Reinders J H, de Groot P G, Sixma J J, van Mourik J A
Central Laboratory of The Netherlands Red Cross Blood Transfusion Service, Amsterdam.
Haemostasis. 1988;18(4-6):246-61. doi: 10.1159/000215811.
Endothelial cells synthesize and store von Willebrand factor. We have studied the storage and secretion of von Willebrand factor in cultured human umbilical vein endothelial cells. In particular, we were interested in the nature of the storage compartment and the effects of perturbation on the storage and secretion processes. The storage compartment for von Willebrand factor was isolated from homogenates of endothelial cells. By an immunostaining technique the isolated vesicles stained for von Willebrand factor. The staining pattern was similar to that of Weibel-Palade bodies in intact endothelial cells. We concluded that the storage compartment containing von Willebrand factor is identical to the Weibel-Palade body. The von Willebrand factor of the isolated storage vesicles is predominantly constructed of polypeptide chains with a Mr of 220 kD. On the other hand, von Willebrand factor continuously secreted by endothelial cells is constructed of both a 220 kD and a larger precursor (apparent Mr of 275 kD) subunit. The storage vesicles contain von Willebrand factor that supports ristocetin-induced platelet aggregation. Thus, endothelial cells store fully processed, biologically active von Willebrand factor within Weibel-Palade bodies. Short-term (less than 1 h) treatment of endothelial cells with the perturbing phorbol ester 4 beta-phorbol-12-myristate-13-acetate (PMA) results in release of cellular stored von Willebrand factor. 24-48 h after exposure to PMA the endothelial cell distribution of von Willebrand factor is changed distinctly. While the contents of the von Willebrand factor storage sites in the cells are gradually restored within 48 h, enhanced amounts of von Willebrand factor are secreted into the medium. The number as well as the size of von Willebrand factor storage granules in the endothelial cells increase after exposure to phorbol ester, as determined by immunofluorescence microscopy. Phorbol ester treated cells release stored von Willebrand factor 48 h after they have been stimulated. PMA decreases the von Willebrand factor contents of the extracellular matrix; the deposition of von Willebrand factor in the subendothelium is blocked by PMA, whereas the degradation of matrix von Willebrand factor is not affected. Thus, perturbation of endothelial cells changes the cellular distribution of von Willebrand factor.
内皮细胞合成并储存血管性血友病因子。我们研究了培养的人脐静脉内皮细胞中血管性血友病因子的储存和分泌情况。特别地，我们关注储存区室的性质以及干扰对储存和分泌过程的影响。从内皮细胞匀浆中分离出血管性血友病因子的储存区室。通过免疫染色技术，分离出的囊泡被染成血管性血友病因子阳性。染色模式与完整内皮细胞中的魏尔-帕拉德小体相似。我们得出结论，含有血管性血友病因子的储存区室与魏尔-帕拉德小体相同。分离出的储存囊泡中的血管性血友病因子主要由分子量为220kD的多肽链构成。另一方面，内皮细胞持续分泌的血管性血友病因子由220kD和更大的前体（表观分子量为275kD）亚基构成。储存囊泡含有支持瑞斯托霉素诱导的血小板聚集的血管性血友病因子。因此，内皮细胞在魏尔-帕拉德小体内储存完全加工的、具有生物活性的血管性血友病因子。用干扰性佛波酯4β-佛波醇-12-肉豆蔻酸酯-13-乙酸酯（PMA）对内皮细胞进行短期（少于1小时）处理，会导致细胞内储存的血管性血友病因子释放。暴露于PMA后24 - 48小时，血管性血友病因子的内皮细胞分布明显改变。虽然细胞内血管性血友病因子储存位点的内容物在48小时内逐渐恢复，但分泌到培养基中的血管性血友病因子量增加。通过免疫荧光显微镜检查确定，暴露于佛波酯后，内皮细胞中血管性血友病因子储存颗粒的数量和大小增加。经佛波酯处理的细胞在受到刺激后48小时释放储存的血管性血友病因子。PMA降低细胞外基质中血管性血友病因子的含量；PMA阻断血管性血友病因子在血管内皮下层的沉积，而基质血管性血友病因子的降解不受影响。因此，内皮细胞的干扰改变了血管性血友病因子的细胞分布。
