NOVEL PROINFLAMMATORY CYTOKINE AND ENDOTHELIUM

Regulation of endothelial cell (EC) coagulant, barrier, adhesive and proliferative functions is central to inflammation, and the host response to tumors. We have identified, cloned and characterized a novel polypeptide, designated Endothelial-Monocyte Activating Polypeptide II (EMAPII), which modulates these EC functions, and is phlogogenic and angiogenic. EMAPII is synthesized by activated murine mononuclear phagocytes (MPs) and constitutively by murine Meth A sarcoma (Meth A) cells, and is secreted as an about 18-20 kDA single chain molecule. EMAPII also activates both MPs, stimulating cell migration and tissue factor synthesis, and polymorphonuclear leukocytes (PMNs), promoting chemotaxis and superoxide generation. In vivo, subcutaneously administered EMAPII elicits inflammation; but Meth A tumor cells transfected to constitutively overexpress EMAPII exhibit increased growth. We hypothesize that the context and kinetics of EMAPII production determine its biologic properties, either as phlogogenic agent in inflammatory settings, or as inducer of neoangiogenesis in tumors. The specific aims are to understand mechanisms by which EMAPII elicits inflammation, and influences tumor-host reactions. Using cDNA probes and antibodies to EMAPII (murine and human), we will identify sites of synthesis and accumulation in a spectrum of tumors, inflammatory and vascular lesions. Functional activity of a region of mature EMAPII near the HN2-terminus, which, in synthetic peptides, stimulates PMN and MP migration, binds specifically to cells, and cross-links to an about 73 kDa cell surface polypeptide will be examined using additional peptides and site-directed mutagenesis. These reagents will also be used to characterize and isolate the EMAPII cell surface binding site. Based on strong sequence homology between the NH2-terminal region of EMAPII and von Willebrand antigen II (vWAgII, a polypeptide in platelet alpha- granules and released by stimulated ECs), we have shown that vWAgII has properties resembling EMAPII. Because of the role of secreted platelet proteins in tissue repair, we will assess the effects of vWAgII on ECs, MPs, and PMNs, and determine its contribution to inflammation and wound repair in animal models. And since elevated levels of vWAgII, consequent on infusion of the arginine vasopressin analog, DDAVP, induced MP tissue factor, we will find out whether vWAgII is capable of initiating activation of procoagulant mechanisms in vivo, and the thrombosis complicating DDAVP therapy. These studies offer new means of linking coagulation with inflammation, and define a new ligand-receptor interaction, involving EMAPII, relevant to host responses in inflammation, neoplasia and, vasculopathies, such as atherosclerosis.