Mechanisms of Induction of Skin Cancers By UV Light

描述（由申请人提供）：本补助金中提出的研究 应用程序的重点是调查参与的机制 CD 1基因敲除小鼠对紫外线致癌作用的抗性。这些理由 研究是基于一个新的和有趣的观察，CD 1基因敲除小鼠 对紫外线致癌作用和光毒性具有极强的抵抗力， 与CD 1 +/+小鼠相比，含有大量凋亡的角质形成细胞。我们 最近的研究表明，紫外线诱导CD 1蛋白的表达， 野生型小鼠皮肤，表明CD 1是UV的靶点。基于这些 结果，我们假设CD 1-/-小鼠对紫外线致癌的抵抗力 可能与持续消除紫外线损伤有关， 恶性角质细胞以下具体目标将用于测试这一点 假设：（1）确定是否将CD 1基因转染入CD 1-/- 小鼠角质形成细胞恢复对紫外线诱导的凋亡的抗性， 肿瘤转化的易感性，（2）确定是否 CD 1-/-小鼠对紫外线致癌作用的抵抗力增加是由于持续的 从皮肤中消除UV损伤的角质形成细胞，和（3）确定 CD 1-/-小鼠皮肤是否对UV诱导的p53突变具有抗性， CD 1 +/+小鼠皮肤。 为了研究特定目标1，新生儿CD 1-/-和+/+的角质形成细胞培养物 将建立和测试其敏感性紫外线诱导 凋亡、终末分化和肿瘤转化。在 此外，将检测来自CD 1-/-和+/+小鼠的成纤维细胞，以确定 该缺陷是普遍的或局限于角质形成细胞。更重要的是，CD 1基因 将其转染到CD 1-/-角质形成细胞中，以研究CD 1在 紫外线诱导的细胞凋亡、终末分化和肿瘤转化。 为了研究特异性目的2和3，将CD 1-/-和+/+小鼠暴露于急性或慢性炎症。 慢性紫外线照射和他们的皮肤分析（i）晒伤和 TUNEL阳性细胞，（ii）细胞周期和凋亡相关基因的表达 蛋白质如p53、p21 Waf 1/Cip 1、Fas、Fas-L、Bax和Bcl-2，和（ii） UV特异性p53诱导的动力学（发生率、潜伏期和持续性） 通过等位基因特异性聚合酶链反应检测突变。 这些研究产生的数据应该为CD 1在以下方面的作用提供新的线索： 光致癌作用这些信息对于理解 紫外线，CD 1基因和皮肤癌之间的相互作用。虽然重要的 CD 1分子在抗原提呈和免疫调节中的作用 虽然已知，但实际上对其在皮肤癌中的作用一无所知 发展我们的初步发现，抗紫外线致癌作用， CD 1-/-小鼠中凋亡增加，表明CD 1在细胞周期中的作用 调控因为紫外线会导致免疫抑制和诱发皮肤癌， CD 1蛋白在抗原呈递和免疫调节中起作用， 了解CD 1-/-小鼠对紫外线的抵抗机制 致癌作用也可能有助于理解 紫外线致癌和紫外线诱导的免疫抑制之间的联系。此外该 结果可能表明，皮肤癌也涉及类似的机制， 人类的发展。

DESCRIPTION (provided by applicant): The studies proposed in this grant application are focused on investigating the mechanisms involved in the resistance of CD1 knockout mice to UV carcinogenesis. The rationale for these studies is based on a novel and interesting observation that CD1 knockout mice are extremely resistant to UV carcinogenesis and phototoxicity, and their skin contains a high number of apoptotic keratinocytes compared to CD1+/+ mice. Our recent studies indicate that UV induces the expression of CD1 protein in wild-type mouse skin, suggesting that CD1 is a target for UV. Based on these results, we hypothesize that the resistance of CD1-/- mice to UV carcinogenesis may be associated with persistent elimination of UV damaged, potentially malignant keratinocytes. The following specific aims will be used to test this hypothesis: (1) Determine whether transfection of the CD1 gene into CD1-/- mouse keratinocytes restores resistance to UV-induced apoptosis and susceptibility to neoplastic transformation, (2) Determine whether the increased resistance of CD1-/- mice to UV carcinogenesis is due to persistent elimination of UV-damaged keratinocytes from the skin, and (3) Determine whether CD1-/-mouse skin is resistant to UV-induced p53 mutations compared to CD1+/+ mouse skin. To study Specific Aim 1, keratinocyte cultures from neonatal CD1-/- and +/+ will be established and tested for their susceptibility to UV-induced apoptosis, terminal differentiation, and neoplastic transformation. In addition, fibroblasts from CD1-/- and +/+ mice will be tested to determine if the defect is general or limited to keratinocytes. More importantly, CD1 gene will be transfected into CD1-/- keratinocytes to investigate the role of CD1 in UV-induced apoptosis, terminal differentiation and neoplastic transformation. To study Specific Aims 2 and 3, CD1-/- and +/+ mice will be exposed to acute or chronic UV irradiation and their skin analyzed for (i) sunburn and TUNEL-positive cells, (ii) expression of cell cycle and apoptosis-associated proteins such as p53, p21 Waf1/Cip1, Fas, Fas-L, Bax, and Bcl-2, and (ii) kinetics (incidence, latency, and persistence) of induction of UV-specific p53 mutations by allele-specific polymerase chain reaction. Data generated from these studies should shed new light on the role of CD1 in photocarcinogenesis. This information is important for understanding the interactions between UV, the CD1 gene, and skin cancer. Although the important role of CD1 molecule in antigen presentation and immune regulation is well known, virtually nothing is known in regard to its role in skin cancer development. Our preliminary findings, resistance to UV carcinogenesis and increased apoptosis in CD1-/- mice, suggest a role for CD1 in cell cycle regulation. Because UV causes immunosuppression and induces skin cancer, and CD1 protein plays a role in antigen presentation and immune regulation, understanding the mechanisms involved in the resistance of CD1-/- mice to UV carcinogenesis may also contribute to the understanding of the relationship between UV carcinogenesis and UV-induced immunosuppression. In addition, the results may suggest that similar mechanisms be involved in skin cancer development in humans.