Techniques for studying mechanisms of autophagy in RPE cells

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Abstract

Autophagy is basic catabolic mechanism which ”self eats” cellular components that are unnecessary or dysfunctional to the cell. Autophagy comprises three intracellular pathways in eukaryotic cells, which are macroautophagy, microautophagy and chaperone-mediated autophagy. These forms of autophagy are mechanistically different from each other, but finally they all lead to lysosomal degradation of intracellular material. Autophagy is triggered as an adaptive response during AMD-associated stress conditions such as hypoxia, oxidative stress, the unfolded protein response or inflammation. Macroautophagy process begins with the formation of isolation membranes called phagophores at the phagophore assembly site. The phagophores then become elongated and surround a portion of the cytosol to form mature double membrane autophagosomes that can engulf portions of cytoplasm containing oligomeric protein complexes and organelles. The autophagosomes fuse with the lysosomes and their content is then degraded by lysosomal enzymes. Failure of autophagy in aged cells may accelerates degenerative processes. Recent methods to study autophagy are discussed with details in the course.

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