PROLACTIN AN ANTIANGIOGENIC FACTOR

催乳素是一种抗血管生成因子

• 批准号: 3247711
• 负责人: RICHARD Ira WEINER
• 金额: $ 16.22万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-06-11 至 1996-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3247711
• 关键词: 3T3 cells; B lymphocyte; SDS polyacrylamide gel electrophoresis; angiogenesis; angiogenesis factor; athymic mouse; chick embryo; chorioallantoic membrane; epidermal growth factor; fibroblast growth factor; fusion gene; genetic library; genetically modified animals; growth factor receptors; growth inhibitors; hormone regulation /control mechanism; laboratory mouse; neoplastic cell; neoplastic growth; polymerase chain reaction; prolactin; protease inhibitor; recombinant proteins; transfection /expression vector; vascular endothelium

An essential component of the multistep process of tumorigenesis is the development of a new blood supply, angiogenesis. A strategy in inhibition of tumor formation and growth is to inhibit neovascularization. We have shown that the 16 kd n terminal fragment of prolactin (16K PRL) is a specific angiolytic factor in vitro. Both rat and recombinant human 16K PRL inhibits basal and bFGF or VEGF stimulated growth of bovine and human vascular endothelial cells. 16K PRL also inhibits organization of capillary endothelial cells into capillary like structures when cultured in collagen gels. Finally 16K PRL inhibits development of the microvasculature in the chick chorioallantoic membrane. We will determine if 16K PRL inhibits activation of proteases in capillary endothelial cells by BFGF. In the chick chorioallantoic membrane we will determine if 16K PRL also inhibits the angiogenic actions of bFGF and VEGF. We will determine if 16K PRL inhibits tumor growth and associated angiogenesis in two model systems. NIH 3T3 cells transformed by transfections with a signal peptide FGF expression vector or the HCT 16 human colon carcinoma cells will be transplanted subcutaneously into nude athymic mice. 16K PRL will be administered systemically or intratumorally and the growth and vascularization of tumor histologically evaluated. In another model lines of transgenic mice will be developed in which 16K hPRL expression is directed to beta cells of the pancreas. This will be accomplished by constructing a chimeric gene consisting of the insulin promoter/enhancer elements (In) upstream from the coding region for 16K hPRL. These mice will then be crossed with transgenic mice expressing SV40 T antigen in the beta cells of the pancreas (In-Tag). The In-Tag mice are well characterized and develop time dependent tumors of the pancreas. Frequency, rate of growth and vascularization of tumors will be evaluated. The angiolytic action of 16K PRL is mediated via a novel receptor which is not the prolactin or FGF receptors. We will attempt to clone the receptor by two approaches. (1) Expression cloning in COS cells. A bovine brain capillary endothelial cell (BBE) cDNA library in pcDNA1 vector will be expressed in COS cells and screened by autoradiographic analysis of I-125 16K PRL binding. (2) Homology to GH/PRL/Interleukin receptor family: PCR fragments obtained with primers to conserved region of the receptor family will be used to screen cDNA libraries of BBE or human umbilical vein endothelial cells.

肿瘤发生的多步骤过程中的一个重要组成部分是 开发一种新的血液供应，血管生成。在中国的战略 抑制肿瘤的形成和生长就是抑制 新生血管。我们已经证明了16kdn末端片段 催乳素(16K-PRL)在体外是一种特异的血管溶解因子。两者都有 大鼠和重组人16K催乳素抑制碱性成纤维细胞生长因子和血管内皮生长因子 刺激牛和人血管内皮细胞的生长。16K 催乳素还抑制毛细血管内皮细胞的组织 在胶原胶中培养时，可形成毛细血管样结构。最后是16K 催乳素抑制雏鸡微血管的发育 绒毛尿囊膜。 我们将确定16K PRL是否抑制蛋白水解酶的激活 碱性成纤维细胞生长因子诱导的毛细血管内皮细胞。在鸡绒毛膜尿囊症中 我们将确定16K PRL是否也抑制血管生成 碱性成纤维细胞生长因子和血管内皮生长因子的作用。 我们将确定16K催乳素是否抑制肿瘤生长并与 两个模型系统中的血管生成。转化的NIH3T3细胞 信号肽成纤维细胞生长因子表达载体或HCT的转染法 16个人结肠癌细胞将被移植到皮下 裸体裸鼠。16K PRL将系统管理或 肿瘤内与肿瘤生长和血运的关系 组织学评估。在另一种转基因小鼠模型系中 将开发其中16K hPRL表达定向到测试版 胰腺细胞。这将通过构造一个 由胰岛素启动子/增强子元件(In)组成的嵌合基因 16K hPRL编码区的上游。然后这些老鼠就会被 与表达SV40T抗原的转基因小鼠杂交 胰腺细胞(标记内)。标签内的小鼠有很好的特征 并发展成时间依赖性的胰腺肿瘤。频率、频率、频率 将对肿瘤的生长和血管形成进行评估。 16K PRL的血管溶解作用是通过一种新的受体介导的，该受体 不是催乳素或成纤维细胞生长因子受体。我们将尝试克隆 受体通过两种途径。(1)COS细胞表达克隆。一个 PcDNA1中的牛脑毛细血管内皮细胞(BBE)cDNA文库 载体将在COS细胞中表达，并通过放射自显影筛选 I-125 16K催乳素结合分析。(2)生长激素/催乳素/白介素同源 受体家族：用聚合酶链式反应扩增片段 该受体家族的区域将被用来筛选 BBE或人脐静脉内皮细胞。