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Mice were infected with Listeria monocytogenes and Lyt-2+ T cell clones capable of lysing Ag-primed bone marrow macrophages were established. In accordance with earlier findings obtained at the population level, some T cell clones were identified which lysed bone marrow macrophages of different MHC type provided the relevant Ag was present. This unusual target cell recognition was further analyzed using a T3+, L3T4-, Lyt-2+, F23+, KJ16+ T cell clone, designated L-28. Target cell lysis by this clone was Ag specific, apparently non-MHC restricted. In contrast, YAC cells and P815 cells were not lysed by clone L-28. However, lysis of irrelevant targets could be induced by anti-T3, F23, or KJ16 mAb. Furthermore, Ag-specific lysis was blocked by anti-Lyt-2 mAb and by F(ab)2 fragments of F23 mAb. In addition to its cytolytic activity, clone L-28 produced IFN-gamma after co-stimulation with accessory cells, Ag, and rIL-2 and conferred significant protection on recipient mice when given together with rIL-2. These data suggest that non-MHC-restricted Lyt-2+ killer cells generated during listeriosis are cytolytic T lymphocytes that interact with their target Ag via the T cell receptor/T3 complex and the Lyt-2 molecule and, furthermore, that these cells play a role in anti-listerial resistance. The possible relevance of IFN-gamma secretion and target cell lysis for antibacterial protection is discussed.

In vitro expanded T cell lines were used to determine whether antigen-specific cytolytic T lymphocytes are generated after infection with the intracellular bacterium, Listeria monocytogenes. Spleen cells from infected mice were cultured in the presence of syngeneic accessory cells, listerial antigen, and interleukin 2 containing supernatants. Cell lines were greater than 98% Thy-1+, L3T4-, Lyt-2+. Bone-marrow macrophages were used as target cells in two in vitro cytolytic assay systems. The Lyt-2+ T cells killed bone marrow macrophages only when infected with L. monocytogenes as assessed in a 4-hr 51Cr release assay and in an 18-hr neutral red uptake assay. Cytolysis was blocked by anti-LFA-1 and anti-Lyt-2 monoclonal antibodies. These cytolytic T cells produced interferon-gamma after co-stimulation with antigen, accessory cells, and recombinant interleukin 2. Bone marrow macrophages infected with Mycobacterium bovis were not killed by T cells from L. monocytogenes-infected mice but by T cell lines from M. bovis-infected mice, indicating that cytolysis was antigen specific. L. monocytogenes-infected target cells of different haplotype were lysed by the Lyt-2+ T cells. By using a low cell density split culture system, antigen-specific, H-2-restricted cytolytic T cells could be identified. These findings demonstrate that during infection with intracellular bacteria, Lyt-2+ T cells with cytolytic activity are generated that may be involved in antibacterial protection.

The effect of tuberculin purified protein derivative (PPD) on the development of plaque-forming cells (PFC) against bromelain-treated syngeneic mouse red blood cells (Br-MRBC) was studied in peritoneal cell cultures. The finding that PPD enhances the development of PFC to Br-MRBC, even under conditions where cell division is blocked by mitomycin C treatment, suggests that cell proliferation does not represent a necessary prerequisite for differentiation of precursor cells into autoantibody-forming cells.

Cytokines have been suggested to act as intermediates between the immune and the central nervous system, but little is known about the type of cells synthesizing them in the brain. We have immortalized with oncogenic retroviruses primary brain cell cultures from mouse embryos and have generated clones of microglial cells that have been characterized. Three of the clones studied produce interleukin 1 (IL 1); IL 6 and tumor necrosis factor-alpha as assessed by biological assays and by Northern blot analysis. Our data raise the question on the role of these cytokines in the brain and suggest that early resident microglial cells might play an important role in development processes and in the adult brain.

Mice were infected with the intracellular bacterium, Listeria monocytogenes, and T cell clones from spleens, lymph nodes and peritoneal exudates were established. The capacity of L3T4+, Lyt2- T-cell clones to specifically lyse L. monocytogenes-infected macrophages was analyzed. As a source of target cells, bone marrow macrophages (BMM phi) after 9 days of culture in hydrophobic teflon bags were used. These BMM phi were totally Ia-; however, significant Ia-expression could be induced by interferon-gamma (IFN-gamma). IFN-gamma-stimulated BMM phi, after priming with live or killed L. monocytogenes organisms were effectively lysed by the vast majority of L3T4+ T cell clones. In the absence of either IFN-gamma stimulation or antigen priming, no lysis occurred. Cytolysis was demonstrable in a conventional 4-h 51Cr-release assay and in an 18-h neutral red uptake assay and was antigen specific and class II restricted. Native T cells from L. monocytogenes-infected mice failed to lyse stimulated, L. monocytogenes-primed BMM phi and gained their cytolytic activity after antigenic restimulation in vitro. These data demonstrate that L. monocytogenes-specific L3T4+ T cells could lyse M phi presenting listerial antigens provided that Ia antigen expression had been induced. L3T4+ T cell clones produced IFN-phi after restimulation with antigen plus accessory cells in vitro and IFN-gamma secretion could be increased by costimulation with recombinant IL 2. These T cell clones conferred significant protection upon recipient mice which was more pronounced in the liver. The possible relevance of lysis by L3T4+ T cells of infected M phi to protection against and pathogenesis of intracellular bacterial infections is discussed.

Human thymocytes were sorted according to the expression of CD4 and CD8 molecules and clones representing the four subpopulations (DP, DN, and single CD4 or CD8 positive) were established. DP clones can be maintained for long periods in tissue culture and give rise to a variable percentage of SP variants. These variants, when isolated and further expanded, do not revert to a DP phenotype. DP clones express a functional TCR-CD3 complex, suggesting that this molecule can interact with the thymic microenvironment during T cell differentiation.

Killed Staphylococcus aureus strain Cowan I cells inhibit contact sensitivity to oxazolone in mice, when given intravenously 24-72 h before sensitization. With transfer experiments it was found that the cells responsible for the suppression are antigen-specific, nylon-adherent, resistant to antitheta serum + C, and sensitive to anti-mouse Ig serum + C. These suppressor B cells bear anti-oxazolone immunoglobulins and appear to exert their suppressive activity by preventing the contact sensitizer from reaching the specific reactive T cells.