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Ingrid Porpino Meschede, Tanya Tolmachova, Miguel C Seabra, Clare Futter; Mechanisms of photoreceptor degeneration in choroideremia. Invest. Ophthalmol. Vis. Sci. 2014;55(13):4367.
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© ARVO (1962-2015); The Authors (2016-present)
Choroideremia (CHM) is an X-linked retinitis pigmentosa-related disease characterized by gradual degeneration of the photoreceptors (PR), retinal pigment epithelium and choroid leading ultimately to vision loss. CHM is caused by loss-of-function mutation of Rab Escort Protein-1 (Rep1), a protein involved in lipid modification (prenylation) and function of Rab GTPases, a protein family that regulates multiple intracellular trafficking pathways. The purpose of this study was to use our mouse CHM models to define how PR die and to identify defects in membrane traffic pathways that could lead to PR death.
We have focused on our CHM mouse model, Chmnull/WT heterozygous female carriers, where nearly 50% of cells lack Rep1 and PR gradually degenerate. 10 µm retinal sections and whole mounts of 4-7 month old female carrier eyes, were subjected to TUNEL assays and immunofluorescent staining for rhodopsin and autophagy markers. In addition, the PR ultrastructure was analysed by transmission electron microscopy (TEM) and rhodopsin distribution was analysed by cryo-immunoEM.
Retinal sections showed a higher fluorescent staining intensity for the autophagy markers LC3 and Beclin-1 in the PR layer of female carriers, compared with controls, suggestive of upregulated autophagy. Retinal sections showed very few TUNEL positive PR nuclei but whole mounts revealed increased numbers of TUNEL positive nuclei in female carriers, compared with controls. TEM showed swollen mitochondria and fragmented outer segments (OS) in female carriers, indicative of dying cells. TEM also demonstrated patchy disorganisation of OS and a general shortening of OS throughout the PR layer, suggesting defects in transport to the outer segments. However there was no clear build-up of transport vesicles within the inner segment and connecting cilium morphology appeared normal. Analysis of rhodopsin distribution by immunofluorescence showed no clear rhodopsin accumulation in the inner segment in female carriers.
There is evidence of both increased apoptosis and autophagy in PR of Chmnull/WT heterozygous female carriers, accompanied by general shortening of OS and patchy OS disorganisation. Further ultrastructural analysis of apoptosis, autophagy, OS morphology and rhodopsin distribution will elucidate the relationship between defects in transport to the outer segment and PR cell death pathways in CHM.
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