Low-dose juglone attenuates pressure overload-induced cardiac hypertrophy through Lys207-dependent FoxO1 activation and mitochondrial protection.

Pressure overload-induced cardiac hypertrophy is characterized by mitochondrial dysfunction and redox imbalance. Here, we investigated the efficacy, safety window, and redox-regulatory mechanism of juglone, a naturally occurring 5-hydroxy-1,4-naphthoquinone. In mice subjected to transverse aortic constriction (TAC), high-dose juglone induced cardiac, hepatic, and renal toxicity, whereas 1 mg/kg/day was well tolerated and attenuated cardiac hypertrophy. Juglone also suppressed Ang II-induced cardiomyocyte hypertrophy in vitro and preserved mitochondrial ultrastructure and mitochondrial membrane potential, respiration, ATP production, and redox homeostasis. Single-nucleus RNA sequencing (snRNA-seq) revealed that juglone reduced stress-associated cardiomyocyte states and enriched calcium-handling cardiomyocytes. Integrated transcriptomic analyses, Biotin-Juglone pull-down assay, surface plasmon resonance, cellular thermal shift assays, molecular docking, and mutational analyses identified FoxO1 as a functional target of juglone. Juglone engaged the FoxO1 DNA-binding domain in a Lys207-dependent manner, and was associated with FoxO1 deacetylation, nuclear accumulation, and transcriptional activation. Activated FoxO1 induced Sesn3 transcription and restored AMPK phosphorylation at Thr172. Sesn3 silencing markedly attenuated juglone-induced AMPK activation and impaired its protective effects against cardiomyocyte hypertrophy and mitochondrial dysfunction. FoxO1 silencing, FoxO1-K207Q mutation, blockade of FoxO1 deacetylation, or inhibition of FoxO1 transcriptional activity impaired these benefits. Importantly, myocardium from patients with aortic valve stenosis-associated hypertrophy exhibited increased FoxO1 acetylation, reduced cardiomyocyte nuclear FoxO1 localization, decreased Sesn3 expression and AMPK phosphorylation, and increased 4-HNE. Collectively, low-dose juglone attenuates pathological hypertrophy and preserves mitochondrial homeostasis through the FoxO1-Sesn3-AMPK axis, while highlighting the importance of dose-dependent redox effects in determining its therapeutic efficacy and systemic toxicity.