title: Ferritin-stimulated lipid peroxidation, lysosomal leak, and macroautophagy promote lysosomal "metastability" in primary hepatocytes determining in vitro cell survival.
abstract: Several pathologies are associated with elevated levels of serum ferritin, for which growth inhibitory properties have been reported; however, the underlying mechanisms are still poorly defined. Previously we have described cytotoxic properties of isoferritins released from primary hepatocytes in vitro, which induce apoptosis in an iron and oxidative stress-dependent mode. Here we show that this ferritin species stimulates endosome clustering and giant endosome formation in primary hepatocytes accompanied by enhanced lysosomal membrane permeability (LMP). In parallel, protein modification by lipid peroxidation-derived 4-hydroxynonenal (HNE) is strongly promoted by ferritin, the HNE-modified proteins (HNE-P) showing remarkable aggregation. Emphasizing the prooxidant context, GSH is rapidly depleted and the GSH/GSSG ratio is substantially declining in ferritin-treated cells. Furthermore, ferritin triggers a transient upregulation of macroautophagy which is abolished by iron chelation and apparently supports HNE-P clearance. Macroautophagy inhibition by 3-methyladenine strongly amplifies ferritin cytotoxicity in a time- and concentration-dependent mode, suggesting an important role of macroautophagy on cellular responses to ferritin endocytosis. Moreover, pointing at an involvement of lysosomal proteolysis, ferritin cytotoxicity and lysosome fragility are aggravated by the protease inhibitor leupeptin. In contrast, EGF which suppresses ferritin-induced cell death attenuates ferritin-mediated LMP. In conclusion, we propose that HNE-P accumulation, lysosome dysfunction, and macroautophagy stimulated by ferritin endocytosis provoke lysosomal "metastability" in primary hepatocytes which permits cell survival as long as in- and extrinsic determinants (e.g., antioxidant availability, damage repair, EGF signaling) keep the degree of lysosomal destabilization below cell death-inducing thresholds.
authors.affiliation: University of Salzburg, Department of Cell Biology, Hellbrunnerstrasse 34, A-5020 Salzburg, Austria.
authors.forename: Margit A
authors.initials: MA
authors.lastname: Krenn
authors.affiliation: University of Salzburg, Department of Cell Biology, Hellbrunnerstrasse 34, A-5020 Salzburg, Austria.
authors.forename: Melanie
authors.initials: M
authors.lastname: Schürz
authors.affiliation: University of Salzburg, Department of Cell Biology, Hellbrunnerstrasse 34, A-5020 Salzburg, Austria.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Teufl
authors.affiliation: Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Uchida
authors.affiliation: University of Salzburg, Department of Cell Biology, Hellbrunnerstrasse 34, A-5020 Salzburg, Austria.
authors.forename: Peter M
authors.initials: PM
authors.lastname: Eckl
authors.affiliation: University of Salzburg, Department of Cell Biology, Hellbrunnerstrasse 34, A-5020 Salzburg, Austria. Electronic address: nikolaus.bresgen@sbg.ac.at.
authors.forename: Nikolaus
authors.initials: N
authors.lastname: Bresgen
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000447:Aldehydes / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001343:Autophagy / Q000187:drug effects*; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000494:pharmacology*; D004705:Endocytosis; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005260:Female; D005293:Ferritins / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism; D019803:Glutathione Disulfide / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D007425:Intracellular Membranes / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D007502:Iron Chelating Agents / Q000494:pharmacology; D007976:Leupeptins / Q000494:pharmacology; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D008247:Lysosomes / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D057054:Molecular Imaging; D010539:Permeability / Q000187:drug effects; D061251:Primary Cell Culture; D011480:Protease Inhibitors / Q000494:pharmacology; D066329:Protein Aggregates; D051381:Rats; D011916:Rats, Inbred F344
chemical_list: D000447:Aldehydes; D017077:Culture Media, Conditioned; D007502:Iron Chelating Agents; D007976:Leupeptins; D011480:Protease Inhibitors; D066329:Protein Aggregates; C025946:3-methyladenine; D004815:Epidermal Growth Factor; D005293:Ferritins; D005978:Glutathione; C032854:leupeptin; D000225:Adenine; C027576:4-hydroxy-2-nonenal; D019803:Glutathione Disulfide
keywords: Autophagy; Cell death; Ferritin; HNE; Hepatocytes; Lysosome