To explore which cytokine or cell is essential for the production of antibodies (Abs) of the IgE class in allergic diseases, we injected cedar pollen into wild-type, interferon-gamma(-/-) (IFN-gamma(/)), or interleukin-4(-/-) (IL-4(-/-)) BALB/c mice through four (i.n., i.p., s.c., and i.v.) different routes without adjuvant. Wild-type or IFN-gamma(-/-), but not IL-4(-/-), mice sensitized once or twice showed a significant increase in total IgE Ab in their serum, revealing the essential role of IL-4 in the production of total IgE Ab. We separated peripheral blood mononuclear cells (PBMCs) from untreated or sensitized mice into monocyte-rich, lymphocyte-rich, and granulocyterich populations by Percoll density-gradient centrifugation or into specific antigen cells by flow cytometry, cultured the cells in various combinations, and examined the levels of cytokines and IgE Ab released into the medium. The PBMCs from mice sensitized s.c. once, but not those from untreated animals, produced significant amounts of IL-4 and total IgE Ab, whereas the lymphocyte-rich population alone did not. Unexpectedly, IL-4 and IgE Ab production was restored by the addition of Mac-1(+) cells in the monocyte-rich fraction to the lymphocyte-rich fraction. These results indicate the essential role of monocytes in the production of IL-4 and total IgE Ab by lymphocytes during the initial stage of sensitization.

The generation of knockout mice demonstrated that CD4+, but not CD8+, T cells were essential for the rejection of allografted skin or heart, presumably because these targets were CTL resistant. In the case of CTL-susceptible targets (e.g., P815 mastocytoma cells and EL-4 or RLmale1 T lymphoma cells), however, it is assumed that the CTL is the effector cell responsible for allograft rejection and that perforin and Fas ligand (FasL) pathways are the killing mechanisms. In the present study, we examined the role of these cytotoxic molecules in the rejection of i.p. allografted CTL-susceptible leukemia cells. Unexpectedly, the allografted leukemia cells were acutely rejected from gld (a mutation of FasL), perforin−/−, or double-deficient mice. The peritoneal exudate cells from gld or normal mice showed T cell-, TCRαβ-, and perforin-dependent cytotoxic activity against the allograft, whereas the exudate cells from perforin−/− mice exhibited almost full cytotoxic activity in the presence of Fas-Fc. Furthermore, the infiltrates from double-deficient mice showed a high cytotoxic activity against the allografted cells even in the presence of anti-TCRαβ Ab or in the absence of T cells. The cytotoxic cells appeared to be macrophages, because they were Mac-1+ mononuclear cells with a kidney- or horseshoe-shaped nucleus and because the cytotoxic activity was completely suppressed by the addition of NG-monomethyl-l-arginine, an inhibitor of inducible NO synthase. These results indicate that macrophages are ready and available to kill CTL-susceptible allografts when CTLs lack both perforin and FasL molecules.

The production of allergen‐specific IgE antibodies (Abs) in allergen‐sensitized patients or animals has a mutual relationship with the immunologic response leading to allergic rhinitis. We recently reported that, after an intranasal injection of cedar pollen into mice, an interleukin‐4 (IL‐4)‐dependent increase in serum nonspecific IgE Abs was a prerequisite for the production of serum allergen‐specific IgE Abs. Here, we explored which lymphoid organs were responsive to the intranasally injected allergen and how IL‐4 and IgE Abs were produced in the lymphocytes. Time‐dependent changes in the total cell numbers and in in vitro IgE Ab production in various lymphoid organs revealed that the submandibular lymph nodes were the main responsible organ. After treatment with allergen (for IgE production) or allergen and complete Freund's adjuvant (for IgG production), we separated submandibular lymph node cells into macrophage‐, lymphocyte‐, and granulocyte‐rich populations by discontinuous Percoll density‐gradient centrifugation. Unexpectedly, bulk cells, but not the lymphocyte‐ or macrophage‐rich populations, produced significant amounts of IL‐4, IgE, and IgG; whereas production was restored by addition of Mac‐1+ cells from the macrophage‐rich to the lymphocyte‐rich fraction. Furthermore, a combination of the lymphocyte‐rich population (for IgG [or IgE]) production) and the macrophage‐rich population (for IgE [or IgG]) production) produced a large amount of IgE (or IgG). These results indicate that, in the initiation of allergic rhinitis, macrophages in the submandibular lymph nodes are essential not only for IL‐4 or immunoglobulin production, but also for class switching of immunoglobulin in lymphocytes.

No abstract