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.