In vitro models of allograft reactions suggest central roles for IL-2-induced proliferation of T helper (TH) cells and IL-2/IL-4-induced maturation of cytotoxic T cells CTL). The sponge matrix model of class I plus class II major histocompatibility complex (MHC) allograft rejection in vivo confirms the presence of TH cells and CTL maturation from pre-CTL at the graft site, but other evidence to support the conventional paradigm is absent. Bioassays show evidence of accessory cell cytokines (IL-1,TNFa M-CSF) and the TH type 2 (TH2) product IL-6, but proliferation of TH cells is absent, and TH1 cytokines (IL-2, interferon gamma) cannot be found. Furthermore, the TH2 product IL-4 is absent by bioassay. In order to test the hypothesis that TH1 activity is absent in the in vivo model and that some TH2 functions are downregulated, we plan to further analyze the allograft for critical TH1 (IL-2, IFNG) and TH2 (IL-4, IL-5) cytokines by bioassay, enzyme-immune assay, and messenger RNA in sponge cells. Evidence for actual selection of TH2 vs TH1 subtypes in vivo will be sought by limiting dilution analysis of sponge cells under conditions which permit each of the subtypes to be cloned. The role of TH2 cytokines in CTL maturation will be determined utilizing pre-CTL from the sponge allograft in an assay of antigen-specific CTL maturation. Because the sponge allograft is rejected in the context of an acute inflammatory response plan to analyze the effects of several selected components of that response (prostaglandin E2, nitric oxide, arginine, tumor necrosis factor alpha, and transforming growth factor beta) on the cellular proliferation and cytokine production by TH1 and TH2 subsets, as well as on the maturation of pre-CTL to specific CTL. We hypothesize that some combination of acute inflammatory factors will downregulate cellular proliferation and foster selective cytokine production, while permitting CTL maturation in vivo. Considerable information concerning the discrepancies between in vivo events and in vitro models of allograft rejection should result.
