We address the mechanism of hybrid resistance (HR) in vitro using NK effector cells and target lymphoblasts from H-2b, H-2d, and H-2b/d mice. The 5E6 (Ly49C)+ subset of F1 NK cells lyse BALB/c (H-2d) but not B6 (H-2b) targets unless either anti-5E6 or anti-H-2Kb MAbs are present. H-2Dd transgenic B6 (D8) targets are not susceptible to F1 Ly49A+ effectors. Furthermore, 5E6+ Ly49A+ F1 effectors lyse B6 and BALB/c targets only in the presence of anti-5E6 and anti-Ly49A MAbs, respectively. Thus, recognition of H-2Kb by 5E6 and H-2Dd by Ly49A transduce independent inhibitory signals. Moreover, anti-5E6 MAbs enable 5E6+ BALB/c NK cells to lyse (BALB/c x B6)F1 targets. These data support the "missing self" and not the "hemopoietic histocompatibility antigen" hypothesis for HR. In addition, 5E6+ NK cells from BALB/c and BALB.B, but not B6 or (BALB/c x B6)F1, mice receive negative signals from both H-2d and Kb class I antigens. Thus, allelic differences in 5E6 (C57BL versus BALB) may regulate recognition events by NK cells.

A rabbit was immunized with rabbit immunoglobulins of a different allotype. The anti‐allotypic antibodies produced by this rabbit were used to immunize a second rabbit which produced anti‐idiotypic antibodies. To explain the occurrence, among these anti‐idiotypic antibodies, of “internal images” of the original immunizing allotype, a restricted and a more general hypothesis are developed. The first assumes that B‐cells can be triggered when idiotopes on their receptor molecules are recognized by the paratopes of the immunizing antibody. The second denies the existence of a specially constructed combining site on the variable domain of an antibody molecule.

An important feature of natural killer (NK) cell activity is the lysis of cells that have extinguished expression of some or all class I major histocompatibility (MHC) molecules. Accordingly, the Ly49A NK-cell antigen receptor has been shown to deliver an inhibitory signal to NK cells on encounter with Dd or Dk class I MHC on target cells. Ly49A belongs to a family of eight or more highly related, tightly linked genes. Expression of Ly49A and Ly49C, another member of the Ly49 family with distinct MHC specificity, define subpopulations of NK cells that are only partly overlapping. The mechanisms regulating the expression of LY49 family members are unknown. We show here that the Ly49A and Ly49C NK-cell receptors are each subject to allelic exclusion. Because Ly49 genes are not thought to undergo DNA rearrangement, allelic exclusion of Ly49 genes could involve a mechanism distinct from that used by B and T lymphocytes and is likely to play an important role in the genesis of a putative NK-cell repertoire specific for class I molecules.

Experimental infection of C57BL/6 mice by Plasmodium yoelii sporozoites induced an increase of CD4−CD8− NK1.1+ TCRαβint cells and a down-regulation of CD4+ NK1.1+ TCRαβint cells in the liver during the acute phase of the infection. These cells showed an activated CD69+, CD122+, CD44high, and CD62Lhigh surface phenotype. Analysis of the expressed TCRVβ segment repertoire revealed that most of the expanded CD4−CD8− (double-negative) T cells presented a skewed TCRVβ repertoire and preferentially used Vβ2 and Vβ7 rather than Vβ8. To get an insight into the function of expanded NK1.1+ T cells, experiments were designed in vitro to study their activity against P. yoelii liver stage development. P. yoelii-primed CD3+ NK1.1+ intrahepatic lymphocytes inhibited parasite growth within the hepatocyte. The antiplasmodial effector function of the parasite-induced NK1.1+ liver T cells was almost totally reversed with an anti-CD3 Ab. Moreover, IFN-γ was in part involved in this antiparasite activity. These results suggest that up-regulation of CD4−CD8− NK1.1+ αβ T cells and down-regulation of CD4+ NK1.1+ TCRαβint cells may contribute to the early immune response induced by the Plasmodium during the prime infection.

Various components of innate and adaptive immunity contribute to host defenses against Plasmodium infection. We investigated the contribution of NK cells to the immune response to primary infection with Plasmodium yoelii sporozoites in C57BL/6 mice. We found that hepatic and splenic NK cells were activated during infection and displayed different phenotypic and functional properties. The number of hepatic NK cells increased whereas the number of splenic NK cells decreased. Expression of the Ly49 repertoire was modified in the spleen but not in the liver. Splenic and hepatic NK cells have a different inflammatory cytokines profile production. In addition, liver NK cells were cytotoxic to YAC-1 cells and P. yoelii liver stages in vitro but not to erythrocytic stages. No such activity was observed with splenic NK cells from infected mice. These in vitro results were confirmed by the in vivo observation that Rag2−/− mice were more resistant to sporozoite infection than Rag2−/− γ c−/− mice, whereas survival rates were similar for the two strains following blood-stage infection. Thus, NK cells are involved in early immune mechanisms controlling Plasmodium infection, mostly at the pre-erythrocytic stage.

In this paper we describe a monoclonal antibody, JR9-318, which was obtained from a wild-derived mouse immunized with the M14T cell line (CD3+CD4-CD8-) in order to describe antigenic molecules expressed during T cell differentiation. This mAb precipitated a protein from the M14T cell line or subclones that migrates like a T cell receptor heterodimer at 85-90 kDa under non-reducing conditions and 45-50 kDa under reducing conditions. The molecular characteristics of this antigen and the data obtained by sequential immunoprecipitation led us to conclude that JR9-318 mAb recognizes the Ly-49 molecule already described. Using direct immunofluorescence staining, this mAb was shown to identify a small population of cells in the adult thymus but not in the spleen, lymph node or bone marrow of most laboratory mouse strains. The phenotypic characteristics of this thymic subpopulation do not correspond to those of any other known classical population. A more prominent population of CD3+ alpha beta+ T cells is stained by this mAb in intestinal intra-epithelial lymphocytes with a peculiar phenotype, different from the one found in the thymus. These cells express V beta families usually deleted in the thymus of BALB/c mice. We discuss here the origin of these T cell subpopulations and the possible maturation of T cells without thymic influence.

NKT cells are a population of innate-like lymphocytes that display effector functions and immunoregulatory properties. We characterized the NKT cell response induced in C57BL/6 mice during a primary infection with Plasmodium yoelii sporozoites. We observed a heterogeneous NKT cell response that differed between liver and spleen. Hepatic NKT cells found in infected livers consisted mainly of CD1d-dependent CD4؉ and doublenegative (DN) NKT cells, whereas CD1d-independent NKT cells exhibiting a TCR high CD4 high phenotype were prominent among splenic NKT cells during the infection. Hepatic and splenic NKT cells isolated from infected mice were activated and secreted mainly gamma interferon and tumor necrosis factor alpha in response to stimulation. Finally, P. yoelii-activated hepatic DN NKT cells inhibited the parasite's liver stage in a CD1d-dependent manner in vitro. However, experiments using B6.CD1d-deficient mice showed that CD1d and CD1d-restricted NKT cells are not necessary to control the parasite's development in vivo during neither the preerythrocytic stage nor the erythrocytic stage. Thus, our results show that a primary P. yoelii infection induces a heterogeneous and organ-specific response of NKT cells and that CD1d-dependent NKT cells play a minor role in the control of the development of Plasmodium in vivo in our model.

The Ly49 family of natural killer (NK) cell receptors is encoded by a polygenic genetic locus. Allelic forms have been described and their expression appears to be regulated. The best-characterized Ly49 molecule, the C57BL/6 form of Ly49A, is an NK cell inhibitory receptor that binds H2Dd. To determine whether differences between Ly49a alleles may have functional consequences, allelic variants of Ly49a were cloned from several inbred mouse strains. Stable transfectants expressing each Ly49a allelic variant were generated and tested for reactivity with a panel of monoclonal antibodies (mAbs A1, JR9.318, YE1/32, and YE1/48) that recognize the C57BL/6 form of Ly49A. Binding to H2Dd was also assessed using fluorescently labeled H2Dd tetramers. Furthermore, cytotoxicity assays were performed using anti-Ly49A mAb-separated interleukin-2-activated NK cells. We show that despite binding to fluorescently labeled H2Dd tetramers, the Ly49A+ NK cells from representative mouse strains displayed significantly different degrees of inhibition with H2Dd targets. These results can be interpreted in the light of recent structural data on the Ly49A-H2Dd complex. Thus, the Ly49 family displays functionally significant allelic polymorphism which adds to the repertoire of NK cell receptors.