title: 3-Aminobenzamide protects primary human keratinocytes from UV-induced cell death by a poly(ADP-ribosyl)ation independent mechanism.
abstract: Poly(ADP-ribosyl)ation (PARylation) is a NAD(+)-dependent protein modification carried out by PARP [poly(ADP-ribose) polymerase] enzymes. Here we set out to investigate whether PARylation regulates UVB-induced cell death in primary human keratinocytes. We used the benchmark PARP inhibitor 3-aminobenzamide (3AB) and a more potent and specific inhibitor PJ34 and found that UVB (0.05-0.2J/cm(2)) induced a dose dependent loss of viability that was prevented by 3AB but not by PJ34. Similarly to PJ34, two other new generation PARP inhibitors also failed to protect keratinocytes from UVB-induced loss of viability. Moreover, silencing PARP-1 in HaCaT human keratinocytes sensitized cells to UVB toxicity but 3AB provided protection to both control HaCaT cells and to PARP-1 silenced cells indicating that the photoprotective effect of 3AB is independent of PARP inhibition. Lower UVB doses (0.0125-0.05J/cm(2)) caused inhibition of proliferation of keratinocytes which was prevented by 3AB but augmented by PJ34. UVB-induced keratinocyte death displayed the characteristics of both apoptosis (morphology, caspase activity, DNA fragmentation) and necrosis (morphology, LDH release) with all of these parameters being inhibited by 3AB and apoptotic parameters slightly enhanced by PJ34. UVA also caused apoptotic and necrotic cell death in keratinocytes with 3AB protecting and PJ34 sensitizing cells to UVA-induced toxicity. 3AB prevented UVB-induced mitochondrial membrane depolarization and generation of hydrogen peroxide. In summary, PARylation is a survival mechanism in UV-treated keratinocytes. Moreover, 3-aminobenzamide is photoprotective and acts by a PARP-independent mechanism at a premitochondrial step of phototoxicity.
authors.affiliation: Department of Medical Chemistry, Medical and Health Science Center, University of Debrecen, Debrecen, Hungary.
authors.forename: Petra
authors.initials: P
authors.lastname: Lakatos
authors.forename: Éva
authors.initials: É
authors.lastname: Szabó
authors.forename: Csaba
authors.initials: C
authors.lastname: Hegedűs
authors.forename: György
authors.initials: G
authors.lastname: Haskó
authors.forename: Pál
authors.initials: P
authors.lastname: Gergely
authors.forename: Péter
authors.initials: P
authors.lastname: Bai
authors.forename: László
authors.initials: L
authors.lastname: Virág
mesh_terms: D017209:Apoptosis / Q000187:drug effects* / Q000528:radiation effects; D001549:Benzamides / Q000494:pharmacology*; D015153:Blotting, Western; D020169:Caspases / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D007124:Immunoenzyme Techniques; D015603:Keratinocytes / Q000166:cytology / Q000187:drug effects* / Q000528:radiation effects; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D009336:Necrosis; D010616:Phenanthrenes / Q000494:pharmacology*; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D014466:Ultraviolet Rays
chemical_list: D001549:Benzamides; D004791:Enzyme Inhibitors; C434926:N-(oxo-5,6-dihydrophenanthridin-2-yl)-N,N-dimethylacetamide hydrochloride; D010616:Phenanthrenes; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D034741:RNA, Small Interfering; D011838:Radiation-Sensitizing Agents; C025160:3-aminobenzamide; D006861:Hydrogen Peroxide; D011065:Poly(ADP-ribose) Polymerases; D020169:Caspases