Impacts of ciliary neurotrophic factor on the retinal transcriptome in a mouse model of photoreceptor degeneration

Abstract

ciliary neurotrophic factor (cntf) has been tested in clinical trials for human retinal degeneration due to its potent neuroprotective effects in various animal models. To decipher CNTF-triggered molecular events in the degenerating retina, we performed high-throughput RnA sequencing analyses using the Rds/Prph2 (P216L) transgenic mouse as a preclinical model for retinitis pigmentosa. In the absence of cntf treatment, transcriptome alterations were detected at the onset of rod degeneration compared with wild type mice, including reduction of key photoreceptor transcription factors Crx, Nrl, and rod phototransduction genes. Short-term CNTF treatments caused further declines of photoreceptor transcription factors accompanied by marked decreases of both rod-and cone-specific gene expression. in addition, cntf triggered acute elevation of transcripts in the innate immune system and growth factor signaling. These immune responses were sustained after long-term CNTF exposures that also affected neuronal transmission and metabolism. Comparisons of transcriptomes also uncovered common pathways shared with other retinal degeneration models. Cross referencing bulk RNA-seq with single-cell RnA-seq data revealed the cntf responsive cell types, including Müller glia, rod and cone photoreceptors, and bipolar cells. Together, these results demonstrate the influence of exogenous cntf on the retinal transcriptome landscape and illuminate likely cntf impacts in degenerating human retinas. Retinal degeneration (RD) is known as an irreversible, progressive neurologic disorder caused by genetic mutations and/or environmental or pathological damage to the retina. One therapeutic strategy to attenuate vision loss in disease conditions is treating the retina with neuroprotective agents that promote neuronal viability. Ciliary neurotrophic factor (CNTF) has been shown to be a potent neuroprotective agent for various types of retinal degenerations, including those caused by gene mutations, light-induced damage, and physical injury 1. Since CNTF enhances the survival of both photoreceptors and retinal ganglion cells, the two major cell classes damaged in retinal degenerative diseases, a CNTF-based therapy has been developed and tested in several clinical trials. To date, outcomes of the trials for retinitis pigmentosa, geographic atrophy, and achromatopsia have not demonstrated sufficient therapeutic benefits for human patients 2-5. However, CNTF therapy for macular telangiectasia type 2 has shown efficacy 6,7 , and a clinical trial for the second largest blinding disease glaucoma 8 is ongoing (https://clinicaltrials.gov/ct2/show/NCT01408472). To shed light on the molecular and cellular mechanisms of CNTF-mediated neuroprotection, we have delivered the same secreted recombinant CNTF used in human trials to a mouse model expressing the Rds/peripherin2 (Prph2) transgene with the P216L mutation 9. Pheripherin2 is a structural protein of the photoreceptor outer segment disc, and over 80 PRPH2 pathological mutations have been linked to RD, with...

Ciliary neurotrophic factor-mediated neuroprotection involves enhanced glycolysis and anabolism in degenerating mouse retinas

“…To evaluate whether CNTF signaling influenced metabolism at the transcription level, we performed high throughput RNA-sequencing and analyzed retinal transcriptome of the wild type and rds mutants treated with LV-IG or LV-CNTF from P25 to P35 [44]. Compared to the wild type, LV-CNTF treatment led to significant elevation of the majority of the glycolysis pathway transcripts, whereas LV-IG injection induced moderate increases of the glycolytic pathway gene expression (Figure 6A).…”

“…A previous study has shown that elevating wild type or a constitutively active STAT3 in mutant mouse rod photoreceptors can delay degeneration, and that the pY705 phosphorylation of STAT3 mediates this protective effect [83]. Since CNTF signaling asserts a strong influence on the retinal transcriptome [44], the precise functions of pY705 and pS727 STAT3 in CNTF-mediated metabolic modulation require further investigations. Consistent with our results that exogenous CNTF suppresses mitochondrial respiratory chain activity and promotes cell viability, accumulating evidence indicates that partial uncoupling of mitochondrial respiration can reduce oxidative stress and support neuronal survival under stress conditions [84–88].…”

“…Previous studies have also shown that the dosage and duration of CNTF treatment critically affect the rescued neurons, as a single dose injection of CNTF transiently affects the length of photoreceptor outer segments while prolonged exposure to high levels of CNTF can result in decreased visual function despite robust rescue of photoreceptors [7, 41–43]. Molecular analyses have revealed that CNTF-induced STAT3 phosphorylation followed by its dimerization and nuclear entry significantly influences the retinal transcriptome, resulting in rapid elevation of transcripts involved in innate immunity and growth factor signaling, as well as reduced expression of genes involved in phototransduction and maintenance of photoreceptor identities [44].…”

Risk Factors for Retinal Ganglion Cell Distress in Glaucoma and Neuroprotective Potential Intervention

“…In addition to enhancing the survival of both photoreceptors and RGCs, CNTF is capable of stimulating axonal regeneration [ 69 ]. The specific binding of CNTF to the receptor complex, which includes the CNTF receptor-α (CNTFRα), gp130 and Leukemia Inhibitory Factor receptor (LIFR), activates Jak-STAT, MEK-ERK and PI3K/Akt signaling pathways [ 70 , 71 ].…”

Plagl1 is part of the mammalian retinal injury response and a critical regulator of Müller glial cell quiescence

“…Included was Vim, a marker of reactive gliosis, as well as Sox9, a glialspecific retinal gene that is upregulated in injury 8 . Finally, in Rds/Prph2 mutant mice, treatment with CNTF induced GFAP expression and reactive gliosis, and transcriptomic analyses identified 4 Müller glia-enriched genes 59 , including Vsx2, a retinal progenitor cell (RPC) gene upregulated in injury, which was amongst the in P7 Plagl1 +/-pat retinal DEGs (Supplementary Fig. 5e ).…”