We have demonstrated that in human epidermis and in sweat on the skin surface nitrate is the predominant NO-related product, followed by nitrite, with only very low concentrations of RSNOs being present. Both nitrate and nitrite are available from exogenous and endogenous sources. Green leafy vegetables make up 60–80% of the daily nitrate intake of those on a typical Western diet (<!--bib66-->
Ysart et al., 1999) whereas nitrite is used as a preservative for meat and fish (<!--bib29-->
Lundberg et al., 2004). The main endogenous source of nitrate and nitrite is the L-arginine–NO pathway, which is constitutively active in all cell types in the body. L-arginine is converted to NO by inducible nitric oxide synthase, an enzyme which is upregulated in human skin 8–10 hours after UV exposure (<!--bib24-->
Kuhn et al., 1998). Nitrite can also be generated from commensal bacteria in the digestive system by nitrate reduction. An enterosalivary recirculation pathway exists, with 25% of all circulating nitrate being taken up by the salivary glands and secreted in saliva. The oral cavity contains large numbers of nitrate-reducing bacteria, with the result that saliva is the main source of plasma nitrite (<!--bib4-->
Benjamin et al., 1994). <!--bib28-->
Lundberg and Govoni (2004) have demonstrated that dietary nitrate ingestion influences both plasma nitrate and nitrite but not RSNO levels. Analysis of our data suggests that the concentration of NO-related products in an individual's plasma will influence the concentration of NO-related products found in their sweat and superficial vascular dermis. Thus, dietary nitrate may offer a source for manipulation of cutaneous NO-related products.
Historically, evidence for the existence of stored forms of NO dates back to experiments performed in the 1950s on the relaxant effect of light on vascular smooth muscle (<!--bib15-->
Furchgott et al., 1961). Photorelaxation in rabbit aorta was shown by Furchgott
et al. to peak near 310 nm with a shoulder near 350 nm. The role of nitrite in photorelaxation was implicated when a dominant peak was shown at 355 nm following incubation with nitrite (<!--bib14-->
Furchgott, 1971). More recently, <!--bib43-->
Rodriguez et al. (2003) have shown that in rat vascular tissue RSNOs, RNNOs, and nitrite have photoactivity
in vitro but nitrate shows no appreciable activity. They calculated the action spectra for NO release from RSNOs to be 310–340 nm and from nitrite 310 and 350 nm.
It has been hypothesized that NO-related products stored in human skin may be involved in the acute response to UV (<!--bib38-->
Paunel et al., 2005). Paunel
et al. demonstrated formation of NO due to photodecomposition of nitrite and RSNOs maximal within 20 minutes of UVA exposure of
ex vivo full-thickness human skin (<!--bib38-->
Paunel et al., 2005). We have now demonstrated this effect in human skin
in vivo with cutaneous microdialysis. Dialysate sampling of the superficial vascular dermis revealed an increase in NO-related products maximal 30 minutes after exposure to a biologically relevant dose of UVA1 (30 J cm[SUP]
−2[/SUP]). Gaseous NO released within the dermis will undergo oxidative decomposition on entering the extracellular vascular space forming aqueous nitrite. Although both RSNOs and nitrite will release NO in the UVA spectrum, the relative concentrations of nitrite and RSNOs that we demonstrated in human skin suggest that nitrite is likely to be the predominant photoactive NO-related product. NO may play an anti-apoptotic role in human skin following UVR (<!--bib51-->
Suschek et al., 1999, <!--bib52-->
2003; <!--bib59-->
Weller et al., 2003) and it is likely that cutaneous NO-related products offer an immediate enzyme-independent source of NO, allowing protection within 30 minutes of exposure to UVR. As proposed by <!--bib38-->
Paunel et al. (2005), this enzyme-independent NO release bridges the time gap following UV exposure before the upregulation of inducible nitric oxide synthase and enzyme-dependent NO release is maximal.
Interindividual variation in the concentration of NO-related products in human saliva, plasma, sweat, epidermis, and superficial vascular dermis was seen in our subjects and was similar in each of the different biological samples. Gladwin (<!--bib16-->
Gladwin et al., 2005) has suggested that the majority of nitrite in tissues originates from the exogenous intake of nitrite and nitrate and not from endogenous sources, thus resulting in great variation in tissue nitrite levels with nitrate and nitrite intake. In contrast plasma levels of nitrite vary only slightly suggesting the existence of regulatory pathways in blood (<!--bib8-->
Bryan et al., 2005).
Enzyme-dependent NO production occurs in all cell types of human skin by at least one of the three NO synthase isoenzymes. NO is produced constantly by the endothelium at 4 nM per second (<!--bib32-->
Marley et al., 2001). We have confirmed the presence of NO-related products in the dermis and epidermis but have not isolated the storage site to a particular cell type. On exposure to UVA a significant increase in aqueous NO-related products is detected by cutaneous microdialysis, and this increase is reduced in the presence of a local vasoconstrictor. Both nitrite and RSNOs are transported in the bloodstream and are susceptible to photolysis by UVA. We suspect that much of the observed increase in NO-related products following UVA exposure comes directly from the vasculature, but with locally bound stores also contributing. Hypertension and ischemic heart disease both correlate with latitude (<!--bib13-->
Fleck, 1989; <!--bib46-->
Rostand, 1997), increasing in incidence with distance from the equator. Much of this is probably due to racial and dietary factors, and a correlation has been also recently shown between UV-induced vitamin D synthesis and reduced ischemic heart disease (<!--bib56-->
Wang et al., 2008). Our data, showing UV release of skin-bound NO-related products, suggest this as an alternative mechanism by which UV exposure may have beneficial cardiovascular effects.
We present data that confirm and quantify the presence of NO-related products both in human epidermis, superficial vascular dermis and skin surface sweat. These species have the capability of releasing NO, and we have demonstrated such release within 30 minutes of UVA exposure. We believe these findings to be of great significance, in the context of the skin as the largest “organ” of the human body (10–20 dm[SUP]
3[/SUP]), which thus offers a considerable store. We believe that enzyme-independent NO release is an acute mechanism preventing UV-induced keratinocyte apoptosis (<!--bib38-->
Paunel et al., 2005). It is known that NO-related products in plasma can be influenced by dietary intake of nitrate and we have demonstrated a relationship between the concentration of NO-related products in plasma and that of the superficial vascular dermis and sweat. We postulate that an individual's dietary consumption of green leafy vegetables may in part influence their cutaneous response to UVR.
