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Actinic Keratosis (AK) arises from sun-damaged skin and is the first clinical manifestation in the multistep process of skin carcinogenesis to invasive squamous cell carcinoma. Thus, it is an ideal target for chemopreventive efforts. Noninvasive measures of AK severity are needed to assess the efficacy of chemoprevention agents. We performed a pilot study on 20 participants to investigate the OCT appearance of sun-protected skin of the upper inner arm as well as sun-damaged skin and early AKs of the dorsal forearms, and to determine if features or quantitative measures in Optical Coherence Tomography (OCT) images could be used to reliably differentiate between these categories. OCT images of upper inner arm (normal appearing skin) showed skin layers and features (stratum corneum, epidermis, dermis, blood vessels) seen in previous studies; additionally in this participant group the subcutaneous fat layer was usually identified. Sun-damaged skin was characterized by increased signal in the epidermis and rapid attenuation of light. AKs were diverse in appearance but frequently characterized by high surface reflection, the presence of a low-signal band in the stratum corneum, and heterogeneous appearance in the epidermis/dermis. Significant differences were found between skin categories using measures of stratum corneum and epidermal/dermal depths and intensities. The presence of a dark band in the stratum corneum was 79% sensitive and 100% specific for AK. This study indicates that OCT holds promise as a useful technique for identifying and characterizing AKs and monitoring their response to chemoprevention agents.

Seven normal volunteers (six males and one female) with tanning skin types III or IV (Fitzpatrick scale) were given 10 daily subcutaneous injections of a superpotent synthetic analog of alpha-melanocyte stimulating hormone (alpha-MSH) over two weeks. This agent, [Nle4-D-Phe7]alpha-MSH, also called Melanotan-I (MT-I), was administered at a dose of 0.16 mg/kg/day (Monday-Friday), over a two week period. Tanning was measured serially using computerized light reflectance. This regimen induced tanning at 3 of 8 anatomic sites including the face, neck and forearm by comparison of baseline to (1) the end of the daily dosing period, (day 14), and (2) one week later, (day 21). Shave biopsies of the forearm taken at baseline and day 21 were analyzed by high performance liquid chromatography for eumelanin content which was measured as the permanganate oxidation product, pyrrole-2,3,5-tricarboxylic acid or PTCA. Pheomelanin content was measured as the hydroiodic acid digestion product, aminohydroxyphenylalanine (AHP). Eumelanin was also measured in the forehead skin samples of three subjects. The HPLC results show that mean (+/- SD) baseline eumelanin (PTCA) levels in forehead skin (n = 3) averaged 1.38 (+/- 0.87) ng/mg of wet skin tissue weight. Higher mean baseline levels of PTCA were detected in forearm skin (2.06 +/- 0.28 ng/mg wet weight, n = 7). One week after MT-I treatments ended, there was a mean (SD) 49% (+/- 17.6%) increase in forehead skin PTCA levels compared to baseline (P = 0.019, n = 3, by paired sample T-test). The mean (SD) increase in forearm skin PTCA levels was 98% (+/- 25.4%) over the same period (P = 0.003). In contrast, forearm pheomelanin expression following MT-I treatment did not significantly change from baseline. Overall, the MT-I regimen increased the eumelanin: pheomelanin ratio in forearm skin from 51:1 at baseline to 86:1 following MT-I (P = 0.054 by paired sample T-test). These results show that the tanning induced by MT-I in the face and forearm is associated with a significant increase in the eumelanin content of the human skin.

The incidence of skin cancer is higher than all other cancers and continues to increase, with an average annual cost over $8B in the United States. As a result, identifying molecular pathway alterations that occur with UV exposure to strategize more effective preventive and therapeutic approaches is essential. To that end, we evaluated phosphorylation of proteins within the PI3K/Akt and MAPK pathways by immunohistochemistry in sun-protected skin after acute doses of physiologically relevant solar simulated ultraviolet light (SSL) in 24 volunteers. Biopsies were performed at baseline, 5-minute, 1-, 5-, and 24-hour post SSL. Within the PI3K/Akt pathway, we found activation of Akt (Serine473) to be significantly increased at 5 hrs while mTOR (Serine2448) was strongly activated early and was sustained over 24-hour post SSL. Downstream we observed a marked and sustained increase in phospho-S6 (Serine235/S236) whereas phospho-4E-BP1 (Threonines37/46) was increased only at 24 hours. Within the MAPK pathway, SSL-induced expression of phospho-p38 (Threonine180/Tyrosine182) peaked at 1–5 hrs. ERK 1/2 was observed to be immediate and sustained post-SSL. Phosphorylation of histone H3 (Serine10), a core structural protein of the nucleosome, peaked at 5-hour post SSL. The expression of both p53 and COX-2 was increased at 5-hour and was maximal at 24-hour post SSL. Apoptosis was significantly increased at 24 hrs as expected and indicative of a sunburn-type response to SSL. Understanding the timing of key protein expression changes in response to SSL will aid in development of mechanistic-based approaches for the prevention and control of skin cancers.

Acute UVB irradiation of mouse skin results in activation of phospatidyinositol-3 (PI-3) kinase and mitogen-activated protein kinase (MAPK) pathways leading to altered protein phosphorylation and downstream transcription of genes. We determined whether activation of these pathways also occurs in human skin exposed to 4x minimal erythemic dose of UVB in 23 volunteers. Biopsies were taken prior to, at 30 min, 1 and 24 h post-UVB. In agreement with mouse studies, the earliest UV-induced changes in epidermis were seen in phospho-CREB (two- and five-fold at 30 min and 1 h) and in phospho-MAPKAPK-2 (three-fold at both 30 min and 1 h). At 1 h, phospho-c-JUN and phospho-p38 were increased five- and two-fold, respectively. Moreover, phospho-c-JUN and phospho-p38 were further increased at 24 h (12- and six-fold, respectively). Phospho-GSK-3beta was similarly increased at all time points. Increases in phospho-p53 (12-fold), COX-2 (four-fold), c-FOS (14-fold) and apoptosis were not seen until 24 h. Our data suggest that UVB acts through MAPK p38 and PI-3 kinase with phosphorylation of MAPKAPK-2, CREB, c-JUN, p38, GSK-3beta and p53 leading to marked increases in c-FOS, COX-2 and apoptosis. Validation of murine models in human skin will aid in development of effective skin cancer chemoprevention and prevention strategies.

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Unlike gravity feedings, the timed feedings resulted in a statistically significant loss of fat as compared with their controls. These findings should raise questions about how those infants in the neonatal intensive care unit are routinely gavage fed.

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