Histological, immunocytochemical and biochemical analyses on nature and releasing mechanism of Weibel-Palade body.

众所周知，Webel-Palade小体(WP)中含有von Willebrand因子，在血管损伤后血小板与内皮的黏附中起重要作用。随着内皮衍生松弛因子(EDRF)和内皮衍生收缩因子(EDCF)概念的提出，内皮细胞在血管运动中的作用越来越受到重视，因此高尔基复合体中的WP不仅含有von Willebrand因子，而且还含有其他一些多肽。在组胺诱导的收缩过程中，超微结构的变化是细胞内空泡的数量和大小的增加，内皮细胞间裂隙的扩张，以及通过辣根过氧化物酶标记实验确定的内皮通透性的增加。基于这些原因，我们为本项目进行了以下四个实验。1.猪冠状动脉内皮细胞在含终浓度为100uci/ml的蛋氨酸的MEM中培养16小时，并用2D-PAGE分析内皮细胞释放的放射性多肽。2.上述动脉段的培养和内皮源性多肽的分析。3.上述动脉段的培养加10-3M组胺。4.加入组胺释放剂48/80、钙离子载体(A23187)和聚甲基丙烯酸甲酯(PMA)，培养冠状动脉节段，分析WP在冠脉收缩过程中释放的多肽(120K)。这些多肽是否参与血管收缩正在调查中。

It is well known that Weibel-Palade bodies (WP) contain von Willebrand factor and have an important role in the adhesion of blood platelets to the endothelium after vascular injuries.Since the concept as to endothelial derived relaxing factor (EDRF) and endothelial derived constricting factor (EDCF) has progressed, a role of endothelial cells in vasomotion becomes more and more notified and, therefore WP derived from Golgi complex is supposed to contain not only von Willebrand factor but also some other polypeptides.In the previous study by ours, the in vitro swine coronary artery remarkably contracts by addition of 10^<-3>M histamine. Ultrastructural alterations during the histamine-induced contraction are the increase in number and size of the intraendocellular vacuoles, the dilatation of interendothelial clefts, and the enhancement of the endothelial permeability as determined by marker experiments using horseradish peroxidase.On these grounds, we performed the following four experiments for the present project. 1. Culture of endothelial cells of the swine coronary artery in MEM containing [^<35>S] methionine at a final concentration of 100uci/ml for 16 hr and analysis of radioactive polypeptides released from the endothelial cells by 2D-PAGE. 2. Culture of the arterial segments as described above and analysis of endothelial-derived polypeptides. 3. Culture of the arterial segments as described above with an addition of 10-3m histamine. 4. Culture of the arterial segments with and without an addition of histamine releasers such as compound 48/80, calcium ionophore (A23187) and PMA.From these experiments, we analyze polypeptides (120K) released from WP during the coronary artery contraction. Whether these polypeptides are involved in the vasocontraction is under investigation.