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.

肿瘤发生多步过程的重要组成部分是 开发新的血液供应，血管生成。 一种策略 抑制肿瘤形成和生长是抑制 新血管形成。 我们已经证明了16 kD N末端片段 催乳素（16K PRL）的体外是特定的血管细胞因子。 两个都 大鼠和重组人16K PRL抑制基础和BFGF或VEGF 牛和人血管内皮细胞的刺激生长。 16K PRL还抑制毛细管内皮细胞的组织 在胶原蛋白凝胶中培养时，毛细管类似结构。 终于16K PRL抑制雏鸡微脉管系统的发展 绒毛膜膜。 我们将确定16K PRL是否抑制蛋白酶的激活 BFGF毛细血管内皮细胞。 在小鸡绒毛膜中 膜我们将确定16K PRL是否也抑制了血管生成 BFGF和VEGF的动作。 我们将确定16K PRL是否抑制肿瘤生长和相关 两个模型系统中的血管生成。 NIH 3T3细胞通过 用信号肽FGF表达载体或HCT转染 16人类结肠癌细胞将皮下移植到 裸露小鼠。 16K PRL将被系统地管理或 肿瘤内以及肿瘤的生长和血管形成 组织学评估。 在另一个模型的转基因小鼠中 将开发16k HPRL表达式定向到beta 胰腺细胞。 这将通过构建 由胰岛素启动子/增强子元素组成的嵌合基因（IN） 从编码区域上游的16K HPRL。 这些老鼠将是 与表达SV40 T抗原的转基因小鼠交叉在β 胰腺的细胞（IN-TAG）。 标签小鼠的特征很好 并发展胰腺的时间依赖性肿瘤。 频率，速率 将评估肿瘤的生长和血管化。 16K PRL的血管元动作是通过一种新型受体介导的 不是催乳素或FGF受体。 我们将尝试克隆 通过两种方法的受体。 （1）在cos细胞中表达克隆。一个 PCDNA1中的牛脑毛细管内皮细胞（BBE）cDNA库 向量将在COS细胞中表达，并通过放射自显影筛选 I-125 16K PRL结合的分析。 （2）与GH/PRL/白介素同源 受体家族：用底漆获得的PCR片段 受体家族的区域将用于筛选cDNA库 BBE或人脐静脉内皮细胞。