title: Polyethylenimine architecture-dependent metabolic imprints and perturbation of cellular redox homeostasis.
abstract: Polyethylenimines (PEIs) are among the most efficient polycationic non-viral transfectants. PEI architecture and size not only modulate transfection efficiency, but also cytotoxicity. However, the underlying mechanisms of PEI-induced multifaceted cell damage and death are largely unknown. Here, we demonstrate that the central mechanisms of PEI architecture- and size-dependent perturbations of integrated cellular metabolomics involve destabilization of plasma membrane and mitochondrial membranes with consequences on mitochondrial oxidative phosphorylation (OXPHOS), glycolytic flux and redox homeostasis that ultimately modulate cell death. In comparison to linear PEI, the branched architectures induced greater plasma membrane destabilization and were more detrimental to glycolytic activity and OXPHOS capacity as well as being a more potent inhibitor of the cytochrome c oxidase. Accordingly, the branched architectures caused a greater lactate dehydrogenase (LDH) and ATP depletion, activated AMP kinase (AMPK) and disturbed redox homeostasis through diminished availability of nicotinamide adenine dinucleotide phosphate (NADPH), reduced antioxidant capacity of glutathione (GSH) and increased burden of reactive oxygen species (ROS). The differences in metabolic and redox imprints were further reflected in the transfection performance of the polycations, but co-treatment with the GSH precursor N-acetyl-cysteine (NAC) counteracted redox dysregulation and increased the number of viable transfected cells. Integrated biomembrane integrity and metabolomic analysis provides a rapid approach for mechanistic understanding of multifactorial polycation-mediated cytotoxicity, and could form the basis for combinatorial throughput platforms for improved design and selection of safer polymeric vectors.
authors.affiliation: Nanomedicine Research Group and Centre for Pharmaceutical Nanotechnology and Nanotoxicology, Department of Pharmacy, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen Ø, Denmark; NanoScience Centre, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen Ø, Denmark; Genome Integrity Unit, Danish Cancer Society Research Center, Copenhagen, Denmark. Electronic address: arnaldur.hall@sund.ku.dk.
authors.forename: Arnaldur
authors.initials: A
authors.lastname: Hall
authors.affiliation: Nanomedicine Research Group and Centre for Pharmaceutical Nanotechnology and Nanotoxicology, Department of Pharmacy, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen Ø, Denmark; NanoScience Centre, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen Ø, Denmark.
authors.forename: Ladan
authors.initials: L
authors.lastname: Parhamifar
authors.affiliation: Genome Integrity Unit, Danish Cancer Society Research Center, Copenhagen, Denmark.
authors.forename: Marina Krarup
authors.initials: MK
authors.lastname: Lange
authors.affiliation: Institute of Preventive Medicine, Bispebjerg and Frederiksberg Hospitals, The Capital Region, Copenhagen, Denmark.
authors.forename: Kathrine Damm
authors.initials: KD
authors.lastname: Meyle
authors.affiliation: Seahorse Bioscience Europa, Copenhagen, Denmark.
authors.forename: May
authors.initials: M
authors.lastname: Sanderhoff
authors.affiliation: Nanomedicine Research Group and Centre for Pharmaceutical Nanotechnology and Nanotoxicology, Department of Pharmacy, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen Ø, Denmark; NanoScience Centre, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen Ø, Denmark.
authors.forename: Helene
authors.initials: H
authors.lastname: Andersen
authors.affiliation: Section of Environmental Health, Department of Public Health, University of Copenhagen, DK-1014 Copenhagen K, Denmark.
authors.forename: Martin
authors.initials: M
authors.lastname: Roursgaard
authors.affiliation: Nanomedicine Research Group and Centre for Pharmaceutical Nanotechnology and Nanotoxicology, Department of Pharmacy, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen Ø, Denmark; NanoScience Centre, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen Ø, Denmark.
authors.forename: Anna Karina
authors.initials: AK
authors.lastname: Larsen
authors.affiliation: Seahorse Bioscience Europa, Copenhagen, Denmark.
authors.forename: Per Bo
authors.initials: PB
authors.lastname: Jensen
authors.affiliation: Genome Integrity Unit, Danish Cancer Society Research Center, Copenhagen, Denmark.
authors.forename: Claus
authors.initials: C
authors.lastname: Christensen
authors.affiliation: Genome Integrity Unit, Danish Cancer Society Research Center, Copenhagen, Denmark; Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacky University, CZ-775 15 Olomouc, Czech Republic.
authors.forename: Jiri
authors.initials: J
authors.lastname: Bartek
authors.affiliation: Nanomedicine Research Group and Centre for Pharmaceutical Nanotechnology and Nanotoxicology, Department of Pharmacy, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen Ø, Denmark; NanoScience Centre, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen Ø, Denmark. Electronic address: moien.moghimi@sund.ku.dk.
authors.forename: Seyed Moein
authors.initials: SM
authors.lastname: Moghimi
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000975:Antioxidants / Q000378:metabolism / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D019069:Cell Respiration / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004734:Energy Metabolism / Q000187:drug effects*; D005978:Glutathione / Q000378:metabolism; D006706:Homeostasis; D006801:Humans; D007700:Kinetics; D051336:Mitochondrial Membranes / Q000187:drug effects* / Q000378:metabolism; D015394:Molecular Structure; D008970:Molecular Weight; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000187:drug effects*; D010101:Oxygen Consumption / Q000187:drug effects; D011094:Polyethyleneimine / Q000737:chemistry / Q000633:toxicity*; D017382:Reactive Oxygen Species / Q000378:metabolism; D013329:Structure-Activity Relationship; D014162:Transfection / Q000379:methods*
chemical_list: D000975:Antioxidants; D017382:Reactive Oxygen Species; D000255:Adenosine Triphosphate; D011094:Polyethyleneimine; D005978:Glutathione
keywords: Bioenergetics; Cell death; Glycolytic flux; Mitochondrial dysfunction; Oxidative stress; Polyethylenimine