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

从曼氏血吸虫感染的小鼠肝肉芽肿中提纯的惊厥诱导脂蛋白X因子(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诱导的信号转导途径。1.体外分析表明，巨噬细胞和树突状细胞在几种Toll样受体的配体如脂多糖、LTA、肽聚糖、多聚IC和CpGDNA的刺激下，与脾≒NK细胞和/或经抗原交换的CD4T细胞刺激，可以产生Toll样受体的CILIP。SWAP致敏的淋巴细胞诱导巨噬细胞FX激活Pca需要从头开始的γ和蛋白质合成及激活PTK和PKC，而不需要MAPK的激活、钙调素或细胞内钙离子浓度的增加。此外，细胞内cAMP的增加下调了巨噬细胞FX激活PCA的诱导。大鼠的单抗对天然CILIP表现出高度的特异性和敏感性，并呈剂量依赖性地抑制CILIP的FX激活活性，但不与经洗涤剂处理的变性CILIP发生反应。建立了两种抗CILIP单抗的夹心ELISA法，并将其应用于CILIP的定量检测。4.尽管利用抗组织因子的单抗和抗各种凝血因子的特异性抗体进行了各种生化分析，以确定CILIP的前激活分子，但尚未得到确凿的证据。

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.