IL-12 initiates Th1 cell development and cell-mediated immunity, but whether IL-12 contributes to the maintenance of a Th1 response is unclear. To address this question, we infected IL-12 p40−/− C57BL/6 mice with Leishmania major, an intracellular protozoan parasite controlled by a cell-mediated immune response, and simultaneously administered IL-12. Whereas untreated p40−/− mice developed an uncontrolled infection, p40−/− mice treated with IL-12 for the first 2 or 4 wk of infection developed a Th1 response and resolved their lesions. However, the induction of this protective Th1 cell response by IL-12 treatment was not associated with long term immunity. We observed that on rechallenge in the absence of IL-12, the mice exhibited a susceptible phenotype. In addition, without rechallenge, lesions in the IL-12-treated p40−/− mice developed several weeks after cessation of IL-12 treatment. In both cases, disease was associated with the loss of a Th1 response and the development of a Th2 response. Our observations are not limited to the C57BL/6 strain, because IL-12 treatment was also unable to provide lasting protection to p40−/− BALB/c mice. Finally, we found that although Th1 cells from healed wild-type C57BL/6 mice adoptively transferred protection to L. major-infected RAG−/− mice, they were unable to protect p40−/− mice. In conclusion, these studies provide the first demonstration that IL-12 is required not only to initiate Th1 cell development but also throughout infection to maintain a Th1 cell response and resistance to L. major.

IL-12p40 is required for the maintenance of resistance during Leishmania major infection. In this study, we addressed how IL-12 mediates this function. First, we demonstrated that both subunits of IL-12, p40 and p35, were required for continued resistance to L. major. Second, using IL-12, IL-4 doubly deficient mice, we investigated the possibility that IL-12 inhibits IL-4-induced outgrowth of Th2 cells that might compete with Th1 cells. We found that even in the absence of a Th2 response, IL-12 was still required to maintain resistance. Next, using adoptive transfer of Thy-1 disparate CD4+ T cells from L. major-healed mice, we were able to show that the loss of a protective response in L. major-infected IL-12-deficient mice is linked with the loss of Th1 cells. In contrast, there was an equal recovery of CD4+ Th1 cells from wild-type and IL-12-deficient mice when transferred into mice that were not challenged with L. major. The ability of Th1 cells to survive regardless of IL-12 levels in the absence of Ag stimulation was confirmed by adoptive transfer studies of CD4+ Th1 cells from DO11.10 TCR transgenic mice. Taken together, these results indicate that, rather than modulating Th2 responses or optimizing IFN-γ production, the critical role for IL-12 in maintaining cell-mediated immunity may be to prevent the loss of Th1 cells during a challenge infection.

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IL-2-deficient (IL-2−/−) mice develop disorders of the hemopoietic and immune systems characterized by anemia, lymphocytic hyperplasia, and colitis. The mechanisms responsible for these abnormalities remain unclear. To investigate the underlying basis of autoimmunity, the particular role of commensal gut flora in the initiation of colitis, and the role of IL-2 in the development of intestinal intraepithelial lymphocytes (iIEL), we evaluated IL-2−/− mice reared and maintained under gnotobiotic (germfree) conditions. By 8 wk of age, 80% (20 of 25) of germfree IL-2−/− mice show signs of disease, including anemia, disturbances in bone marrow hemopoietic cells, lymphocytic hyperplasia, and generalized autoimmunity, similar to those seen in specific pathogen-free (SPF) IL-2−/− mice. In striking contrast to SPF IL-2−/− mice, germfree IL-2−/− mice do not develop colitis. However, the numbers of γδ+ and TCRαβ+CD8αα+ iIELs are reduced, and in lethally irradiated SPF IL-2+/+ mice, reconstituted with IL-2−/− bone marrow TCRγδ+ iIELs fail to develop, consistent with an important role of IL-2/IL-2R signaling in the development of γδ iIELs. Consequently, our findings demonstrate that the colitis seen in SPF IL-2−/− mice depends upon the presence of intestinal bacterial flora and that environmental Ags are not responsible for the anemia and extraintestinal lymphoid hyperplasia that occur in IL-2−/− mice. Thus, germfree IL-2−/− mice represent a unique system in which the role of IL-2 deficiency in hemopoietic and immune system disorders can be investigated in dissociation from complications that may arise due to colitis.