Nox1-based NADPH oxidase is the major source of UVA-induced reactive oxygen species in human keratinocytes.
UVA radiation is a major environmental stress on skin, causing acute and chronic photodamage. These responses are mediated by reactive oxygen species (ROS), although the cellular source of these ROS is unknown. We tested the hypotheses that UVA-induced activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase is required for ROS generation in human keratinocytes (HK) and that these ROS initiate rapid prostaglandin E2 (PGE2) synthesis. Treatment of HK with a non-toxic dose of UVA rapidly increased NADPH oxidase activity and intracellular ROS, which were partially blocked by an inhibitor of NADPH oxidase and by a mitochondria-selective antioxidant. Depleting the Nox1 isoform of the catalytic subunit of NADPH oxidase using small interfering RNA (siRNA) blocked the UVA-induced ROS increase, indicating that ROS produced by mitochondria or other sources are downstream from Nox1. Nox1 siRNA also blocked UVA-initiated PGE2 synthesis. The mechanism for activation of Nox1 is mediated by an increase in intracellular calcium. Ceramide, which has been proposed to mediate responses to UVA in HK, also activated NADPH oxidase. These results indicate that UVA activates Nox1-based NADPH oxidase to produce ROS that stimulate PGE2 synthesis, and that Nox1 may be an appropriate target for agents designed to block UVA-induced skin injury.
Valencia
Antonio
A
Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02114, USA.
Kochevar
Irene E
IE
D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002518:Ceramides / Q000494:pharmacology; D015232:Dinoprostone / Q000096:biosynthesis; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism / Q000528:radiation effects*; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000502:physiology*; D009861:Onium Compounds / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D014466:Ultraviolet Rays
D002518:Ceramides; D009861:Onium Compounds; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; C007517:diphenyleneiodonium; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases; C402799:NOX1 protein, human; D015232:Dinoprostone; D002118:Calcium