CHARACTERIZATION OF THE PRODUCTION OF MONOCYTE CHEMOATTRACTANT PROTEIN-1 AND IL-8 IN AN ALLOGENEIC IMMUNE-RESPONSE

Delayed-type hypersensitivity and allograft rejection are dependent up on the generation of Ag-specific T cell immune responses. The mixed ly mphocyte reaction is a model of alloantigen-driven immunity and has pr ovided significant insight into the mechanisms of T cell proliferation . Recently, soluble mediators, including TNF-alpha have been shown to be involved in the full expression of this response. In order to eluci date the potential mechanisms of cellular recruitment in the context o f either delayed-type hypersensitivity or allograft rejection, we empl oyed a mixed lymphocyte reaction to study the production of chemokines , monocyte chemoattractant protein-1 (MCP-1), and IL-8. Time-course ex periments demonstrated that significant levels of MCP-1 and IL-8 were produced in allogeneic cultures as early as day 1, and that this relat ionship persisted over 6 days in culture. Northern blot analysis of ch emokine mRNA confirmed the early induction of these genes in the allog eneic response. The levels of MCP-1 and IL-8 protein from mixed lympho cyte reaction supernatants as measured by specific ELISA were positive ly correlated with the proliferative response as measured by [H-3]TdR uptake. Although significant MCP-1 and IL-8 were produced during a mix ed lymphocyte reaction, these molecules did not participate in the pro liferative response, as neutralizing antibodies to either MCP-1 or IL- 8 had no effect on proliferation. In order to ascertain the mechanism for the induction of MCP-1 and IL-8 during the mixed lymphocyte reacti on, experiments were performed in the presence of neutralizing anti-hu man TNF antibodies. Neutralization of TNF resulted in significant abro gation of MCP-1 and IL-8 production (75 +/- 5 and 87 +/- 2 percent, re spectively). Cells isolated from the mixed lymphocyte reaction at day 6, demonstrated that MCP-1 and IL-8 Ag were localized to mononuclear p hagocytes. These results demonstrate that potent chemokines, MCP-1 and IL-8, are produced during the evolution of a mixed lymphocyte reactio n, and their induction is TNF-dependent.