Extensive documentation has validated the role of UV irradiation as a tumor initiator and promoter, inducing both squamous and basal cell carcinomas. Human epidermis is a tissue which undergoes active metabolism of arachidonic acid to prostaglandins which is regulated by the action of prostaglandin H synthase (also known as cyclooxygenase). One mechanism for the promotional activity of UV light may involve its ability to induce prostaglandin formation. Work in our laboratory has demonstrated that acute exposure of human keratinocytes to UVB irradiation results in increased production of prostaglandin E2 (PGE2). When cultured human keratinocytes were examined after irradiation with 30 mJ/cm2 UVB in vitro, Western blot analysis showed a 6-fold increase in COX-2 protein which was evident at 6 h and peaked 24 h after irradiation. Furthermore, when human subjects were irradiated on sun-protected skin with up to four times their minimal erythema dosage (MED) and biopsied 24 h later, upregulation of COX-2 protein expression was observed via immunofluorescence microscopy. RNAase protection assays supported this observation, showing induction of COX-2 message which peaked at approximately 12 h following irradiation in vitro. Furthermore, human squamous cell carcinoma biopsies exhibited strongly enhanced staining for COX-2 protein via immunohistochemistry and Western analysis when compared to normal non-sun-exposed control skin. Together, these data demonstrate acute upregulation of COX-2 via UVB irradiation and suggest the need for further studies of COX-2 expression as a potential pharmacological target mediating human skin tumor development.

UV light is a complete carcinogen, inducing both basal and squamous cell skin cancers. The work described uses the selective COX-2 inhibitor celecoxib to examine the efficacy of COX-2 inhibition in the reduction of UV light-induced skin tumor formation in hairless mice. UVA-340 sun lamps were chosen as a light source that effectively mimics the solar UVA and UVB spectrum. Hairless mice were irradiated for 5 days a week for a total dose of 2.62 J/cm(2). When 90% of the animals had at least one tumor, the mice were divided into two groups so that the tumor number and multiplicity were the same (P < 0.31). Half of the mice were then fed a diet containing 1500 p.p.m. celecoxib. Tumor number, multiplicity and size were then observed for the next 10 weeks. Ninety-five percent of the tumors formed were histopathologically evaluated as squamous cell carcinoma. COX-2 expression and activity were increased in tumors. After 10 weeks, the difference in tumor number and multiplicity in the drug-treated group was 56% of UV controls (P < 0.001). The results show that the orally administered selective COX-2 inhibitor celecoxib prevents new tumor formation after the onset of photocarcinogenesis and suggest that treatment with celecoxib may be very useful in preventing UV-induced skin tumors in humans.

We reported that interstitial collagenase is produced by keratinocytes at the edge of ulcers in pyogenic granuloma, and in this report, we assessed if production of this metalloproteinase is a common feature of the epidermal response in a variety of wounds. In all samples of chronic ulcers, regardless of etiology, and in incision wounds, collagenase mRNA, localized by in situ hybridization, was prominently expressed by basal keratinocytes bordering the sites ofactive re-epithelialization indicating that collagenolytic activity is a characteristic response of the epidermis to wounding. No expression of mRNAs for 72-and 92-kD gelatinases or matrilysin was seen in keratinocytes, and no signal for any metalloproteinase was detected in normal epidermis. Immunostaining for type IV collagen showed that collagenase-positive keratinocytes were not in contact with an intact basement membrane and, unlike normal keratinocytes, expressed a5jh receptors. These observations suggest that cellmatrix interactions influence collagenase expression by epidermal cells. Indeed, as determined by ELISA, primary cultures of human keratinocytes grown on basement membrane proteins (Matrigel; Collaborative Research Inc., Bedford, MA) did not express significant levels of collagenase, whereas cells grown on type I collagen produced markedly increased levels. These results suggest that migrating keratinocytes actively involved in re-epithelialization acquire a collagenolytic phenotype upon contact with the dermal matrix. (J. Clin. Invest. 1993. 92:2858-2866

We present a cascade of proteolytic events catalyzed by the proteases secreted by cultured keratinocytes and fibroblasts that results in the activation of interstitial procollagenase. Cultured human skin fibroblasts constitutively secrete interstitial collagenase and stromelysin as proenzymes. In contrast, interstitial collagenase found in serum-free skin organ culture conditioned medium is activated. Cocultivation of the major cellular components of skin organ culture, dermal fibroblasts and epidermal keratinocytes, induces activation of interstitial procollagenase and prostromelysin in the presence of plasminogen. This activation occurs through a urokinasedependent pathway where added keratinocytes secrete the plasminogen activator urokinase, which converts p!asminogen into plasmin. Plasmin is capable of activating purified procollagenase and prostromelysin. Plasmin-dependent activation of procollagenase generates an enzyme species, by amino-terminal processing, identical to those generated by limited proteolysis with trypsin or treatment with organomercurial compounds. Catalytic amounts of activated stromelysin can in turn convert plasmin-or trypsin-activated collagenase into a fully active enzyme by removal of 15 amino acid residues from the carboxyl end of the enzyme. This results in a 5-to 8-fold increase in collagenase specific activity that is due to its proteolytic cleavage and not to the presence of the activator stromelysin. Stromelysin alone in both pro-and activated forms is not capable of efficient activation of human fibroblast interstitial procollagenase.Intensified tissue remodeling during morphogenesis, wound healing, and tumor invasion requires the presence of secreted metalloproteases capable of initiating the degradation of macromolecules ofthe extracellular matrix. Several enzymes of this class have been identified: fibroblast (1-3) and granulocyte (4,5) collagenases, which degrade interstitial collagens, type IV collagenase, which degrades both basement membrane collagen and denatured collagen (gelatin) (6-8), and stromelysin, which degrades proteoglycans (9-13). We have determined the primary structure of fibroblast interstitial collagenase (14), stromelysin (12), and type IV collagenase (7), demonstrated a close structural relationship between these proteins (7), and compared their substrate specificities. Analysis of the genomic organization of the human fibroblast interstitial collagenase gene (15) in comparison with that of rabbit collagenase (16) and rat stromelysin (17) revealed that a close structural relationship on the protein level is reflected in the very similar genomic organization of at least two members of the secreted metalloprotease gene family. Interstitial collagenase (1, 18), stromelysin (12), and type IV collagenase (7) are constitutively secreted by cultured human skin fibroblasts in a proenzyme form and are subject to activation extracellularly. Both procollagenase and prostromelysin are secreted in two forms, one of which contains N-linked complex oligosacc...

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Ultraviolet radiation (UVR) has widespread effects on the biology and integrity of the skin barrier. Research on the mechanisms that drive these changes, as well as their effect on skin barrier function has been ongoing since the 1980s. However, no studies have examined the impact of UVR on nanoparticle skin penetration. Nanoparticles (NP) are commonly used in sunscreens and other cosmetics, and since consumer use of sunscreen is often applied to sun damaged skin, the effect of UVR on NP skin penetration is a concern due to potential toxicity. In this study we investigate nanoparticle skin penetration by employing an in vivo semiconductor quantum dot nanoparticle (QD) model system. This model system improves NP imaging capabilities and provides additional primary interest due to widespread and expanding use of QD in research applications and manufacturing. In our experiments, carboxylated QD were applied to the skin of SKH-1 mice in a glycerol vehicle with and without UVR exposure. The skin collection and penetration patterns were evaluated 8 and 24 hours after QD application using tissue histology, confocal microscopy, and transmission electron microscopy (TEM) and EDAX analysis. Low levels of penetration were seen in both the non-UVR exposed mice and the UVR exposed mice. Qualitatively higher levels of penetration were observable in the UVR exposed mice. These results are the first for in vivo QD skin penetration, and provide important insight into the ability of QD to penetrate intact and UVR compromised skin barrier. Our findings raise concern that NP of similar size and surface chemistry, such as metal oxide NP found in sunscreens, may also penetrate UV damaged skin.

Background Preclinical studies indicate that the enzyme cyclooxygenase 2 plays an important role in ultraviolet-induced skin cancers. We evaluated the efficacy and safety of celecoxib, a cyclooxygenase 2 inhibitor, as a chemopreventive agent for actinic keratoses, the premalignant precursor of nonmelanoma skin cancers, and for nonmelanoma skin cancers, including cutaneous squamous cell carcinomas (SCCs) and basal cell carcinomas (BCCs). Methods A double-blind placebo-controlled randomized trial involving 240 subjects aged 37–87 years with 10–40 actinic keratoses was conducted at eight US academic medical centers. Patients were randomly assigned to receive 200 mg of celecoxib or placebo administered orally twice daily for 9 months. Subjects were evaluated at 3, 6, 9 (ie, completion of treatment), and 11 months after randomization. The primary endpoint was the number of new actinic keratoses at the 9-month visit as a percentage of the number at the time of randomization. In an intent-to-treat analysis, the incidence of actinic keratoses was compared between the two groups using t tests. In exploratory analyses, we evaluated the number of nonmelanoma skin cancers combined and SCCs and BCCs separately per patient at 11 months after randomization using Poisson regression, after adjustment for patient characteristics and time on study. The numbers of adverse events in the two treatment arms were compared using χ 2 or Fisher exact tests. All statistical tests were two-sided. Results There was no difference in the incidence of actinic keratoses between the two groups at 9 months after randomization. However, at 11 months after randomization, there were fewer nonmelanoma skin cancers in the celecoxib arm than in the placebo arm (mean cumulative tumor number per patient 0.14 vs 0.35; rate ratio [RR] = .43, 95% confidence interval [CI] = 0.24 to 0.75; P = .003). After adjusting for age, sex, Fitzpatrick skin type, history of actinic keratosis at randomization, nonmelanoma skin cancer history, and patient time on study, the number of nonmelanoma skin cancers was lower in the celecoxib arm than in the placebo arm (RR = 0.41, 95% CI = 0.23 to 0.72, P = .002) as were the numbers of BCCs (RR = 0.40, 95% CI = 0.18 to 0.93, P = .032) and SCCs (RR = 0.42, 95% CI = 0.19 to 0.93, P = .032). Serious and cardiovascular adverse events were similar in the two groups. Conclusions Celecoxib may be effective for prevention of SCCs and BCCs in individuals who have extensive actinic damage and are at high risk for development of nonmelanoma skin cancers.

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