PROLIFERATION AND FUNCTION IN HUMAN ENDOTHELIAL CELLS

该研究计划的目的是研究生长和功能 从不同血管中分离出的培养人内皮细胞 (EC) 网站。 血管组织将从脑死亡、心跳中获得 尸体、大血管和毛细血管EC将被分离。 在最初的 实验优先考虑人髂静脉内皮细胞 血统源自年轻、健康的个体。 将实现以下目标 追求： 1. 成人血管 EC 培养物的培养和表征 关于 EC 标记功能、核型和培养寿命。 2. 将研究人类 EC 体外增殖的影响 肝素、脑源性内皮细胞之间的协同相互作用 有丝分裂原、纤连蛋白和胶原蛋白。 将采用以下方法 使用： a) 组分的纯化和分馏； b) EC 的定义 对来自a)的组件的响应； c) 下组件的交互 无细胞条件； d) 血清对效果和相互作用的影响 的组件。 3. 人体EC功能，血管紧张素I转换酶的表达 （ACE）在体外，将被研究，因为它受到合作的影响 肝素、脑源性内皮细胞有丝分裂原、 纤连蛋白和胶原蛋白。 将使用以下方法：a) EC 对培养物成分的反应（根据 ACE 活性）的定义 目标#2 中纯化并分级； b) 特定多克隆的开发 或单克隆抗体来定量 ACE 的量、合成和 退化； c) 克隆和非克隆培养物中 ACE 表达的分析 在单个继代生长周期内。 4. 在实现上述目标取得足够进展后， 血管部位、供体年龄和性别以及体外可能的影响 培养人类 EC 的功能和增殖 调查了。

The objective of this research program is to study the growth and function of cultured human endothelial cells (EC) isolated from diverse vascular sites. Vascular tissue will be obtained from brain-dead, heart-beating cadavers, and large vessel and capillary EC will be isolated. In initial experiments priority will be given to a human iliac vein endothelial cell line derived from a young, healthy individual. The following goals will be pursued: 1. Cultures of adult human vascular EC will be cultivated and characterized with respect to EC marker functions, karyotypes, and culture lifespans. 2. Human EC proliferation in vitro will be investigated as effected by cooperative interactions between heparin, brain-derived endothelial mitogen, fibronectin, and collagen. The following approaches will be used: a) Purification and fractionation of components; b) Definition of EC response to components from a); c) Interaction of components under cell-free conditions; d) Influence of serum on the effects and interaction of components. 3. A human EC function, the expression of angiotensin I-converting enzyme (ACE) in vitro, will be investigated as it is affected by cooperative interactions between heparin, brain-derived endothelial mitogen, fibronectin, and collagen. The following approaches will be used: a) Definition of EC response (in terms of ACE activity) to culture components purified and fractionated in Aim #2; b) Development of specific polyclonal or monoclonal antibodies to quantitate ACE amounts, synthesis and degradation; c) Analyses of ACE expression in cloned and uncloned cultures within a single subcultivation growth cycle. 4. After sufficient progress has been made in achieving the above goals, possible effects of vascular site, donor age and sex, and in vitro cultivation on function and proliferation in culture human EC will be investigated.