ISG15+ Müller glial subtype contributes to early blood–retinal barrier breakdown in diabetic retinopathy

Diabetic retinopathy features early glial stress and neurovascular dysfunction, yet Müller glial programs linking inflammatory cues to oxidative injury remain unclear. We identify a spatially clustered interferon-stimulated gene 15 (ISG15) + Müller glial subtype expanded in streptozotocin-induced diabetic mice. High glucose alone did not increase basal free ISG15 or detectable ISGylation (ISG15-dependent protein modification) in primary Müller glia, whereas conditioned medium from high glucose–exposed microglia induced both, recapitulated by IFN-β and attenuated by type I interferon receptor 1 inhibition. In vivo, Ifnb1 RNAscope signals were associated with microglia, retinal Ifnb1 expression increased in diabetic retinas, and microglial depletion reduced Ifnb1 expression and limited the expansion of ISG15 + Müller glia. Müller glia–specific Isg15 deletion preserved blood–retinal barrier integrity, reduced acellular capillaries and Evans blue leakage, and mitigated oxidative stress, lipid peroxidation, and glutathione redox imbalance. Loss of ISG15 or disruption of ISGylation blunted interferon-driven oxidative stress in Müller glia and reduced Müller glia–conditioned medium–dependent endothelial migration and tube formation. Mechanistically, IFN-β promoted peroxiredoxin 6 (PRDX6) ISGylation and proteasome-dependent degradation without altering Prdx6 transcripts. Thus, microglial IFN-β promotes an ISG15 + Müller glial subtype that compromises antioxidant defense and promotes early barrier injury in diabetic retinopathy.