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Identifier	000421788
Title	Involvement of macroautophagy in nigrostriatal axonal degeneration induced by inhibition of chaperone-mediated autophagy
Alternative Title	Η συμμετοχή της μακροαυτοφαγίας στον εκφυλισμό της μελαινοραβδωτής οδού που προκαλείται απο την αναστολή της αυτοφαγίας διαμεσολαβούμενης από Σαπερόνες
Author	Φούκα, Μαρία
Thesis advisor	Στεφανής, Λεωνίδας
Reviewer	Καραγωγέως, Δόμνα Νικολετοπούλου, Βασιλική
Abstract	Chaperone-mediated autophagy (CMA) is one of the main autophagic proteolytic pathways involving the selective targeting of cytosolic proteins to the lysosomes. It is widely believed that dysfunction of the CMA pathway is important in the pathogenesis of Parkinson’s disease (PD). In order to model this, we have recently generated a novel rat model, in which downregulation of CMA within the nigra, via AAV-shRNA-mediated targeting of endogenous Lamp2a, the rate-limiting step in the CMA pathway, leads to loss of striatal dopamine, nigral neuron degeneration, and accumulation of alpha-synuclein, 8 weeks post injection, thus mimicking aspects of PD. In this system, we have observed a marked accumulation of autophagic vacuoles and biochemical indices suggestive of enhanced productive macroautophagy in the nigral cell bodies of dopaminergic neurons. In view of these features, aim of the current study was to assess the contribution of macroautophagy to the dopaminergic axonal degeneration that precedes nigral cell death, evoked by inhibition of the Chaperone-mediated autophagy (CMA) pathway. In order to inhibit CMA, we have stereotaxically injected adeno-associated viruses expressing shRNAs targeting LAMP2A receptor or scrambled shRNAs in the rat substantia nigra (SN). At 2 and 3 weeks post-injection, we examined indices of macroautophagy induction and the formation of autophagic vacuoles (AVs) in the nigrostriatal axis by Confocal and Electron Microscopy. The integrity of the nigrostriatal projections and the astro- and micro-gliosis in both striatum and SN at these time-points were also assessed by Confocal Microcscopy. LAMP2A down-regulation was accompanied by abnormal accumulation of AVs at synaptic nerve terminals, prior to dopaminergic degeneration at 3 weeks post-injection. At this early time point, the levels of Bassoon, a negative regulator of autophagy and a marker for the active synaptic zone, were decreased, whereas levels of ULK-1 were increased. Increased astro- and micro-gliosis was observed in both SN and striatum. Our data suggest that uncontrolled induction of macroautophagy may, at least in part, be responsible for the nigrostriatal terminal degeneration that occurs early in this model, well before cell soma degeneration. Further, our results provide the first in vivo evidence that ULK-1 is a CMA substrate and may act as a link between CMA and macroautophagy. Therefore, down-regulation of macroautophagy may represent a promising target to reverse the damage and rescue the deteriorating dopaminergic neurons.
Language	English
Subject	Neurodegeneration
	Νευροεκφύλιση
Issue date	2019-03-27
Collection	School/Department--School of Medicine--Department of Medicine--Post-graduate theses
	Type of Work--Post-graduate theses
Permanent Link	https://elocus.lib.uoc.gr//dlib/e/5/3/metadata-dlib-1554197487-423306-17653.tkl
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