Phloretin inhibits ferroptosis by restoring the antioxidant capacity of bovine adipose and muscle cells via the AMPK-PPAR signaling pathway.

Ferroptosis has been increasingly implicated in adipose and muscle dysfunction, systemic metabolic disturbances, and several diseases in livestock, which necessitates effective and side-effect-free inhibition strategies. Phloretin, a dihydrochalcone with excellent antioxidant and anti-inflammatory properties, may have the potential to restrain cell ferroptosis. Herein, phloretin was verified to significantly inhibit (1S,3R)-RSL3-induced ferroptosis by reducing intracellular MDA, Fe2⁺, and ROS levels and restoring cell total antioxidant capacity in bovine and mouse preadipocytes or myoblasts. It also alleviated oxidative stress (OS), a vital inducer of ferroptosis, by restoring antioxidant enzyme activity in the above cells and obese mice. In vivo, phloretin gavage significantly reversed the trend where high-fat diet (HFD)-induced OS promoted the expression of ferroptosis-promoting genes and proteins (e.g., ACSL4 and PTGS2) while inhibiting the expression of ferroptosis-negative regulators (e.g., Fth1 and Gpx4). Unlike most flavonoids that exert anti-inflammatory or antioxidant activities by altering the gut microbiota composition, metagenomic sequencing analysis of cecal contents from phloretin-gavaged and HFD mice revealed that phloretin exerts its antioxidative and ferroptosis-inhibitory effects independent of modulating gut microbiota diversity. Further transcriptomic analyses of mouse adipose tissues revealed that phloretin alleviated ferroptosis in adipocytes by modulating the transcription of genes enriched in AMPK and PPAR signaling pathways, such as Camkk2. Hence, based on multi-omics analysis combined with in vivo and in vitro verification, phloretin effectively alleviated the OS to further inhibit ferroptosis of adipose or muscle cells through the AMPK-PPAR pathway, which can provide new research ideas for ameliorating adipose or myocyte dysfunction induced by ferroptosis in animals.