Abstract
Purpose :
Retinitis pigmentosa (RP) refers to a group of inherited disorders that lead to the dysfunction and death of photoreceptors, which causes progressive vision loss, and there is in principle no treatment available. The cGMP-PKG system has been suggested as a disease driver in several mouse RP models, although the downstream signaling of this enzyme system has not been clarified. Here we investigate the cGMP-PKG-system with the aim to reveal its potential target(s), that may have a direct detrimental effect during retinal degeneration.
Methods :
With the aid of organotypic retinal explant cultures from a mouse-based disease model, i.e. the rd1 mouse, we added cGMP analogues to inhibit PKG in retinal explants from rd1. Mass spectrometry followed by peptide enrichment was then done to study the cGMP-PKG dependent signaling. Via a bioinformatics analysis, we identified downstream targets regulated by this system, and selected cyclin dependent kinase 1 (CDK1) for further validation. The expression of CDK1 within photoreceptors was compared between the rd1 model and its healthy, normal wild-type counterparts. CDK1 expression was also observed after various lengths of pharmacological inhibition of PKG during culturing of rd1 explants, to further assess the association between cGMP-PKG and CDK1. Co-staining with two cell death markers, namely acetylated lysine and TUNEL, was performed to study the possible involvement of CDK1 during photoreceptor degeneration.
Results :
We observed that CDK1 was expressed in rd1 photoreceptors while no expression was noticed in the wt peers. With PKG manipulation of rd1 retinal explants, we could not find any alternation of CDK1 expression after 2 hours nor after 2 days of PKG inhibition, respectively, while CDK1 was lower expressed after 4 days treatment, compared to non-treated counterparts. Also, we found a high overlap of CDK1-positive and TUNEL-positive cells, as well as a partial overlap of CDK1-positive and acetylated lysine-negatively cells, according to the data from the co-staining of in vivo retinas.
Conclusions :
Our results, particularly the overlap of CDK1 and TUNEL positive cells, suggest that CDK1 could be a downstream effector of the cGMP-PKG system and that it may be directly connected to the cell death process. Further studies of the CDK1 enzyme during retinal degeneration may therefore put it forward as a potential target for neuroprotection.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.