism in DIC pathogenesis. Human cardiomyocytes (AC16), mouse atrial myocytes (HL-1), and C57BL/6 mice were used to establish the DIC model. The role and mechanism of NAD+ in DIC were investigated using a range of methods. Using integrated in vitro and in vivo models, we demonstrated that DOX induces myocardial oxidative damage accompanied by NAD+ depletion. Exogenous NAD+ supplementation mitigated the DOX-induced cardiomyocyte death and redox imbalance. Mechanistically, pharmacological CD38 inhibition with 78C or genetic silencing failed to restore the NAD+ pool, whereas nicotinamide mononucleotide adenylyltransferase 3 (NMNAT3) overexpression, combined with nicotinamide mononucleotide (NMN) administration, effectively rescued NAD+ levels and attenuated oxidative stress. Computational and functional analyses identified FOXO1 as a transcriptional repressor of NMNAT3 following DOX exposure. This study establishes the dysregulation of the FOXO1-NMNAT3 axis as a key mechanism underlying NAD+ depletion in DIC. Targeting this axis through NAD+ replenishment, particularly by activating NMNAT3, offers a novel redox-based therapeutic strategy against DIC.阿霉素（Doxorubicin, DOX）可诱导剂量依赖性心脏毒性，其主要致病机制为氧化应激与代谢失调。尽管烟酰胺腺嘌呤二核苷酸（nicotinamide adenine dinucleotide, NAD+）缺乏已被证实与心血管病理过程密切相关，但其在阿霉素诱导的心脏毒性（Doxorubicin-induced cardiotoxicity, DIC）中的具体作用仍尚不明确。 本研究聚焦NAD+代谢紊乱作为DIC发病机制中的氧化还原敏感通路展开探究。实验采用人心肌细胞（AC16）、小鼠心房肌细胞（HL-1）及C57BL/6小鼠构建DIC模型，通过多维度实验方法系统解析NAD+在DIC中的作用及潜在分子机制。 通过整合体外与体内实验模型，本研究证实DOX可诱导心肌氧化损伤并伴随NAD+水平显著耗竭。外源性补充NAD+能够有效缓解DOX介导的心肌细胞死亡与氧化还原失衡。机制研究显示，采用78C进行CD38药理学抑制或基因沉默均无法恢复细胞内NAD+池水平；而烟酰胺单核苷酸腺苷酰转移酶3（nicotinamide mononucleotide adenylyltransferase 3, NMNAT3）过表达联合烟酰胺单核苷酸（nicotinamide mononucleotide, NMN）给药，则可有效挽救NAD+水平并减轻氧化应激损伤。通过生物信息学与功能实验分析，本研究鉴定出叉头框蛋白O1（forkhead box O1, FOXO1）为DOX暴露后NMNAT3的转录抑制因子。 本研究明确了FOXO1-NMNAT3轴的紊乱是DIC中NAD+耗竭的关键核心机制。通过NAD+补充策略靶向该轴，尤其是激活NMNAT3的表达与功能，可为DIC提供一种基于氧化还原调控的新型治疗方案。