Release by ultraviolet B (u.v.B) radiation of nitric oxide (NO) from human keratinocytes: a potential role for nitric oxide in erythema production.
1. The mechanism of human sunburn is poorly understood but its characteristic features include the development of erythema. In this study we attempted to determine whether human keratinocytes possess a nitric oxide (NO) synthase (NOS), if this enzyme could be activated to release NO following exposure to ultraviolet B (u.v.B) and to define whether this photo-induced response could be involved in the pathogenesis of sunburn erythema. 2. Treatment of human keratinocytes with various doses of u.v.B (290-320 nm) radiation (up to 100 mJ cm-2) resulted in a dose-dependent release of NO and cyclic GMP production that was reduced by NG-monomethyl-L-arginine (L-NMMA). 3. u.v.B irradiation of keratinocyte cytosol at varying doses (up to 50 mJ cm-2), resulted in a gradual rise in NO production, with a concomitant increase in soluble guanylate cyclase activity (sGC). 4. NOS isolated from the keratinocyte cytosol was constitutively expressed and was dependent on NADPH, Ca2+/calmodulin, tetrahydrobiopterin and flavins. 5. In reconstitution experiments, when purified NOS was added to purified sGC, both isolated from keratinocyte cytosol, a four fold increase in cyclic GMP was observed. The GMP was increased by NO synthesized following u.v.B radiation (up to 20 mJ cm-2) of NOS. 6. In in vivo experiments, guinea-pigs were subjected to u.v.B light. A Protection Factor (PF) of 8.71 +/- 2.85 was calculated when an emulsified cream formulation containing L-NMMA (2%) was applied to their skin. 7. The present results indicate that u.v.B radiation acts as a potent stimulator of NOS in keratinocytes. NO is lipophilic and may diffuse out of the keratinocytes, activating sGC in endothelial cells and neighbouring smooth muscle cells. This may be a major part of the integrated response of the skin leading to vasodilatation and erythema.
Deliconstantinos
G
G
Department of Physiology, University of Athens Medical School, Greece.
Villiotou
V
V
Stravrides
J C
JC
D000818:Animals; D002956:Citrulline / Q000096:biosynthesis; D003600:Cytosol / Q000378:metabolism / Q000528:radiation effects; D004890:Erythema / Q000209:etiology / Q000378:metabolism / Q000503:physiopathology*; D006157:Guanosine Monophosphate / Q000378:metabolism; D006168:Guinea Pigs; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism / Q000528:radiation effects*; D009569:Nitric Oxide / Q000378:metabolism / Q000502:physiology*; D012867:Skin / Q000098:blood supply; D014407:Tumor Cells, Cultured; D014466:Ultraviolet Rays
D002956:Citrulline; D009569:Nitric Oxide; D006157:Guanosine Monophosphate