IMMUNOLOGY OF RECOMBINANT MIGRATION INHIBITORY FACTOR

使用来自转染的COS-1细胞的人重组MIF，我们已经显示， 这种淋巴因子上调了几种巨噬细胞分子， 激活（HLA-DR，IL-1 β，TNF 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），并增强巨噬细胞的凝集和粘附 彼此之间。 对于对照，我们使用含有非活性 “stopMIF”来自转染了突变的MIF-cDNA的COS-1细胞， 终止密码子。 人rMIF在体外作用于鼠细胞， 与小鼠体内的抗原结合， 淋巴结T细胞增殖反应与 完全弗氏佐剂。 最后，初步研究表明， 完全弗氏佐剂体内佐剂作用完全 被rMIF抗体消除。 这项建议的具体目标是： 1.目的在大肠杆菌中生产和纯化具有生物学活性的重组MIF。 合理的数量。 2.探讨rMIF的佐剂作用机制。 这将 包括研究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.为了确定最近发现的机制，当给予 在小鼠中，rMIF的抗体消除了完全免疫抑制剂的佐剂作用。 弗氏佐剂。 4.利用纯化的标记rMIF，通过直接测序法获得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.