STROMAL INFLUENCES ON BREAST CANCER PROGRESSION

间质对乳腺癌进展的影响

• 批准号: 2109411
• 负责人: SANDRA W MCLESKEY
• 金额: $ 10.73万
• 依托单位: GEORGETOWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-01-01 至 1999-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2109411
• 关键词: MCF7 cell; RNase protection assay; angiogenesis; angiogenesis factor; biomarker; breast neoplasms; fibroblast growth factor; genetic transcription; immunocytochemistry; in situ hybridization; microcirculation; neoplasm /cancer blood supply; neoplasm /cancer pharmacology; neoplastic process; pentosan; polymerase chain reaction; receptor expression; stainings; stromal cells; tamoxifen; transfection; vascular endothelium

Transfection of MCF-7 breast carcinoma cells with fibroblast growth factor 4 (FGF-4), an angiogenic growth factor, produces a dramatic change in tumor phenotype when the cells are injected into the mammary fat pads of ovariectomized nude mice. While the parental cells are estrogen- dependent for tumor growth, poorly invasive, and rarely metastatic, the transfectants produce large, progressively growing tumors which are invasive, frequently metastatic and growth-stimulated by tamoxifen, an estrogen antagonist. This in vivo behavior of the transfectants is in contrast to their in vitro behavior, which is not substantially different from the parental MCF-7 cells (specifically, the transfectants are not growth-stimulated by tamoxifen in vitro). Thus, it would seem likely that at least some portion of the change in in vivo phenotype is due to an interaction between the transfected gene product and stromal components of the tumor. Since the transfected gene product is a known angiogenic growth factor, it seems likely that increased angiogenesis in tumors produced by-the transfected cells is at least partially responsible for the change in in vivo phenotype produced by the transfection. A number of FGF receptor (FGFR) genes have been identified which may each encode multiple receptors via alternative mRNA splicing. Evidence is emerging which attributes specific responses or affinity for specific FGF ligands to specific FGFR isoforms. Specific FGFRs in tumors produced by transfected cells may respond to the FGF-4 ligand and/or be up- or down- regulated. Evidence is also accumulating that endothelial cells from specific tissues differ, and that endothelial cells from tumors are different from those in normal tissues. Cultured endothelial cells are quite plastic in their gene expression and behavior depending on their passage and growth conditions. Therefore, attempts to study tumor angiogenesis in vitro must use appropriate endothelial cells under appropriate conditions. This project seeks to relate changes in the process of angiogenesis in tumors produced by parental and FGF-4 transfected MCF-7 cells to the change in in vivo behavior. We will relate microvessel density and total vascularity to tumor size in tumors produced by parental and transfected cells. We will describe the process of angiogenesis in tumors produced by parental and transfected cells by techniques which identify proliferating or immature endothelial cells in tumor sections obtained at very early and later time points after injection of tumor cells. The project will also seek to characterize specific in situ FGFR expression by endothelial and other cells contained within the tumor. We will isolate endothelial cells from tumors produced by parental or FGF-4 transfected MCF-7 cells as well as endothelial cells from normal mammary fat pads. Growth assays in which quiescent cells are stimulated to invade a 3-dimensional matrix will be done utilizing very early passage tumor-derived and normal endothelial cells to identify differences in behavior between the three types of endothelial cells. Tumor-derived and mammary fat pad endothelial cells will be subjected to differential display PCR in a search for genes which are preferentially expressed in tumors produced by FGF-4 transfected MCF-7 cells. Transcription of such genes might be an important downstream event of FGFR stimulation and the protein products of such genes might be important in determining the phenotype of the tumor.

转染 MCF-7 乳腺癌细胞并促进成纤维细胞生长 因子 4 (FGF-4) 是一种血管生成因子，可产生巨大的变化 当细胞被注射到乳腺脂肪垫时，肿瘤表型 卵巢切除的裸鼠。虽然亲本细胞具有雌激素- 依赖于肿瘤生长，侵袭性差，且很少发生转移， 转染子产生大的、逐渐生长的肿瘤 他莫昔芬具有侵袭性、经常转移性和生长刺激性， 雌激素拮抗剂。转染子的这种体内行为是 与它们的体外行为相比，没有显着差异 来自亲本 MCF-7 细胞（具体来说，转染子不是 体外由他莫昔芬刺激生长）。因此，看起来很可能 体内表型的变化至少有一部分是由于 转染基因产物和基质成分之间的相互作用 肿瘤的。由于转染的基因产物是已知的血管生成 生长因子，似乎可能增加肿瘤的血管生成 由转染细胞产生的至少部分负责 转染产生的体内表型的变化。一个数字 已鉴定出 FGF 受体 (FGFR) 基因，每个基因可能编码 通过选择性 mRNA 剪接形成多个受体。 证据正在出现 其归因于特定 FGF 配体的特定反应或亲和力 特定 FGFR 亚型。肿瘤中产生的特定 FGFR 转染细胞可能会对 FGF-4 配体产生反应和/或上调或下调 受监管。越来越多的证据表明，内皮细胞来自 特定组织有所不同，来自肿瘤的内皮细胞是 与正常组织中的不同。培养的内皮细胞是 他们的基因表达和行为具有相当的可塑性，具体取决于他们的 传代和生长条件。因此，尝试研究肿瘤 体外血管生成必须使用适当的内皮细胞 适当的条件。该项目旨在将变化联系起来 亲本和 FGF-4 产生的肿瘤中血管生成的过程 转染MCF-7细胞使其体内行为发生改变。 我们将 将微血管密度和总血管分布与肿瘤中的肿瘤大小联系起来 由亲代细胞和转染细胞产生。我们将描述该过程 亲代细胞和转染细胞产生的肿瘤中血管生成的影响 识别增殖或未成熟内皮细胞的技术 在术后极早和较晚时间点获得的肿瘤切片 注射肿瘤细胞。该项目还将寻求表征 内皮细胞和其他细胞特异性原位 FGFR 表达 肿瘤内。我们将从产生的肿瘤中分离出内皮细胞 由亲本或 FGF-4 转染的 MCF-7 细胞以及内皮细胞 来自正常乳腺脂肪垫。静止细胞的生长测定 刺激入侵 3 维矩阵将利用非常 早期传代肿瘤来源的内皮细胞和正常内皮细胞以鉴定 三种类型内皮细胞之间的行为差​​异。 肿瘤来源的内皮细胞和乳腺脂肪垫内皮细胞将受到 差异显示PCR以寻找优先的基因 在 FGF-4 转染的 MCF-7 细胞产生的肿瘤中表达。 这些基因的转录可能是一个重要的下游事件 FGFR 刺激和此类基因的蛋白质产物可能是 对于确定肿瘤的表型很重要。