Study of pro-apoptotic and pro-survival pathways at late stage disease in two non-allelic forms of photoreceptor degeneration

摘要

Purpose : To examine by RNA-seq analysis the predominant pro-survival and pro-apoptotic transcriptomic profiles in two canine models of photoreceptor degeneration (rcd1/PDE6B mutation and xlpra2/RPGR mutation) at late stage disease. Methods : The retinal transcriptome of 3 rcd1 (age: 22 wks) and 3 xlpra2 (age: 41 wks) female dogs at a disease stage with about 75% photoreceptor loss was compared to that of 3 normal females (age: 24 wks) by RNA-seq analysis. Retinas were collected for RNA extraction (OS) or OCT embedding without fixation (OD). RNA-seq libraries were sequenced with a minimum read-depth of 25 million per sample. EdgeR and limma packages (Bioconductor) were used for normalization and differential gene expression analysis. P-value for differential gene expression was calculated by a likelihood ratio test. All differentially expressed genes with a P a?¤ 0.05 and a a?￥ 2 fold-change value were considered for pathway analysis. Further characterization and pathway analysis was performed using Gene Set Enrichment Analysis (GSEA) and Ingenuity Pathway Analysis (IPA, Qiagen). Fluorescent immunohistochemistry was performed on OCT embedded sections to evaluate retinal localization and expression of some of the genes identified by RNA-seq analysis. Results : Both pro-apoptotic and pro-survival pathways were active at late stages of rcd1 and xlpra2 diseases. A number of genes belonging to Tumor Necrosis Factor (TNF) superfamily pathway that have recently been shown to be activated in early stages of both diseases (Genini et. al., PLoS ONE 2013) continued to be up-regulated at late stage. In addition genes associated with the mitochondria-dependent apoptosis pathway, as well as a pro-survival and anti-apoptotic genes, and genes involved in the complement cascade, angiogenesis, autophagy and inflammation were up-regulated during the late disease stage. Conclusions : Disease progression to later stages of photoreceptor degeneration is likely to be controlled by a balance of more than one pro-death and pro-survival pathways. Further examination of these pathways will enhance our understanding of disease processes and possibly identify therapeutic targets for late stage intervention. This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.