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NCL Method GTA-12

By Stephan Stern, Barry Neun

Autophagic Dysfunction in LLC-PK1 Cells

Listed in Datasets | publication by group NCL Protocols

Version 2.0 - published on 09 Jul 2020 doi:10.17917/YPTH-N396 - cite this

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Lysosomal dysfunction is recognized as a potential toxic mechanism for xenobiotics that can result in various pathological states (1).  There is concern that nanoparticles, in particular, may cause lysosomal pathologies, since they are likely to accumulate within lysosomes (2).  Lysosomal dysfunction could potentially result from nanoparticle biopersistance, or inhibition of lysosomal enzymes, such as inhibition of phospholipase resulting in phospholipidosis, or inhibition of lysosomal protein degradation resulting in lysosomal overload (1).  Nanoparticle exposure has also been shown to cause autophagic dysfunction (3), resulting in increased lysosomal mediated degradation of cellular organelles.  Common methods used to characterize autophagic dysfunction include direct morphological assessment via light and electron microscopy, in both cell culture and tissue samples, as well as use of lysosomal dyes (4, 5).  The method detailed in this protocol utilizes the latter lysosomal dye staining method, as it is suited to high throughput screening.

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