Abstract
Purpose :
We recently proposed that lipoxins are synthesized in the inner retina, are downregulated following acute and chronic injury, and that restoring their function is neuroprotective. In this study, we characterize the cell-specific regulation and functional expression of lipoxin pathways in response to ocular hypertension (OHT) induced injury to the retina.
Methods :
OHT was induced in the C57BL/6J mice by injection of silicone oil (SO) in the anterior chamber of the eye. In the contralateral eye, sterile PBS was injected and served as the control. Intraocular pressure (IOP) was measured weekly in both eyes and mice were maintained for 10 weeks (n=48). At weeks 4, 8, and 10, mice were euthanized, and the retina from both eyes was harvested for qRT-PCR and HPLC/LC/MS/MS-based lipidomic analysis. For cell-specific gene expression of Alox5, Alox15, and lipoxin receptors (Fpr2 and Fpr3), RNAscope in-situ hybridization was performed. GFAP and RBPMS expression were assessed by immunofluorescence of cryosections.
Results :
Lipidomic analysis of 4-week retinas showed changes in the activity of the 15-LOX pathway (15HETE and 17HDHA) in the SO injected group when compared with the PBS injected group and age-matched controls. qRT-PCR analysis revealed a significant up-regulation of Alox15, Fpr1, Fpr2, and Fpr3 genes (>2.5-fold induction) at 4, 8, and 10 weeks and a significant down-regulation of Alox5 gene (p<0.05) at 8 weeks after SO injection when compared with the PBS injected eyes. Interestingly, this upregulation of Alox15 (~2-fold, p<0.05), Fpr1, Fpr2, and Fpr3 genes were also observed in 8-week IOP-induced Alox5 KO mice. RNAscope data showed that Alox5, Alox15, Fpr2, and Fpr3 were localized in GCL along with INL and ONL of the retina and expressed in the optic nerve. At 10 weeks, loss of RGCs was observed as indicated by the expression of RBPMS, and induced GFAP expression was seen in the SO induced retinal sections (including muller glial activation) when compared with the control.
Conclusions :
We demonstrate profound dysregulation of the lipoxin pathway in response to induced OHT, further confirming our earlier findings of a potential and novel mechanism for RGC death. Moreover, the lipoxin pathway is expressed in the optic nerve, which provides evidence that this neuroprotective pathway role extends beyond the retina.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.