Lipoxin B4 Mitigates TRPV4-Activated Muller Cell Gliosis During Ocular Hypertension

Abstract

PurposeMuller glia play dual roles in glaucoma, contributing to both retinal homeostasis and neuroinflammation; their activation by elevated intraocular pressure through the mechanosensitive channel TRPV4 promotes a reactive state that drives retinal ganglion cell (RGC) loss. Lipoxin B4 (LXB4), an endogenous lipid mediator produced by retinal astrocytes, has been shown to suppress glial reactivity and directly protect RGCs. This study investigated whether LXB4 modulates TRPV4-driven Muller glial activation and inflammation and whether Muller glia themselves contribute to this retinal lipoxin pathway. MethodsOcular hypertension (OHT) was induced in mice via a silicone oil model, and reactive Muller glia were isolated via magnetic sorting for transcriptomic analysis. In vitro, primary and immortalized Muller glia were treated with a TRPV4 agonist with or without LXB4. Glial reactivity was assessed by flow cytometry, immunostaining, qPCR, and western blotting. LC-MS/MS-based lipidomics was used to quantify lipoxin pathway metabolites, and single-cell RNA-seq was used to examine transcriptional responses to LXB4 treatment. GFAP and TRPV4 expression was evaluated via immunohistochemistry in retinal sections. ResultsRNA bulk-sequencing analysis and qPCR revealed that Muller glia express both 5- and 15-lipoxygenase. Lipidomic analysis confirmed that the lipoxin pathway is functional and that Muller glia endogenously produce LXB4, establishing this essential cell type as a source of anti-inflammatory and neuroprotective LXB4 in the retina. TRPV4 activation induced a reactive glial phenotype characterized by increased GFAP and IL6 expression, increased STAT3 phosphorylation, and increased production of lipoxins, suggesting that biomechanical stress simultaneously triggers both gliosis and protective lipid signaling. Treatment with LXB4 suppressed TRPV4-induced gliosis in vitro by downregulating IL6 and inhibiting STAT3 activation, and in vivo by reducing the expression of Stat3, Il6, and TNF- during OHT while attenuating TRPV4 upregulation in Muller glia. ConclusionMuller glia are a significant source of LXB4 in the retina. This neuroprotective Muller glia pathway is amplified during chronic TRPV4 activation to counter-regulate gliosis. The findings support targeting of the TRPV4-lipoxin pathway as a potential approach to protect against OHT-induced neurodegeneration in glaucoma.