![]() ![]() While hfRPE cultures were remarkably resistant to UAM accumulation, cultures with heavy UAM burden had mild defects in tight-junction integrity, ketogenesis, RPE differentiation, and phagocytic capacity. Newer granules resembled those seen in RPE from patients with AMD older granules more closely resembled lipofuscin from healthy, aged patients. We discovered that UAM granules are dynamic, compacting and shifting their spectrum over time, suggesting that newly formed and older granules may have different biologic effects. 17 Here, we develop a model for accumulation of lipofuscin-like material, which we term undigested autofluorescent material (UAM), in hfRPE cultures through repeated OS feedings. 1, 16 However, this has not been done in primary (passage 1) human fetal RPE (hfRPE) cultures, among the most extensively validated systems for in vitro studies of human RPE. An alternate approach is to induce lipofuscinogenesis by OS feedings of cultures. 1, 15 Furthermore, it may not be possible to recapitulate an accurate model of lipofuscinogenesis by feeding RPE with just one type of bisretinoid. 12 – 14 However, controversy exists over which, if any, specific bisretinoids are toxic. Many groups have fed a single bis-retinoid, A2E, to the RPE cultures. To understand the relationship between lipofuscin and RPE toxicity in a more controlled setting, a number of groups have attempted to model lipofuscin accumulation in cell culture. Prior to RPE cell death, the marked accumulation of lipofuscin in normal, healthy, aged RPE creates a strong “background” above which any disease-specific signal is difficult to measure. Thus, any degree of cell death decreases the strength of any putative relationship between lipofuscin and toxicity. As RPE cells die, lipofuscin is lost and fundus autofluorescence decreases. 3 The significance of this difference is unknown.ĭistinguishing whether lipofuscin is toxic in AMD patients in the clinic is difficult. While levels of lipofuscin may not correlate with drusen formation or retinal toxicity, the lipofuscin granules in patients with AMD have been reported to be larger and more irregular than granules simply associated with age. 11 Additionally, there is minimal evidence supporting a link between lipofuscin and the pathologic hallmark of AMD, extracellular accumulation of lipid-rich deposits above (termed reticular pseudodrusen) and below (termed drusen) the RPE while lipofuscin accumulates in the RPE of all persons as they age, only a subset develop pathologic drusen or reticular pseudodrusen. 6 Further, the concentration of A2E or its oxidized components is no higher in RPE underlying the macula compared to the retinal periphery, despite AMD being a macular disease. ![]() ![]() 9, 10 On the other hand, lipofuscin-related fundus autofluorescence from patients with early- or intermediate-stage AMD is no higher than age-matched controls. 8 Numerous components of lipofuscin, in particular the bisretinoid N-retinylidene-N-retinylethanolamine (A2E), have been shown in cell culture and animal models to dysregulate complement activity, induce photo-oxidative damage, and disrupt lysosomal integrity. 6, 7 In support of lipofuscin toxicity, animal models lacking the ABCA4 gene, mimicking the defect seen in Stargardt disease, accumulate lipofuscin at an accelerated rate and demonstrate retinal degeneration. 5 Whether this accumulated waste is toxic to the RPE, and in what diseases, is heavily debated. 2 – 4 It contains numerous components, including a complex mixture of bisretinoids derived from covalent linkage of monoretinoids used in the visual cycle, oxidized lipid products, and cross-linked protein. 1 Lipofuscin accumulates over a lifetime, occupying major portions of the RPE cell in elderly individuals. Incomplete degradation of OS material contributes, in part, to the development of autofluorescent intracellular inclusions termed lipofuscin. As the RPE is a mostly postmitotic tissue, the phagocytic burden for each cell over a lifetime is immense. The daily ingestion of shed OS tips from photoreceptors by the RPE is critical for photoreceptor health and function. ![]()
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