title: Differing molecular mechanisms appear to underlie early toxicity of prefibrillar HypF-N aggregates to different cell types.
abstract: Considerable attention has been paid to the high cytotoxic potential of small, prefibrillar aggregates of proteins/peptides, either associated or not associated with amyloid diseases. Recently, we reported that different cell types are variously affected by early aggregates of the N-terminal domain of the prokaryotic hydrogenase maturation factor HypF (HypF-N), a protein not involved in any disease. In this study, we provide detailed information on a chain of events triggered in Hend murine endothelial cells and IMR90 fibroblasts, which have previously been shown to be highly vulnerable or very resistant, respectively, to HypF-N aggregates. Initially, both cell lines displayed impaired viability upon exposure to HypF-N toxic aggregates; however, at longer exposure times, IMR90 cells recovered completely, whereas Hend cells did not. In particular, significant initial mitochondrial permeability transition (MPT) pore opening was found in IMR90 cells followed by a sudden repair of membrane integrity with rapid and efficient inhibition of cytochrome c and AIF release, and upregulation of Bcl-2. The greater resistance of IMR90 fibroblasts may also be due to a higher cholesterol content in the plasma membrane, which disfavours interaction with the aggregates. In contrast, Hend cells, which have less membrane cholesterol, showed delayed MPT opening with prolonged translocation of cytochrome c into the cytosol. Finally, the caspase 9 active fragment was increased significantly in both Hend and IMR90 cells; however, only Hend cells showed caspase 8 and caspase 3 activation with DNA fragmentation. From our data, the different responses of the two cell types to the same aggregates appear to be associated with two key events: (a) aggregate interaction with the plasma membrane, disfavoured by a high level of membrane cholesterol; and (b) alterations in mitochondrial functionality, leading to the release of pro-apoptotic stimuli, which are counteracted by upregulation of Bcl-2.
authors.affiliation: Department of Biochemical Sciences, University of Florence, Italy. cristina.cecchi@unifi.it
authors.forename: Cristina
authors.initials: C
authors.lastname: Cecchi
authors.forename: Anna
authors.initials: A
authors.lastname: Pensalfini
authors.forename: Serena
authors.initials: S
authors.lastname: Baglioni
authors.forename: Claudia
authors.initials: C
authors.lastname: Fiorillo
authors.forename: Roberto
authors.initials: R
authors.lastname: Caporale
authors.forename: Lucia
authors.initials: L
authors.lastname: Formigli
authors.forename: Gianfranco
authors.initials: G
authors.lastname: Liguri
authors.forename: Massimo
authors.initials: M
authors.lastname: Stefani
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001426:Bacterial Proteins / Q000378:metabolism* / Q000633:toxicity*; D002118:Calcium / Q000378:metabolism; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D051379:Mice; D010084:Oxidation-Reduction; D013997:Time Factors
chemical_list: D001426:Bacterial Proteins; C108878:HypF protein, Bacteria; D000255:Adenosine Triphosphate; D002784:Cholesterol; C505530:CASP8 protein, human; C506433:CASP9 protein, human; C505526:Casp8 protein, mouse; C506434:Casp9 protein, mouse; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D002118:Calcium