ANGIOGENIC FACTORS ASSOCIATED WITH UVEAL MELANOMA

与葡萄膜黑色素瘤相关的血管生成因子

• 批准号: 6024458
• 负责人: ARTHUR S POLANS
• 金额: $ 32.17万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-07-01 至 2003-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6024458
• 关键词: RNase protection assay; SDS polyacrylamide gel electrophoresis; angiogenesis; angiogenesis factor; angiostatins; electron microscopy; eye neoplasms; gene expression; human tissue; immunocytochemistry; immunofluorescence technique; in situ hybridization; laboratory mouse; light microscopy; melanoma; metastasis; northern blottings; nucleic acid sequence; polymerase chain reaction; scintillation counter; thromboplastin; tissue /cell culture; uvea; vascular endothelial growth factors; western blottings

Uveal melanoma is the most prevalent primary ocular tumor found in adults. The annual incidence of uveal melanoma is equivalent to the number of new cases of retinitis pigmentosa. The genetic alterations underlying the disease are unknown, and there are few prognostic indicators that can be observed clinically, contributing to enucleation as a principal method of treatment. Unfortunately, events leading to systemic metastases may have occured by the time the ocular symptoms are recognized, and death due to hepatic disease and other complications usually ensues. Clearly, improved methods are needed for the early detection and treatment of the disease. The current application focuses on the contributions of angiogenic growth factors and inhibitors to the progression of the disease. We hypothesize that the malignant and metastatic properties of such tumors depend on the simultaneous expression of multiple angiogenic factors, specifically Cyr61, Tissue Factor, and VEGF. Further, the balance between the expression of these growth factors and inhibitors determines the quiescent period prior to detectable metastases. Specific Aims include: 1. Verify the expression of Cyr61, Tissue Factor and VEGF in primary ocular melanoma using both archival specimens and fresh biopsies of tumor tissue. Methods of detection will include immunohistochemistry, Northern blot hybridization, RT-PCR, and in situ hybridization. Expression will be correlated with vascular density. 2. Assess the importance of Cyr61, Tissue Factor and VEGF during growth of the primary tumor and its metastases using an animal model. Transfected cell lines of uveal melanoma with varying levels of expression of these three genes will be transplanted into the anterior chambers of nude mice, and the growth of the primary ocular tumors, the extent of hepatic lesions, and the vascular density at both sites will be measured. 3. Determine the cellular localization of Cyr61, Tissue Factor and VEGF using a combination of immunocytochemical methods and cellular fractionation. 4. Identify additional angiogenic growth factors (angiopoietin-1) and inhibitors (angiostatin and endostatin) associated with uveal melanoma by RT_PCR, immunoblotting and Northern blot hybridization.

卵巢黑色素瘤是成年人发现的最普遍的原发性眼肿瘤。 紫veal黑色素瘤的年发病率相当于新的色素性视网膜炎病例的数量。 该疾病的遗传改变是未知的，几乎没有预后指标可以在临床上观察到，从而导致摘除作为主要治疗方法。不幸的是，在识别眼症状时可能发生了导致全身转移的事件，并且通常会导致肝病和其他并发症引起的死亡。显然，需要改进的方法来对疾病进行早期检测和治疗。当前的应用集中在血管生长因子和抑制剂对疾病进展的贡献上。 我们假设这种肿瘤的恶性和转移性特性取决于多种血管生成因子的同时表达，特别是CYR61，组织因子和VEGF。 此外，这些生长因子和抑制剂的表达之间的平衡决定了在可检测到的转移之前的静止期。具体目的包括：1。使用肿瘤组织的档案标本和新鲜的活检，验证原发性眼部黑色素瘤中CYR61，组织因子和VEGF的表达。 检测方法将包括免疫组织化学，北印迹杂交，RT-PCR和原位杂交。 表达将与血管密度相关。 2。使用动物模型评估原发性肿瘤及其转移生长过程中CYR61，组织因子和VEGF的重要性。 卵子黑色素瘤的转染细胞系具有不同水平的这三个基因的表达水平，将移植到裸鼠的前腔中，主要眼肿瘤的生长，肝病变的程度，以及两个部位的血管密度。 3。使用免疫细胞化学方法和细胞分馏的组合确定CYR61，组织因子和VEGF的细胞定位。 4。通过RT_PCR，免疫印迹和北印印迹杂交确定其他血管生成因子（Angiopoietin-1）和抑制剂（Angiostatin和therostatin）。