Na LiJi‐Lin ChenYi‐Jian SunJia‐Fan SunFatin Athirah PauziSong‐Lin Zhu0000-0002-9831-4395Amy Yi Hsan, SaikSaikAmy Yi Hsan0000-0002-1882-7757Alan Ong Han Kiat2026-10-192026-10-192026-03-2610.1002/ibra.70018https://dspace-cris.utar.edu.my/handle/123456789/11790Berberine (BBR) exerts an effective protection for diabetic retinopathy (DR), but the underlying key molecular mechanism remains unknown; this study investigated the protective mechanism of BBR on DR by alleviating cell pyroptosis. A rat DR model was established and treated with BBR, and histological analyses, including hematoxylin and eosin staining, Nissl staining, and immunofluorescence, were executed to evaluate tissue changes. Core target genes were identified using the GeneCards database, Venn diagram analysis, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, protein-protein interaction networks, and molecular docking. Validation of key genes was performed via reverse transcription-quantitative polymerase chain reaction (RT-qPCR), Western blot, and RNA interference. BBR improved retinal morphology, reduced edema, and restored the arrangement of retinal ganglion cells in DR rats. BBR significantly reduced the levels of pyroptosis markers such as IL-1? and IL-18, which were elevated in DR. Network pharmacology identified 10 hub genes, with six genes (JUN, STAT3, AKT1, TP53, IL-1B, EGFR) further analyzed. BBR reversed DR-induced upregulation of JUN, STAT3, and AKT1 at both the mRNA and protein levels, as confirmed by RT-qPCR and Western blot. Silencing these genes enhanced cell viability and amplified BBR's protective effects. Altogether, BBR alleviates retinal inflammation and pyroptosis in diabetic retinal ganglion cells by targeting JUN, STAT3, and AKT1, providing insights into its therapeutic potential for DR. © 2026 The Author(s). Ibrain published by Affiliated Hospital of Zunyi Medical University (AHZMU) and Wiley-VCH GmbH.en-USberberinecell pyroptosisdiabetic retinopathynetwork pharmacologycryopyrininterleukin 18interleukin 1betainterleukin 1beta converting enzymeinterleukin 8messenger RNAprotein p53STAT3 proteinstreptozocinadultanimal experimentanimal tissueArticlecell viabilitygene ontologyhistologyimmunofluorescenceimmunohistochemistryKEGGmalemolecular dockingNissl stainingnonhumanpharmacologyprotein protein interactionpyroptosisratreal time polymerase chain reactionreal time reverse transcription polymerase chain reactionretina ganglion cellRNA interferencesystems pharmacologyWestern blottingjournal-article