EXPERIMENTAL TRANSFORMATION OF HUMAN MELANOCYTES IN VIVO

人类黑色素瘤发展的实验动物模型已经被 很难建立，因为独特的建筑和 人体皮肤的功能特性阻碍了对其的研究 疾病的病因、诊断和预防。流行病学 观察表明，紫外线(UV)照射在 人类黑色素瘤的病因仍然缺乏直接证据。这个 前驱发育不良痣的组织学和生物学诊断 早期原发性黑色素瘤持续存在困难和争议， 即使在专家中，目前对基因的了解也很少 突变及其对黑色素瘤发生的重要性。 为了更好地了解人类黑色素瘤的发展和检测 用新的分子标记在最早的阶段发现黑素细胞病变， 我们在免疫缺陷的RAG-1小鼠身上建立了一种人皮肤移植模型 其中新生儿的技能暴露在化学致癌物中 7，12-二甲基(A)苯并菲(DMBA)紫外光辐照UP 到10个月。对该模型的初步研究揭示了 黑素细胞增生症、雀斑、异型增生和黑色素瘤。使用 精细的移植技术我们现在能够移植成人的皮肤和 将选择具有高度易感性的捐赠者 用于确定最佳紫外线波长的发育不良痣和/或黑色素瘤 用于诱导黑素细胞损伤并制定新的诊断标准 黑素细胞体内转化，包括增殖、逃逸 黑素细胞由角质形成细胞控制，失去与黑素细胞的接触 与基底膜相关的形态变化 黑素细胞异常增殖症。然后我们将定义新的分子标记 基因芯片转化技术在基因表达分析中的应用 来自显微解剖损伤或短期培养的细胞。这些首字母 筛查方法之后是免疫组织化学和原位 杂交分析提供了基因表达的信息 单个细胞分别在RNA和蛋白质水平。去调查 主要检查点基因的作用，我们将转导黑素细胞在 用E6基因或人乳头瘤病毒体外检测P53 隔离或与H-ra一起用于RAS激活。转导黑素细胞 将被包括在皮肤重建中，由真皮和 移植前的表皮进行活体和长期紫外线照射。我们的 独特的体外/体内模型适合于研究病因学， 早期发现和预防人类黑色素瘤。

Experimental animal models for human melanoma development have been difficult to establish because of the unique architectural and functional characteristics of human skin hampering investigations on etiology, diagnosis, and prevention of the disease. Epidemiological observations suggest a role for ultraviolet (UV) light irradiation in the etiology of human melanoma but direct evidence is still missing. The histological diagnosis of precursor dysplastic nevi and biologically early primary melanomas continuous to be difficult and controversial, even among experts, and there is currently little understanding of gene aberrations and their importance for melanoma development. To better understand development of human melanoma and to detect melanocytic lesions at the earliest stages with new molecular markers, we have developed a human skin graft model in immunodeficient RAG-1 mice in which human skill from newborns is exposed to the chemical carcinogen 7,12-dimethyl(a)benzanthracene (DMBA) followed by UV irradiation for up to 10 months. Preliminary studies with this model have revealed melanocytic hyperplasia, lentigo, dysplasia, and melanoma. Using a refined grafting technique we are now able to graft skin from adults and will select donors who have a high susceptibility for developing dysplastic nevi and/or melanomas to determine the optimal UV wavelength for induction of melanocytic lesions and to develop new criteria for melanocyte transformation in vivo, including proliferation, escape of melanocytes from control by keratinocytes, loss of melanocyte contact from basement membrane and morphological changes associated with melanocytic dysplasia. We will then define new molecular markers for transformation using microarrays for analyses of mRNA expression in cells from microdissected lesions or short-term cultures. These initial screening approaches are followed by immunohistochemistry and in situ hybridization analyses to provided information on gene expression in single cells at the RNA and protein levels, respectively. To investigate the role of major check point genes, we will transduce melanocytes in vitro with either the E6 gene or human papilloma virus for p53 sequestration or with H-ra for ras activation. Transduced melanocytes will then be included into skin reconstructs consisting of dermis and epidermis prior to grafting in vivo and long-term UV irradiation. Our unique in vitro/in vivo models are suitable to investigate etiology, early detection, and prevention of human melanoma.