Exposure to IL-4 during activation of naive murine CD8+ T cells leads to generation of IL-4-producing effector cells with reduced surface CD8, low perforin, granzyme B and granzyme C mRNA, and poor cytolytic function. We show in this study that maximal development of these cells depended on exposure to IL-4 for the first 5 days of activation. Although IL-4 was not required at later times, CD8 T cell clones continued to lose surface CD8 expression with prolonged culture, suggesting commitment to the CD8low phenotype. This state was reversible in early differentiation. When single CD8low cells from 4-day cultures were cultured without IL-4, 65% gave rise to clones that partly or wholly comprised CD8high cells; the proportion of reverted clones was reduced or increased when the cells were cloned in the presence of IL-4 or anti-IL-4 Ab, respectively. CD8 expression positively correlated with perforin and granzyme A, B, and C mRNA, and negatively correlated with IL-4 mRNA levels among these clones. By contrast, most CD8low cells isolated at later time points maintained their phenotype, produced IL-4, and exhibited poor cytolytic function after many weeks in the absence of exogenous IL-4. We conclude that IL-4-dependent down-regulation of CD8 is associated with progressive differentiation and commitment to yield IL-4-producing cells with little cytolytic activity. These data suggest that the CD4−CD8− cells identified in some disease states may be the product of a previously unrecognized pathway of effector differentiation from conventional CD8+ T cells.

A carbohydrate enriched soluble fraction (CHP) was prepared by mild treatment of viable Echinococcus granulosus protoscoleces (PSC) with the enzyme endoglycosidase-F (endo-F) and characterized by SDS-PAGE, glycoside- inhibition ELISA, and immunoblotting. Three groups of four BALB/c mice were immunized with the CHP, with the remaining deglycosylated PSC (DGP) and with dead PSC (DPSC) to analyse the relative immunogenicity of carbohydrates on the surface of PSC. A fourth sentinel group was not immunized. The antibody response was analyzed during primary and hyperimmune responses. Specific antibody titres (IgG and IgM) against somatic PSC antigens (PSA) were evaluated by ELISA and their antigen recognition pattern by immunoblotting, discriminating carbohydrate and peptide specific antibody responses by periodate treatment of PSA. The avidity index of those antibodies and the titer of non-specific immunoglobulins during the whole protocol were evaluated by ELISA in vitro mitogenic activity of CHP was also evaluated. The results indicated: 1) immunodominance of surface carbohydrate epitopes from PSC, 2) predominant IgM and low avidity response against these epitopes, 3) a dramatic increase of non-specific antibody titres and 4) in vitro mitogenic activity of CHP.

CD8low ͉ co-receptor tuning ͉ T cell activation ͉ cytotoxic T lymphocytes ͉ cytokines T he CD8␣␤ co-receptor amplifies the CD8 ϩ T cell response to peptide/MHC Class I complexes on antigen-presenting cells (APC) by at least four mechanisms. First, CD8 binding to nonpolymorphic regions of the MHC molecule and ␤2-microglobulin is thought to stabilize the interaction between the TCR and peptide/MHC complexes (1); second, CD8 binding to the MHC augments TCR signaling via activation of p56 lck and LAT (2, 3); third, CD8 is thought to induce the co-localization of receptor complex molecules onto lipid rafts (4); and fourth, CD8 induces a conformational change in CD3 that is necessary for signal transduction (5). The effect can be dramatic: for example, transfection of CD8-negative T cells bearing lowaffinity TCR with CD8␣ or CD8␣/CD8␤ has been reported to amplify peptide sensitivity by factors of 10 7 to 10 9 (6). Modulation of surface CD8 expression on naive and effector cells can therefore alter the threshold peptide/MHC levels required to trigger proliferation, cytokine production and target cell lysis (7, 8), effectively ''tuning'' the T cell response.Surface CD8 levels can be controlled by distinct antigen-and cytokine-dependent mechanisms. The former has been observed as a transient response to TCR activation (9) and in peripheral tolerance where chronic antigen exposure can be associated with stable CD8 down-regulation at the T cell surface (7, 10, 11). Cytokine-dependent CD8 down-regulation was first reported by Erard et al. (12) who demonstrated that exposure of naïve CD8 ϩ T cells to IL-4 during activation with phorbol 12-myristate 13-acetate (PMA) and ionomycin in vitro led to loss of surface CD8 and CD8␣ RNA, induction of a type 2 cytokine profile and the absence of cytolytic activity. We extended these studies to show that the presence of IL-4 in the first few days of primary activation with antibodies to the TCR, CD8 and CD11a (antireceptor Ab) or alloantigens promoted the development of poorly cytolytic, type 2 effector cells that retained their TCR but displayed reduced or undetectable surface CD8 expression; essentially all conventional CD8 ϩ T cells could eventually give rise to committed CD8 low cells that maintained this phenotype in the absence of continued IL-4 exposure (13,14). By contrast, Park et al. have recently reported that IL-4 and other ␥c cytokines up-regulated surface CD8 levels when CD8 ϩ T cells were cultured in vitro without a TCR stimulus (7).IL-4 and IFN-␥ reciprocally regulate a number of lymphocyte functions, including the expression of type 1 and type 2 cytokines by CD4 ϩ and CD8 ϩ effector cells (15,16). The present study was therefore undertaken to determine whether this was also true for CD8 expression in newly activated CD8 ϩ T cells. We show here that IFN-␥ not only counteracts the down-regulatory effect of IL-4 on CD8 mRNA and protein levels but also contributes to the maintenance of CD8 levels in the absence of exogenous IL-4. The reciprocal effects of IL-4 and IFN-␥ we...

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