title: Quantification and Characterization of UVB-Induced Mitochondrial Fragmentation in Normal Primary Human Keratinocytes.
abstract: UV irradiation is a major environmental factor causing skin dryness, aging and cancer. UVB in particular triggers cumulative DNA damage, oxidative stress and mitochondrial dysfunction. The objective of our study was to provide both qualitative and quantitative analysis of how mitochondria respond to UVB irradiation in normal human epidermal keratinocytes (NHEK) of healthy donors, with the rationale that monitoring mitochondrial shape will give an indication of cell population fitness and enable the screening of bioactive agents with UVB-protective properties. Our results show that NHEK undergo dose-dependent mitochondrial fragmentation after exposure to UVB. In order to obtain a quantitative measure of this phenomenon, we implemented a novel tool for automated quantification of mitochondrial morphology in live cells based on confocal microscopy and computational calculations of mitochondrial shape descriptors. This method was used to substantiate the effects on mitochondrial morphology of UVB irradiation and of knocking-down the mitochondrial fission-mediating GTPase Dynamin-related protein 1 (DRP1). Our data further indicate that all the major mitochondrial dynamic proteins are expressed in NHEK but that their level changes were stronger after mitochondrial uncoupler treatment than following UVB irradiation or DRP1 knock-down. Our system and procedures might be of interest for the identification of cosmetic or dermatologic UVB-protective agents.
authors.affiliation: Molecular Biology of the Cell Laboratory, Ecole Normale Supérieure de Lyon, UMR 5239 CNRS - UCBL - ENS Lyon, 46 Allée d'Italie, 69364 Lyon Cedex 07, France.|SILAB, ZAC de la Nau, 19240 Saint-Viance, France.
authors.forename: Romain
authors.initials: R
authors.lastname: Jugé
authors.affiliation: SILAB, ZAC de la Nau, 19240 Saint-Viance, France.
authors.forename: Josselin
authors.initials: J
authors.lastname: Breugnot
authors.affiliation: Molecular Biology of the Cell Laboratory, Ecole Normale Supérieure de Lyon, UMR 5239 CNRS - UCBL - ENS Lyon, 46 Allée d'Italie, 69364 Lyon Cedex 07, France.
authors.forename: Célia
authors.initials: C
authors.lastname: Da Silva
authors.affiliation: SILAB, ZAC de la Nau, 19240 Saint-Viance, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Bordes
authors.affiliation: SILAB, ZAC de la Nau, 19240 Saint-Viance, France.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Closs
authors.affiliation: Molecular Biology of the Cell Laboratory, Ecole Normale Supérieure de Lyon, UMR 5239 CNRS - UCBL - ENS Lyon, 46 Allée d'Italie, 69364 Lyon Cedex 07, France.|ISEM - Institut des Sciences de l'Evolution de Montpellier, UMR 5554, Université de Montpellier | CNRS | IRD | EPHE, Place Eugène Bataillon, 34095 Montpellier, France.
authors.forename: Abdel
authors.initials: A
authors.lastname: Aouacheria
mesh_terms: D017209:Apoptosis; D002470:Cell Survival / Q000528:radiation effects; D002478:Cells, Cultured; D019295:Computational Biology / Q000379:methods; D004249:DNA Damage; D034281:Dynamins; D020558:GTP Phosphohydrolases / Q000235:genetics*; D055785:Gene Knockdown Techniques; D064368:Healthy Volunteers; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000528:radiation effects*; D018613:Microscopy, Confocal; D008869:Microtubule-Associated Proteins / Q000235:genetics*; D008928:Mitochondria / Q000235:genetics / Q000528:radiation effects*; D063154:Mitochondrial Dynamics / Q000528:radiation effects*; D024101:Mitochondrial Proteins / Q000235:genetics*; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D008869:Microtubule-Associated Proteins; D024101:Mitochondrial Proteins; D017382:Reactive Oxygen Species; D020558:GTP Phosphohydrolases; C110768:DNM1L protein, human; D034281:Dynamins