Cutaneous melanoma is the most lethal of human skin cancers in the United States, and its incidence in youngadults as well as in persons over the age of 65 has been steadily increasing over the last 50 years. A largebody of evidence supports the role of solar ultra-violet radiation (UVR) in being a major factor in the rise of thisdisease. However, it is still unclear how UVR, particularly recreational or intermittent exposure to such, causesmelanoma, especially since none of the mutated genes (e.g., N-RAS, BRAF, PTEN, and CDKN2A) that arefrequently found in melanoma typify UV-induced mutations. This incongruity, therefore, raises the possibilitythat these genes associated with melanoma could in fact be collaborating with UV-induced epigenetic changesas imposed by DNA hypo- or hyper-methylation and/or covalent modifications of core histones. Thus, our mainhypothesis is that UVR induces epigenetic alterations in the melanocytes of human skin and that this in turnleads to the activation of oncogenes and/or the silencing of tumor suppressor genes, ultimately resulting in thedevelopment of melanoma. A corollary of this hypothesis is that these types of alterations might also arisefrom indirect UVR-induced changes to the surrounding microenvironment. To establish proof of ourhypotheses, however, we first need to generate data as a point of reference for our future studies. Therefore,the following specific aims are proposed:
SPECIFIC AIM 1 : Evaluating the profiles of gene expression in melanocytes after exposure to UVR invivo.1a. Human volunteers, light source, spectral measurements, and UVR exposure.1b. Sample processing, staining and capture by LCM.1c. The isolation of RNA from captured melanocytes and/or keratinocytes.1d. GeneChip Probe Array and Data Analysis.
SPECIFIC AIM 2 : Constructing a 3-dimensional human-skin model2a. The Isolation of keratinocytes and melanocytes from human tissue.2b. Reconstructing 3-dimensional human-skin.
SPECIFIC AIM 3 : Characterizing gene expression patterns in melanocytes of 3-D culture after UVR.3a. Light source and spectral measurements.3b. Irradiation procedure and microarray analysis.
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