Abstract
Alzheimer's disease (AD) is a progressive neurodegeneration with dysfunctions in both the ubiquitin-proteasome system (UPS) and autophagy. Astroglia participation in AD is an attractive topic of research, but molecular patterns are partially defined and available in vitro models have technical limitations. Immortalized astrocytes from the hippocampus of 3xTg-AD and wild-type mice (3Tg-iAstro and WT-iAstro, respectively) have been obtained as an attempt to overcome primary cell line limitations and this study aims at characterizing their proteolytic systems, focusing on UPS and autophagy. Both 26S and 20S proteasomal activities were downregulated in 3Tg-iAstro, in which a shift in catalytic subunits from constitutive 20S proteasome to immunoproteasome occurred, with consequences on immune functions. In fact, immunoproteasome is the specific complex in charge of clearing damaged proteins under inflammatory conditions. Parallelly, augmented expression and activity of the lysosomal cathepsin B, enhanced levels of lysosomal-associated membrane protein 1, beclin1, and LC3-II, together with an increased uptake of monodansylcadaverine in autophagic vacuoles, suggested autophagy activation in 3Tg-iAstro. The two proteolytic pathways were linked by p62 that accumulated in 3Tg-iAstro due to both increased synthesis and decreased degradation in the UPS defective astrocytes. Treatment with 4-phenylbutyric acid, a neuroprotective small chemical chaperone, partially restored proteasome and autophagy-mediated proteolysis in 3Tg-iAstro. Our data shed light on the impaired proteostasis in 3Tg-iAstro with proteasome inhibition and autophagic compensatory activation, providing additional validation of this AD in vitro model, and propose a new mechanism of action of 4-phenylbutyric acid in neurodegenerative disorders.