LIVER REGENERATION AS A MODEL FOR ANGIOGENESIS

说明：本修订提案的总体目标是确定 调节正常新生血管形成的信号转导途径 在再生的肝脏中。 大鼠肝再生模型 70%部分肝切除术（PHx）后， 系统，通过它来剖析受控血管生成的现象。 PHx后，肝细胞在24小时经历峰值有丝分裂，而 肝窦内皮细胞（SEC）在96小时达到有丝分裂峰值。 肝细胞分裂在没有SEC分裂的情况下，存在大量的 肝脏内非血管化肝岛的百分比。 的 SEC分裂，他们随后迁移到这些岛屿， 肝内开放血管的形成由生长介导 因子和细胞外基质信号尚未研究 并且与阐明控制组织的机制有关 血管化。 细胞外特异性的 基质，以及内皮有丝分裂原和运动原，包括EGF， TGF-α、TGF-β 1、HGF、aFGF、bFGF和VEGF及其磷酸化 在血运重建过程中检查其受体的状态 过程所有这些因素都在PHx后的肝脏中发现 并且可以介导与血管生成有关的特定步骤。 到 研究对启动和进展至关重要的信号 在肝再生的时间框架内血管生成， 将使用体内膜分离技术。 SEC膜 肝脏将通过血液灌注而非共价衍生化 在PHx后的特定时间，将容器与阳离子胶体混合。 这 该技术允许SEC膜的快速、高产率分离 同时保持蛋白质组分的方向， 膜在衍生化的确切时间。 结果导致 作用于细胞膜和细胞溶质的信号转导途径 SEC的分数可以在PHx之后的任何给定时间确定。 平行体外研究将允许详细检查 有丝分裂和运动信号的机制 SEC通过生长因子和细胞外基质的转导 孤立的条件。

DESCRIPTION: The overall aim of this revised proposal is to determine the signal transduction pathways that regulate normal neovascularization in the regenerating liver. The model of liver regeneration in the rat following 70% partial hepatectomy (PHx) provides a hitherto unexploited system by which to dissect the phenomenon of controlled angiogenesis. Following PHx, hepatocytes undergo peak mitoses at 24 hr, while sinusoidal endothelial cells (SEC) achiev peak mitoses at 96 hr. After hepatocyte division in the absence of SEC division, there exist a large percentage of non-vascularized hepatic islands within the liver. The SEC division, their subsequent migration into these islands, and formation of patent vessels within the liver is mediated by growth factor and extracellular matrix signals that have not been investigated and are relevant to elucidating mechanisms that control tissue vascularization in general. The presence of specific extracellular matrices, as well as endothelial mitogens and motogens including EGF, TGF-alpha, TGF-beta1, HGF, aFGF, bFGF and VEGF and the phosphorylation state of their receptors, will be examined during the revascularization process. All of these factors have been found in the liver following PHx and may mediate specific steps pertinent to angiogenesis. To investigate the signals that are critical for initiation and progression of angiogenesis within the time frame of liver regeneration, a novel in vivo membrane isolation technique will be utilized. SEC membranes of the liver will be non-covalently derivatized by perfusion of blood vessels with cationic colloids at specific times following PHx. This technology allows for the rapid, high yield isolation of SEC membranes from liver while maintaining the orientation of component proteins of the membrane at the exact time of derivitization. As a result, the signal transduction pathways acting at the membrane as well as cytosolic fractions of the SEC can be ascertained at any given time following PHx. The parallel in vitro studies will allow for the detailed examination of mechanisms involved in the mitogenic and motogenic signal transduction of SEC by growth factors and extracellular matrices under isolated conditions.