IMMUNOLOGY OF RECOMBINANT MIGRATION INHIBITORY FACTOR

使用来自COS-1细胞的人重组MIF，我们已经证明 这种淋巴因子上调了几个相关的巨噬细胞分子 通过激活(人类白细胞抗原DR、IL-1β、肿瘤坏死因子α和一氧化氮)，激活 巨噬细胞杀死原生动物(细胞内)和肿瘤细胞 (细胞外)，并增强巨噬细胞的聚集和黏附 为了彼此。对于对照组，我们使用了含有非活性物质的上清液 COS-1细胞转导突变的MIF-cDNAs的“stopMIF” 一个终止密码子。人RMIF在体外作用于小鼠细胞，当给予 用小鼠体内的抗原，它会引起抗原依赖的淋巴 结节T细胞增殖反应与其诱导的细胞增殖反应一样强 完全弗洛伊德佐剂。最后，初步研究表明， 完全弗氏佐剂在体内的佐剂作用完全 被RMIF抗体废除。这项建议的具体目标是： 1.制备和纯化具有生物活性的重组MIF 合理的数量。 2.确定RMIF的佐剂作用机制。这将是 包括RMIF对抗原加工、整合素 活性和表达，以及与其他细胞因子的相互作用。 3.确定最近发现的机制，当给予 小鼠，RMIF抗体消除Complete的佐剂作用 弗洛伊德佐剂。 4.用纯化标记的RMIF直接获得RMIF受体 受体表达克隆或通过免疫化学方法。

Using human recombinant MIF from transfected COS-1 cells, we have shown that this lymphokine upregulates several macrophage molecules associated with activation (HLA-DR, IL-1beta, TNFalpha, and nitric oxide), activates macrophage to kill protozoa (intracellularly) and tumor cells (extracellularly), and enhances the clumping and adhesion of macrophages to each other. For controls, we used supernatants containing non-active "stopMIF" from COS-1 cells transfected with a mutated MIF-cDNA containing a stop codon. Human rMIF acts on murine cells in vitro, and when given with an antigen to mice in vivo, it elicits an antigen-dependent lymph node T cell proliferative response in as great as that elicited by complete Freund's adjuvant. Finally, preliminary studies show that the in vivo adjuvant effect of complete Freund's adjuvant is totally abrogated by antibodies to rMIF. The specific aims of this proposal are: 1. To produce and purify biologically active recombinant MIF in reasonable quantity. 2. To determine the mechanism of the adjuvant action of rMIF. This will include studies on the effect of rMIF on antigen processing, on integrin activity and expression, and on its interaction with other cytokines. 3. To ascertain the mechanism for the recent finding that, when given to mice, antibodies to rMIF abrogate the adjuvant action of complete Freund's adjuvant. 4. To use purified labelled rMIF to obtain the rMIF receptor, by direct receptor expression cloning or by immunochemical means.