In developed countries, age-related macular degeneration (AMD) is the most common cause of loss of central vision in people over the age of 65. The retinal pigment epithelium (RPE) appears to be the site of early pathological changes in AMD. Thus, age related structural changes in the RPE, such as the accumulation of lipofuscin 'age pigment', decrease in melanin content, and accumulation of deposits beneath the RPE or -within Bruch's membrane, appear to be strongiy associated with the pathogenesis of AMD. The posunitotic RPE provides essential support for the photoreceptors through daily phagocytosis of the shed photoreceptor outer segment (POS) tips. The area comprised of photoreceptors, RPE, Bruch' s membrane, and the choroid represents a unique environment. The relatively high oxygen tension, and the exposure to short-wavelength light, promotes the generation of reactive oxygen species (ROS) and free radicals. The POS has a high concentration of polyunsatorated fatty acids (PUFA's) and are thus highly susceptible to attack from ROS and free radicals. The resultant peroxides may transform into aldehydes, cross-liok to protein fragments undergoing intralysosomal degradation and form fluorophores, similar to Schiff bases, known components of lipofuscin. Over time, lipofuscin accumulation in the RPE can be substantial and interfere with cellular functions. RPE deterioration and ensning photoreceptor degeneration may contribute to the development of AMD.
The present study focuses on the formation of lipofuscin and on different ways of op- and down-regolating its formation in cultored RPE cells. By developing an experimental model we have shown that the highly metabolically active, posunitotic RPE cells, when cultured nnder hyperoxia (40% O2) with addition of POS, fulfil the prerequisites for massive lipofuscin accumulation. Using this model, the present study shows: (1) that heavily lipofuscin loaded, cultored rabbit RPE cells, have a reduced capacity of phagocytizing both Texas Red labeled latex beads, and Texas Red labeled POS compared to uuloaded control cells after both 2 and 4 weeks of culturing; (2) that melanin-rich RPE cells from pigmented rabbit and calf were better protected against lipofuscin formation than albino rabbit RPE cells and melanin-poor calf RPE cells after 4 weeks of cultoring; (3) that RPE cells from rabbit and calf, supplemented with the antioxidant substances a-tocopherol (vitamin E), lutein, zeaxanthin or lycopene accumulated less lipofuscin compared to control cells after 18 days of culturing; ( 4) that hydroxychloroquine induced less intensive vacuolization of the cytoplasm in calf RPE cells at both 24 h and 2 weeks, and less intensive periodic acid Schiff staining (as an indication of lipofuscin content) at 2 weeks, than did chloroquine.
We conclude that the mvolvement of oxidative processes seems highly plausible to be involved in the formation of RPE lesions associated with aging and AMD. Furthermore, lipofuscin-engorged RPE cells have a reduced capacity for further phagocytosis, in turn resulting in degeneration of photoreceptors and the development of AMD. Melanin possesses aoti-oxidative properties, perhaps through the sequestration of heavy metals, especially traosition metals such as iron aod copper. Apart from quenching of siuglet oxygen and protection against blue light, the carotenoids lutein zeaxanthine and lycopene seem, as a-tocopherol, to have chain breaking abilities in peroxidation reactions, reducing lipofuscin formation. The lysosomotropic weak bases chloroquine, hydroxychloroquine and ammonium chloride compromise the function of lysosomes. An estimated difference in lysosomotropism between the two drugs would correspond to the difference in toxicity reported between chloroquiue and hydroxychloroquine.
Thus, this study has revealed several mechanisms involved in the formation of lipofuscin m RPE, mechanisms that may be iroplicated in the pathogenesis of AMD.
Linköping: Linköpings universitet , 2002. , 43 p.
2002-02-08, Berzeliussalen, Universitetssjukhuset, Linköping, 13:00 (Swedish)