Impaired Cargo Clearance in the Retinal Pigment Epithelium (RPE) Underlies Irreversible Blinding Diseases

Abstract

Chronic degeneration of the Retinal Pigment Epithelium (RPE) is a precursor to pathological changes in the outer retina. The RPE monolayer, which lies beneath the neuroretina, daily internalises and digests large volumes of spent photoreceptor outer segments. Impaired cargo handling and processing in the endocytic/phagosome and autophagy pathways lead to the accumulation of lipofuscin and pyridinium bis-retinoid A2E aggregates and chemically modified compounds such as malondialdehyde and 4-hydroxynonenal within RPE. These contribute to increased proteolytic and oxidative stress, resulting in irreversible damage to post-mitotic RPE cells and development of blinding conditions such as age-related macular degeneration, Stargardt disease and choroideremia. Here, we review how impaired cargo handling in the RPE results in their dysfunction, discuss new findings from our laboratory and consider how newly discovered roles for lysosomes and the autophagy pathway could provide insights into retinopathies. Studies of these dynamic, molecular events have also been spurred on by recent advances in optics and imaging technology. Mechanisms underpinning lysosomal impairment in other degenerative conditions including storage disorders, α-synuclein pathologies and Alzheimer’s disease are also discussed. Collectively, these findings help transcend conventional understanding of these intracellular compartments as simple waste disposal bags to bring about a paradigm shift in the way lysosomes are perceived.

Oxidative Stress and Dysfunctional Intracellular Traffic Linked to an Unhealthy Diet Results in Impaired Cargo Transport in the Retinal Pigment Epithelium (RPE)

“…However, 24 h after the feeding assay, most cargos were localized to LAMP1 early lysosomes (Figure F) or LAMP2 mature lysosomes (Figure G). POS in lysosomes had a predominantly perinuclear arrangement, consistent with findings that cargos intended for degradation are trafficked to lytic compartments deeper in the cell and in proximity to the nucleus . By 48 h, most POS molecules had co‐localized to LC3b labelled compartments, which also had a largely perinuclear distribution (Figure H).…”

“…Although a majority of cargos are transported in this manner, there appears to be some variability, perhaps indicating a degree of flexibility which could help RPE to cope with stressful conditions. The importance of correct cargo trafficking and timely proteolytic degradation is demonstrated by evidence from diverse neurodegenerative and storage diseases which reveal impairment of these mechanisms in early stages of pathology . Our studies reveal divergent outcomes for POS trafficking under conditions of oxidative stress, impaired lysosomal function and disrupted autophagy ( Figure ), all of which can occur simultaneously in the senescent RPE .…”

“…Although Rab7 labelling can distinguish between late phagosomes/endosomes vs lysosomes, there is evidence to suggest that some lysosomes are also positive for Rab7 . In order to assess whether this occurs in the RPE, living cells were transfected with a genetic lysosomal marker after which they were probed with an antibody specific for Rab7.…”

“…For instance, we have shown that loss of MYO1C/myosin 1c involved in the distribution of lipid rafts impairs lysosome‐autophagosome fusion . Whether by an increase in acidity or by alkalinization of lysosomes, fusion with autophagosomes is nonetheless disrupted, since this is dependent on specific intra‐lysosomal conditions . This process is likely to be highly specific as the accumulation of A2‐E, for instance, does not lead to lysosomal pH changes or inhibit activity of lysosomal enzymes in the RPE .…”

“…Such insights contribute to the current understanding of how cargos are trafficked and processed in the RPE. Although we and others have utilized well‐characterized approaches to ensure a tight pulse, the possibility of some premature POS internalization due to leakage cannot be fully excluded. However, it appears that a proportion of POS may be trafficked outside the conventional pathway via other mechanisms.…”