Abstract

Sphingosine-1-phosphate (S1P) is a low-molecular-weight phospholipid derivative released by activated platelets. S1P transduces signals through a family of G protein-coupled receptors to modulate various physiological behaviors of endothelial cells. Platelet/endothelial cell adhesion molecule-1 (PECAM-1; CD31) is a 130-kDa protein expressed on the surfaces of leukocytes, platelets, and endothelial cells. Upon PECAM-1 activation, its cytoplasmic tyrosine residues become phosphorylated and bind with SH2 domain-containing proteins, thus leading to the downstream functions mediated by PECAM-1. In the present study, we found that S1P induced PECAM-1 tyrosine phosphorylation and SHP-2 association in bovine aortic endothelial cells (BAECs) by immunoprecipitation and western blotting. The pretreatment of BAECs with a series of chemical inhibitors to determine the signaling pathway showed that the PECAM-1 phosphorylation was inhibited by PP2, indicating the participation of Src family kinases. These results demonstrated that S1P induced PECAM-1 tyrosine phosphorylation in BAECs through mediation of Src family kinases, and this may regulate the physiological behaviors of endothelial cells.

中文摘要

Sphingosine 1-phosphate (S1P)是一種低分子量的多功能性水解磷酸脂，主要經由被活化的血小板釋放到血液中。S1P 和細胞表面特殊的G 型蛋白耦合受器結合後，會引發下游一連串的細胞訊息傳遞及並影響細胞的基因表現與生理活動，包括：細胞增生、移行、傷口癒合，血管新生及免疫反應的過程。第一型血小板/內皮細胞附著因子(Platelet/endothelial cell adhesion molecule-1; PECAM-1)是一種大量表現在血小板及內皮細胞上的細胞黏著因子。其膜內結構上的酪氨酸在磷酸化後會和具有SH2 結構之蛋白質結合並活化多種下游之細胞生理活動。在本實驗中，我們使用免疫沉澱與西方墨點法證明了S1P 的處理會引發牛主動脈內皮細胞上PECAM-1 酪氨酸的磷酸化以及後續SHP-2的結合。使用化學抑制劑所進行的前處理顯示了此PECAM-1 磷酸化的現象會被PP2 所抑制，顯示Src family kinases 在S1P 所引發的PECAM-1 酪氨酸磷酸化的過程中扮演了仲介者的角色。釐清此機制的細節將有助於未來在內皮細胞生理功能調控過程的研究。

Keyword: S1P, PECAM-1, SHP-2, Src family kinases. S1P、第一型血小板/內皮細胞附著因子、SHP-2、Src family kinases。