MAINTENANCE AND REPAIR OF VASCULAR ENDOTHELIUM

我们的具体目标是分析血管内皮细胞转染术 携带碱性成纤维细胞生长因子基因表达载体的内皮细胞可导致 血管内皮细胞的自主增殖及其机制 转化为肿瘤细胞。我们将首先分析 碱性成纤维细胞生长因子是否会在细胞内起作用(作为大多数癌基因 产品)或需要由细胞释放，以便 与暴露的细胞表面受体相互作用。我们还将分析 它的细胞定位，以确定它是否可以 最终变得与核、胞浆或胞浆有关 膜组分。因为细胞转化可以直接 癌基因表达的结果，我们将在转基因后进行分析 血管内皮细胞是否组成性表达c 可观察到fos、c-myc和c-ras癌基因。无论是 碱性成纤维细胞生长因子基因产物可从细胞中联合释放。 含有ECM成分，如硫酸乙酰肝素，对于这些成分， 有丝分裂原应具有较高的亲和力，将予以评估。其效果 TGFb和肿瘤坏死因子对亲本细胞和转染组细胞生长的影响 在软琼脂中，以及PA的表达将被分析，以便 确定它们是否会产生类似于 当细胞生长在固体基质上时报告。单克隆 将获得抗成纤维细胞生长因子受体的抗体并用于 对成纤维细胞生长因子受体敏感的放射免疫分析的发展，以及 对其表达的研究。它们也将被用来分离成纤维细胞生长因子 用免疫亲和层析法测定受体。成纤维细胞生长因子受体 然后将在结构上进行表征，信息 Gain将用于开发特定的寡核苷酸探针， 为克隆成纤维细胞生长因子受体基因奠定了基础。

Our specific aims are to analyze whether transfection of vascular endothelial cells with bFGF cDNA expression vector could lead to autonomous proliferation of vascular endothelial cells and their transformation into neoplastic cells. We will first analyze whether bFGF would act intracellularly (as most oncogene products do) or need to be released by the cells in order to interact with exposed cell surface receptors. We will also analyze its cellular localization, in order to determine whether it could ultimately become associated with the nuclear, cytosol, or plasma membrane fraction. Since cell transformation can be the direct result of oncogene expression, we will analyze in transfected vascular endothelial cells whether constitutive expression of c fos, c myc and c ras oncogenes can be observed. Whether the bFGF gene product could be released from the cells in association with ECM components such as heparan sulfate GAG, for which the mitogen should have a high affinity, will be assessed. The effect of TGFB and TNF on the growth of parental and transfected cells in soft agar, as well as PA expression, will be analyzed in order to determine whether they could have effects similar to those reported when cells are grown on solid substrates. Monoclonal antibodies against the FGF receptor will be obtained and used for the development of sensitive RIA for the FGF receptor, and studies of its expression. They will also be used to isolate FGF receptors by immunoaffinity chromatography. The FGF receptor will then be characterized structurally, and the information gained will be used to develop sepecific oligonucleotide probes, which will be used for the cloning of 2 FGF receptor cDNA.