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Sriganesh Ramachandra Rao, Lara Ann Skelton, Steven J Pittler, Steven J. Fliesler; Microglial activation in a mouse model of RP59: Contributor or bystander?. Invest. Ophthalmol. Vis. Sci. 2020;61(7):709.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the role of microglial activation in the rapid degeneration of photoreceptors (PRs) observed in a mouse model of RP59, involving rod-specific ablation of the dehydrodolichyl diphosphate synthase (Dhdds) gene.
Rod-specific Dhdds knockout (KO) mice were generated (Rao et al., ARVO, 2017), with Dhddsflx/flx iCre+ (KO) and Dhddsflx/flx iCre- (control) mice selected by PCR tail snip analysis for the loxP-modified Dhdds allele and the Rho-iCre transgene. Retinas harvested from PN 4-wk old KO and control mice were subjected to gene expression analysis using NanoString® neurodegeneration/neuroinflammation panels. Immunohistochemistry (IHC) was performed on eyes from PN 5-wk KO and control mice, probing with anti-microglial markers (Iba-1, CD68, F4/80). Retinal whole mounts were probed by IHC to quantify microglia infiltration into the ONL; particle analysis was performed using ImageJ® software. KO mice were maintained on a diet containing PLX3397 (CSF1R inhibitor, 300 mg/kg chow), starting at PN 3 wk, to deplete retinal microglia. ERG analysis was performed on control and KO mice ± PLX3397 at PN 4/5 wk (N=3-4/group). Statistical analysis: Statistical significance: Student’s t-test, P<0.05.
Retinal gene expression analysis from PN 4-wk old KO and control mice (N=3 ea.) indicated significant changes (fold-change cutoff: 2) in the levels of molecules involved in neuroinflammation and microglial activation (Cxcl-10, TMEM119, C1q, Lif) in KO mice. IHC analysis of KO retinas revealed microglial infiltration into the ONL and subretinal space, with microglia forming phagocytic cups around TUNEL-negative (live) photoreceptors. Quantitative IHC analysis of microglial ONL infiltration demonstrated a significant increase in Iba-1/CD68-positive area in the ONL of KO vs. control retinas (N=3/group, P<0.05). Scotopic ERG analysis at PN 4-5 wk demonstrated rapid rod photoreceptor degeneration in KO mice, regardless of PLX3397 treatment, comparable to that observed in KO mice fed normal chow, at all tested flash intensities.
Gene expression signatures of retinal microglial activation were upregulated in a rod-specific Dhdds-KO mouse model of RP59, at a time point (PN 4 wk) prior to observable retinal degeneration. Microglial infiltration into the ONL was observed at PN 5 wk, with phagocytosis of live PRs. However, dietary PLX3397 treatment did not rescue the retinal degeneration.
This is a 2020 ARVO Annual Meeting abstract.
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