Fasting confers protection against diabetes-induced retinopathy via Akkermansia muciniphila-derived extracellular vesicles

Intermittent fasting is a promising intervention for metabolic disorders. However, whether different fasting regimens exert distinct effects on diabetic retinopathy (DR) remains unknown. Here, we compared alternate-day fasting (ADF) and intermittent energy restriction (IER) in a STZ-induced insulin-deficient mouse model and investigated the role of the gut‑retina axis. A streptozotocin-induced DR mouse model was subjected to ADF or IER. Integrative analyses included gut microbiota sequencing, microbiota depletion, extracellular vesicle (EV) isolation, untargeted metabolomics, and functional studies of CYP2F2 using knockout mice and cellular models. In this model, ADF showed greater efficacy than IER in glycemic control, preservation of retinal architecture, and attenuation of inflammation and oxidative stress, an effect that was accompanied by selective enrichment of Akkermansia muciniphila (Akk), and our depletion and supplementation experiments indicated a necessary role for Akk. Akk supplementation recapitulated these protective effects in an EV‑dependent manner, establishing a causal role of Akk‑derived EVs (Akk‑EVs) natural biological nanoparticles with a mean diameter of 80‑120 nm. Metabolomic profiling identified L-carnitine (LC) as a key metabolite enriched in Akk-EVs, and LC administration improved visual function and vascular integrity. RNA sequencing, molecular docking, thermal shift assays, and enzyme inhibition assays collectively identified CYP2F2 as a direct functional target of L‑carnitine. LC supplementation or CYP2F2 knockdown suppressed VEGF and IL-1β, and enhanced glycolytic and mitochondrial ATP production, whereas CYP2F2 knockout abolished LC-mediated benefits. LC levels in Akk-EVs were reduced by GW4869, and LC transport inhibition abrogated Akk-EV protection, consistent with LC being a necessary mediator of Akk-EV function under the conditions tested. These findings, derived from an STZ‑induced insulin‑deficient model, support a mechanistic cascade in which ADF promotes enrichment of Akk, which delivers L‑carnitine to the retina via extracellular vesicles, leading to inhibition of CYP2F2 and subsequent retinal protection. While the linearity of this gut–retina axis requires further confirmatory studies, our functional experiments establish necessary roles for Akk, LC, and CYP2F2 in ADF‑mediated retinoprotection. This study highlights context-dependent benefits of different fasting regimens and identifies the Akk/L-carnitine/CYP2F2 module as a potential target for further investigation in DR.