Abstract
Purpose :
Subretinal inflammation plays a critical role in retinal degenerative diseases. Although activated macrophages have been shown to play a key role in the progression of retinopathies, little is known about the mechanisms involved in the loss of photoreceptors leading to vision impairment. We investigated the effect of the modulation of CD36 (cluster of differentiation 36) in the inflammatory response involved in retinal degeneration.
Methods :
Retinal degeneration was induced in wild-type (C57Bl6) and CD36-/- mice with a blue-light exposure model. ERG and photoreceptor integrity were analyzed to evaluate the retinal damages. In this context, we study the protective effect of CD36- selective azapeptide ligand (labelled MPE-001). We investigated the subretinal inflammation response by immunofluorescence (iba-1, F4/80) on retinal flatmounts and qPCR analysis (iba-1, iNOS, IL-6, IL-10, IL-12) on laser capture microdissection. Fluorescence Resonance Energy Transfer (FRET) was performed on peritoneal macrophages to study the CD36-TLR2 interactions, respectively using antibodies coupled to energy donor Cy3 and acceptor Cy5. Protein expressions were analyzed by western blot (P65, JNK1/2, P38, IRAK4, IKKαβ, NLRP3, caspase-1, IL-1b) or ELISA (IL-1b). TUNEL assays were used to analyze photoreceptor apoptosis.
Results :
We have observed that CD36-deficient mice featured less subretinal macrophage accumulation with attenuated photoreceptor degeneration compared to wild-type mice (WT). Treatment with MPE-001 as modulator of the inflammatory environment of the retina reduced subretinal macrophage/ activated microglia accumulation with preservation of photoreceptor layers and function assessed by ERG in WT, in a CD36-dependent manner. MPE-001 modulated the transcriptome of subretinal macrophage/activated microglia by reducing pro-inflammatory markers. In isolated macrophages, MPE-001 induced dissociation of the CD36-TLR2/6 heterodimer complex (using FRET) altering the TLR2 signaling pathway, thus decreasing NF-kB activation and inflammasome activity. The azapeptide also incurred cytoprotection against photoreceptor apoptosis elicited by activated macrophages.
Conclusions :
These findings suggest that the azapeptide as ligand of co-receptor CD36 decreases the inflammatory response by modulating CD36-TLR2/6 complex signaling pathway in macrophages, and suggests its potential application in the treatment of retinal degenerative diseases.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.