Abstract
Purpose :
Photoreceptor degeneration in retinitis pigmentosa (RP) is thought to be associated with chronic low grade intraocular inflammation which manifests clinically as vitreous cells, cystoid macular oedema, epiretinal membrane and early-onset cataract. With the emergence of gene therapy for RP, it is unclear if background inflammation might compound any immune response to viral vectors. We explored the immunological state of the retinas of ageing Rpgr-deficient mice as a model for slowly progressive RP.
Methods :
Rpgr-/y mice were assessed at 3, 6, 9 and 12 months by SLO, OCT and ERG. Retinas from 12-month-old Rpgr-/y mice (n=2) and age-matched wildtype (C57BL/6J) controls (n=2) were harvested for IHC to assess microglia and Müller glia morphology. Multicolor flow cytometry (n=8 Rpgr-/y, n=4 WTeyes) was performed on dissociated retinas at 12 months to quantify immune cell populations using a panel detecting CD45, CD11b, CD3, CD4, NK1.1, CD19, TMEM119 and CD11c.
Results :
Rpgr-/y mice develop significant peripheral retinal thinning from 6 months, progressing to outer nuclear layer thinning centrally and peripherally at 9 months. Evenly distributed hyperautofluorescent dots appear on SLO from 3 months, increasing in number with age, but do not correlate with any features on OCT. 12-month aged Rpgr-/y mice demonstrate reduced dark-adapted A-wave and light-adapted B-wave responses compared with age-matched controls. Flow cytometric analysis revealed no significant difference in overall retinal immune cell (CD45+) numbers between Rpgr-/y and WT mice at 12 months. The majority of CD45+ cells in each group were microglia (at 49.05% and 29.03% in Rpgr-/y and WT), reflecting their status as the resident immune cell of the retina. No significant infiltrating leukocyte populations were detected, constituting only 0.42% and 0.46% of live cells in Rpgr-/y and WT retinas, respectively (p=0.72). IHC showed increasing migration of microglia into outer retina from 3 months. No abnormal GFAP staining of Müller glia was detected up to 12 months (Fig.1).
Conclusions :
Lack of significant leukocyte infiltration of Rpgr-/y retina with age is indicative of an intact blood-retinal barrier despite slow progressive retinal degeneration. Further work will assess the role of migrating microglia, compare immunological changes with more rapidly progressive RP, and the impact of gene therapy on the degenerating retina.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.