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Our data suggest a high oxidative stress due to the release of nitric oxide synthase and xanthine oxidase within the dilated spermatic vein. The reaction resulted in dramatic formation of nitric oxide, peroxynitrite and S-nitrosothiols, which are biologically active. Formation of peroxynitrite from the reaction of nitric oxide with superoxide could be a causative factor for impaired sperm function in patients with varicocele.

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Here we demonstrate that human keratinocytes possess a Ca2+/calmodulin-dependent particulate NO synthase that can be activated to release NO after exposure to UVB radiation. UVB irradiation (up to 20 mJ/cm2) of human keratinocyte plasma membranes resulted in a dose-dependent increase in NO and L-[3H]citrulline production that was inhibited by approx. 90% in the presence of N-monomethyl-L-arginine (L-NMMA). In time-course experiments with UVB-irradiated plasma membranes the changes in NO production were followed by analogous changes in soluble guanylate cyclase (sGC) activity. In reconstitution experiments, when particulate NO synthase was added to purified sGC isolated from keratinocyte cytosol, a 4-fold increase in cGMP was observed; the cGMP was increased by NO synthesized after UVB irradiation (up to 20 mJ/cm2) of particulate NO synthase. A 5-fold increase in superoxide (O2-) and a 7-fold increase in NO formation followed by an 8-fold increase in peroxynitrite (ONOO-) production by UVB (20 mJ/cm2)-irradiated keratinocyte microsomes was observed. UVB radiation (20 mJ/cm2) decreased plasma membrane lipid fluidity as indicated by steady-state fluorescence anisotropy. Membrane fluidity changes were prevented by L-NMMA. Changes in Arrhenius plots of particulate NO synthase in combination with changes in its allosteric properties induced by UVB radiation are consistent with a decreased fluidity of the lipid microenvironment of the enzyme. The present studies provide important new clues to the role of NO and ONOO- released by UVB-irradiated human keratinocytes in skin erythema and inflammation.