title: Impaired autophagy flux is associated with neuronal cell death after traumatic brain injury.
abstract: Dysregulation of autophagy contributes to neuronal cell death in several neurodegenerative and lysosomal storage diseases. Markers of autophagy are also increased after traumatic brain injury (TBI), but its mechanisms and function are not known. Following controlled cortical impact (CCI) brain injury in GFP-Lc3 (green fluorescent protein-LC3) transgenic mice, we observed accumulation of autophagosomes in ipsilateral cortex and hippocampus between 1 and 7 d. This accumulation was not due to increased initiation of autophagy but rather to a decrease in clearance of autophagosomes, as reflected by accumulation of the autophagic substrate SQSTM1/p62 (sequestosome 1). This was confirmed by ex vivo studies, which demonstrated impaired autophagic flux in brain slices from injured as compared to control animals. Increased SQSTM1 peaked at d 1-3 but resolved by d 7, suggesting that the defect in autophagy flux is temporary. The early impairment of autophagy is at least in part caused by lysosomal dysfunction, as evidenced by lower protein levels and enzymatic activity of CTSD (cathepsin D). Furthermore, immediately after injury both autophagosomes and SQSTM1 accumulated predominantly in neurons. This was accompanied by appearance of SQSTM1 and ubiquitin-positive puncta in the affected cells, suggesting that, similar to the situation observed in neurodegenerative diseases, impaired autophagy may contribute to neuronal injury. Consistently, GFP-LC3 and SQSTM1 colocalized with markers of both caspase-dependent and caspase-independent cell death in neuronal cells proximal to the injury site. Taken together, our data indicated for the first time that autophagic clearance is impaired early after TBI due to lysosomal dysfunction, and correlates with neuronal cell death.
authors.affiliation: a Shock, Trauma and Anesthesiology Research (STAR) Center; Department of Anesthesiology ; University of Maryland School of Medicine ; Baltimore , MD USA.
authors.forename: Chinmoy
authors.initials: C
authors.lastname: Sarkar
authors.forename: Zaorui
authors.initials: Z
authors.lastname: Zhao
authors.forename: Stephanie
authors.initials: S
authors.lastname: Aungst
authors.forename: Boris
authors.initials: B
authors.lastname: Sabirzhanov
authors.forename: Alan I
authors.initials: AI
authors.lastname: Faden
authors.forename: Marta M
authors.initials: MM
authors.lastname: Lipinski
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D001343:Autophagy / Q000502:physiology*; D001921:Brain / Q000378:metabolism; D001930:Brain Injuries / Q000378:metabolism* / Q000473:pathology; D016923:Cell Death / Q000502:physiology; D004195:Disease Models, Animal; D006360:Heat-Shock Proteins / Q000378:metabolism*; D008810:Mice, Inbred C57BL; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D010588:Phagosomes / Q000378:metabolism; D000071456:Sequestosome-1 Protein
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D006360:Heat-Shock Proteins; D000071456:Sequestosome-1 Protein; C101343:Sqstm1 protein, mouse
keywords: ACTB, actin; AIFM1, apoptosis-inducing factor, mitochondrion-associated, 1; APC, adenomatous polyposis coli; ATG12, autophagy-related 12; ATG5, autophagy-related 5; ATG7, autophagy-related 7; CAPS12, caspase 12; CASP3, caspase 3; CCI, controlled cortical impact; CD68, CD68 molecule; CSPG4, chondroitin sulfate proteoglycan 4; CTSD, cathepsin D; GFP, green fluorescent protein; LAMP1, lysosomal-associated membrane protein 1; LAMP2, lysosomal-associated membrane protein 2; LC3, microtubule associated protein 1 light chain 3; RBFOX3, RNA binding protein, fox-1 homolog (C. elegans) 3; SPTAN1, spectrin, α, non-erythrocytic 1; SQSTM1, sequestosome 1; TBI, traumatic brain injury; ULK1, unc-51 like autophagy activating kinase 1; autophagy; autophagy flux; lysosome; neuronal cell death; traumatic brain injury; β; AIF1/IBA1, allograft inflammatory factor 1