Identification of FX-activating molecules of convulsion-inducing lipoprotein purified from egg granulomas in Schistosoma mansoni-infected mice and of its inducing mechanism

通过生化研究，鉴定了从曼氏血吸虫感染小鼠肝卵肉芽肿中纯化的惊厥诱导脂蛋白（CILIP）的因子-X（FX）激活分子、巨噬细胞和树突状细胞诱导CILIP的机制以及参与CILIP诱导的信号转导通路。 1.体外分析表明巨噬细胞和树突状细胞可以产生CILIP（≒FX激活） PCA) 当用 Toll 样受体的几种配体（如 LPS、LTA、肽聚糖、poly IC 和 CpGDNA）刺激时，用 SWAP 抗原致敏的脾 DX5 NK 细胞和/或 CD4 T 细胞，或用细胞因子（如 IFNγ 和 GM-CSF）刺激。2. 通过用 SWAP 致敏的淋巴细胞刺激来诱导巨噬细胞 FX 激活 PCA 需要从头合成 RNA 和蛋白质，并且PTK 和 PKC 激活，但不激活 MAPK、钙调蛋白或细胞内 Ca 浓度增加。此外，细胞内cAMP的增加可下调巨噬细胞FX激活PCA的诱导。3.从BALB/c小鼠和Wister大鼠中制备了两种抗CILIP单克隆抗体。大鼠的MoAb似乎对天然CILIP具有高度特异性和敏感性，并且抑制CILIP的剂量依赖性FX激活活性，但不与用去垢剂处理的变性CILIP发生反应。开发了使用两种针对CILIP的MoAb的夹心ELISA系统，并证实了其在定量测定CILIP时的有效性。4.尽管已经使用抗组织因子的MoAb和针对各种凝血因子的特异性抗体进行了各种生化分析来鉴定CILIP的EX激活分子，但尚未获得确凿的证据。

Biochemical studies has been performed to identify the factor-X(FX)-activating molecules of convulsion-inducing lipoprotein (CILIP) purified from hepatic egg granulomas in Schistosoma mansoni-infected mice, the inducing mechanism of CILIP by macrophages and dendritic cells, and the signal transduction pathway involved in CILIP induction.1.In vitro analysis demonstrated that macrophages and dendritic cells could produce CILIP(≒FX-activating PCA) when stimulated with several ligands for Toll-like receptors such as LPS, LTA, peptidoglycan, poly IC and CpGDNA, with splenic DX5+ NK cells and/or CD4+ T cell sensitized with SWAP antigen, or with cytokines such as IFNγ and GM-CSF.2.The induction of macrophage FX-activating PCA by stimulation with SWAP sensitized lymphocytes required de novo RNA and protein synthesis and activation of PTK and PKC, but not MAPK activation, calmodulin or increase of intracellular Ca++ concentration. Moreover, the induction of macrophage FX-activating PCAwas down-regulated by increment of intracellular cAMP.3.Two kinds of monoclonal antibodies against CILIP were prepared from BALB/c mice and Wister rats. Rat's MoAb appeared to be highly specific and sensitive to native CILIP, and inhibited dose-dependently FX-activating activity of CILIP, but did not react with denatured CILIP treated with detergents. Sandwich ELISA system using two kinds of MoAbs against CILIP was developed and confirmed its usefulness when applied for quantitative determination of CILIP.4.Although various biochemical analysis using MoAb against tissue factor and specific antibodies against various coagulation factors have been conducted to identify the EX-activating molecules of CILIP, conclusive evidence has not been obtained yet.