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Long-term Rescue of Photoreceptors in a Rodent Model of Retinitis Pigmentosa Associated with MERTK Mutation

Updated: Feb 5

H. Lorach, S. Kang, R. Dalal, M. B. Bhuckory, Y. Quan, D. Palanker | Scientific Reports | 27 July 2018 | https://doi.org/10.1038/s41598-018-29631-z


Abstract

MERTK mutation reduces the ability of retinal pigment epithelial (RPE) cells to phagocytize the photoreceptor outer segments, which leads to accumulation of debris separating photoreceptors from RPE cells, resulting in their degeneration and loss of vision. In a rat model of Retinitis Pigmentosa due to MERTK mutation, we demonstrate that surgical removal of debris performed when about half of photoreceptors are lost (P38), allows the remaining photoreceptor cells to renew their outer segments and survive for at least 6 months – 3 times longer than in untreated eyes. In another set of experiments, patterned laser photocoagulation was performed before the debris formation (P19-25) to destroy a fraction of photoreceptors and thereby reduce the phagocytic load of shed outer segment fragments. This treatment also delayed the degeneration of the remaining photoreceptors. Both approaches were assessed functionally and morphologically, using electroretinography, optical coherence tomography, and histology. The long-term preservation of photoreceptors we observed indicates that MERTK-related form of inherited retinal degeneration, which has currently no cure, could be amenable to laser therapy or subretinal surgery, to extend the visual function, potentially for life.


Introduction

Retinitis pigmentosa (RP) is a group of inherited retinal diseases that can lead to profound loss of vision and eventual blindness due to progressive degeneration of photoreceptor cells. These disorders can be caused by a wide variety of genetic defects. To date, nearly 4300 mutations in 79 genes have been reported to cause RP1. Many of the associated genes encode proteins that are involved in phototransduction, photoreceptor structure, or photoreceptor gene transcription. MER-proto-oncogene, tyrosine kinase (MERTK) gene encodes for a transmembrane protein involved in recognition and phagocytosis of the photoreceptor outer segments, essential for recycling of the phototransduction machinery. Mutations in the MERTK gene cause reduced phagocytic function, which leads to accumulation of photoreceptor outer segment debris in subretinal space. This debris subsequently impedes efficient oxygen and nutrient transport to photoreceptor cells. This mutation was actually identified thanks to a spontaneous rodent model of retinal degeneration: the Royal College of Surgeons (RCS) rat. This first animal model of inherited retinal degeneration, was described by Bourne and coworkers in 19382. Since 1970s, it has been known that the RCS rat has dysfunctional RPE cells, which are unable to phagocytize shed outer segment fragments, leading to their accumulation as a debris between photoreceptors and RPE cells3,4. Recently, mutations in the MERTK gene were identified as the cause of the functional defect in RPE cells of RCS rat5. In humans, mutations of the MERTK gene lead to a loss of night vision in early childhood, gradual constriction of the visual field, and eventual loss of visual acuity before adulthood. Imaging studies using optical coherence tomography (OCT) in these patients revealed the loss of photoreceptors and formation of a hyper-reflective layer, analogous to the debris layer in RCS rats6,7.


In the RCS rats, appearance of the outer segment debris begins around postnatal day (P) 19. These debris accumulate with age8, forming a thick insulating layer between photoreceptors and RPE cells by P35, accompanied by gradual degeneration of photoreceptors, which is complete by P180. RCS animal model is widely used today in research of retinal degeneration, including strategies for RPE cell transplantation9,10,11,12,13,14, gene therapy15,16,17,18 or retinal prosthetics19,20,21,22,23. Many of these studies do not introduce a proper sham procedure group, while a few that do, often report a protective effect in the sham surgery group, but tend to undermine and qualify it as a short-term effect24. One study25, focused on the photoreceptors rescue effect by retinal detachment, demonstrated anatomical rescue comparable to results with RPE transplantation. Unfortunately, this study followed the animals only for 2 months and was not designed to show functional preservation of vision.


Since retinal outer segments are shed daily, but debris accumulation does not start for a few years in patients and for a few weeks in RCS rats, while retina continues to function long after that, we hypothesized that the mutant RPE cells retain some phagocytic activity either by MERTK-independent uptake or microglia-related phagocytosis26, and thereby can sustain a fraction of the outer-segment recycling load27. We also assumed that photoreceptors degeneration is accelerated by accumulation of a thick debris layer, which prevents oxygen and nutrients supply from the choroid. Based on these hypotheses, we designed two strategies to balance the supply and demand of the outer segment recycling and thereby extend the survival of photoreceptors in RCS rats.


Both strategies utilize clinically readily available techniques. First, debris layer can be surgically washed out from subretinal space when a substantial fraction of functional photoreceptors is still present. Alternatively, pattern laser photocoagulation can be applied to selectively destroy a fraction of photoreceptors prior to debris formation. In both approaches, the remaining photoreceptors could be better sustained by the partially functioning RPE cells and survive much longer. In this study, we assess efficacy of these therapies for protection of photoreceptors in RCS rats as a potential therapy for patients with RP due to MERTK mutation.



 

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