Study of regulatory mechanism of inflammatory response by purinergic signaling

我们研究了是否嘌呤信号系统可以改变炎症条件。用IFN-γ处理人脐静脉内皮细胞（HUVEC）可增强ATP诱导的细胞内Ca^<2+>浓度升高。IFN-γ通过上调P2X_4受体的表达和下调三磷酸核苷二磷酸水解酶1（NTPDase 1）的表达而发挥作用。IFN-γ对HUVEC嘌呤能信号系统的作用是通过Jak-STAT途径介导的，并可被IL-1β和TNF-α等其他炎性细胞因子协同增强。ATP刺激静止的HUVEC可迅速诱导环氧合酶（考克斯）2 mRNA、蛋白和酶活性的表达。ATP的作用是通过离子型受体介导的，可能是P2X_4受体，其机制涉及p38 MAP激酶介导的COX 2 mRNA稳定。在IFN-γ处理的HUVEC中，ATP诱导的COX 2诱导增强。与对HUVEC的作用不同，IFN-γ以时间和浓度依赖性方式增加小鼠巨噬细胞J774细胞的ATP水解，并伴随NTPDase 1 mRNA的显著增加。放线菌酮、Jak抑制剂和蛋白酪氨酸磷酸酶抑制剂orthovanadate可抑制IFN-γ对NTPD酶1的上调作用，而酪氨酸激酶抑制剂（除莠霉素A、PP 2和genistein）和p42/44 MAP激酶抑制剂可增强IFN-γ对NTPD酶1的上调作用。这些结果表明IFN-γ以酪氨酸磷酸酶依赖的方式引起NTPDase 1的上调。IFN-γ对巨噬细胞和内皮细胞NTPDase 1表达的差异性调节可能参与了血管炎症条件下巨噬细胞与内皮细胞的相互作用。

We investigated whether the purinergic signaling system could be altered with inflammatory condition. Treatment of human umbilical vein endothelial cells(HUVEC) with IFN-γ enhanced ATP-induced increase in intracellular Ca^<2+> concentration. This effect of IFN-γ was resulted from up-regulation of P2X_4 receptor expression and down-regulation of nucleoside triphosphate diphosphohydrolase 1 (NTPDase1). Effects of IFN-γ on the purinergic signaling system in HUVEC were mediated by Jak-STAT pathway and further enhanced synergistically by other inflammatory cytokines, such as IL-1β and TNF-α. Stimulation of quiescent HUVEC by ATP caused a rapid induction of cyclooxyganase(COX) 2 mRNA, protein and enzyme activity. The effect of ATP was mediated by ionotropic receptors, probably P2X_4 receptors through a mechanism involving p38MAP kinase-mediated COX2 mRNA stabilization. The ATP-induced COX2 induction was augmented in IFN-γ-treated HUVEC. Unlike the effects on HUVEC, IFN-γ increased ATP hydrolysis in murine macrophage J774 cells in a time- and concentration-dependent manner, accompanied by marked increase in NTPDase1 mRNA. Up-regulation of NTPDase1 by IFN-γ was inhibited by cycloheximide, Jak inhibitor and orthovanadate, a protein tyrosine phosphatase inhibitor, but enhanced by tyrosine kinase inhibitors (herbimycin A, PP2 and genistein) and p42/44 MAP kinase inhibitors. These results suggest that IFN-γ causes the up-regulation of NTPDase1 in a tyrosine phosphatase-dependent manner. The differential regulation of NTPDase1 expression in macrophages and EC by IFN-γ may contribute to the macrophage-endothelium interaction under the vascular inflammation.