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L Moretta

Publications and source records attributed to L Moretta.

At least 73 records · Page 4Linked to original sources
PubMed1998

NKp44, a novel triggering surface molecule specifically expressed by activated natural killer cells, is involved in non-major histocompatibility complex-restricted tumor cell lysis.

After culture in interleukin (IL)-2, natural killer (NK) cells acquire an increased capability of mediating non-major histocompatibility complex (MHC)-restricted tumor cell lysis. This may reflect, at least in part, the de novo expression by NK cells of triggering receptors involved in cytolysis. In this study we identified a novel 44-kD surface molecule (NKp44) that is absent in freshly isolated peripheral blood lymphocytes but is progressively expressed by all NK cells in vitro after culture in IL-2. Different from other markers of cell activation such as CD69 or VLA.2, NKp44 is absent in activated T lymphocytes or T cell clones. Since NKp44 was not detected in any of the other cell lineages analyzed, it appears as the first marker specific for activated human NK cells. Monoclonal antibody (mAb)-mediated cross-linking of NKp44 in cloned NK cells resulted in strong activation of target cell lysis in a redirected killing assay. This data indicated that NKp44 can mediate triggering of NK cell cytotoxicity. mAb-mediated masking of NKp44 resulted in partial inhibition of cytolytic activity against certain (FcgammaR-negative) NK-susceptible target cells. This inhibition was greatly increased by the simultaneous masking of p46, another recently identified NK-specific triggering surface molecule. These data strongly suggest that NKp44 functions as a triggering receptor selectively expressed by activated NK cells that, together with p46, may be involved in the process of non-MHC-restricted lysis. Finally, we show that p46 and NKp44 are coupled to the intracytoplasmic transduction machinery via the association with CD3zeta or KARAP/DAP12, respectively; these associated molecules are tyrosine phosphorylated upon NK cell stimulation.

Biomarkers↗
PubMed1998

HLA class I-specific inhibitory receptors in human T lymphocytes: interleukin 15-induced expression of CD94/NKG2A in superantigen- or alloantigen-activated CD8+ T cells.

A fraction of human T lymphocytes, predominantly CD8+, express receptors for HLA class I molecules typical of natural killer cells (natural killer receptors or NKRs) that inhibit T cell receptor-mediated functions. Herein, we analyzed possible mechanism(s) leading to the expression of NKRs by T cells responding to superantigens or allogeneic cells in vitro. We show that, in the presence of interleukin 15 (IL-15), T cells (depleted of NKR+ cells) responding to toxic shock syndrome toxin 1 de novo express CD94, a molecule that is part of a heterodimeric NKR with a broad specificity for different HLA class I alleles. Maximal CD94 expression occurred when IL-15 was added shortly after the cells were placed into culture, and CD94 expression started 4-6 days after addition of IL-15. Although both CD4+ and CD8+ cells expressed CD94, the simultaneous expression of NKG2A (i.e., the other component of the CD94/NKG2A inhibitory NKR) was confined to CD8+ cells. Similar data were obtained in T cell populations activated in mixed lymphocyte cultures in the presence of IL-15. The expression of CD94/NKG2A led to an impairment of allo-specific cytolytic activity by mixed lymphocyte culture-derived T cell populations or clones. Remarkably, cytolysis could be restored by the addition of anti-CD94 mAb, i.e., by masking the inhibitory NKRs.

Antigens, CD↗
PubMed1998

The activating form of CD94 receptor complex: CD94 covalently associates with the Kp39 protein that represents the product of the NKG2-C gene.

Inhibitory receptor complexes formed by CD94 and NKG2-A (Kp43) molecules have been implicated in HLA class I recognition by human natural killer (NK) cells. Additional forms of CD94 receptors have recently been described in NK cells characterized by the lack of NKG2-A expression. These CD94 receptors were shown to display activating functions. Immunoprecipitation with anti-CD94 monoclonal antibodies (mAb) led to the identification, in these cells, of a 39-kDa (Kp39) molecule that was originally believed to represent an activating isoform of the CD94 molecules. In the present study we show that the Kp39 molecule is covalently associated with CD94 and displays a protein backbone (26 kDa) similar to that of NKG2-A (Kp43) glycoproteins. Peptide mapping analysis indicates that Kp39 and NKG2-A glycoproteins belong to the same molecular family. A novel NKG2-specific mAb (termed P25) has been generated that specifically reacts with both NKG2-A and NKG2-C molecules, but fails to recognize NKG2-E molecules. Analysis of polyclonal and clonal NK cells shows that P25 mAb reacts with all NKG2-A+ cells and with a fraction of CD94+ cells lacking the expression of NKG2-A. These data indicate that NKG2-C molecules are indeed expressed only in a subset of cells lacking the expression of NKG2-A. The CD94-associated Kp39 molecule can be detected only in NKG2-A- P25+ cells, i.e. cells expressing NKG2-C molecules. Indeed, reverse transcription-polymerase chain reaction analysis performed on a large panel of NK clones indicates that NKG2-A- P25+ NK clones express the NKG2-C transcript. Notably, the cytolytic activity of these clones can be triggered by the P25 mAb in redirected killing analysis. Finally, biochemical analysis of COS7 cells cotransfected with CD94 and NKG2-C demonstrates the identity between Kp39 and NKG2-C molecules. Altogether, our data demonstrate that NKG2-C molecules associate with CD94 to form an activating receptor complex in a subset of human NK cells.

Amino Acid Sequence↗
PubMed1998

IL-12-induced up-regulation of NKRP1A expression in human NK cells and consequent NKRP1A-mediated down-regulation of NK cell activation.

IL-12, in contrast to IL-2, strongly up-regulated the expression of the NKRP1A lectin molecule on human NK cells. This effect appeared to be specific for NKRP1A as the expression of other functional NK cell surface molecules such as CD16 and different killer inhibitory receptors (KIR) including CD158a and CD158b, p70 and p140 were not affected by culture in IL-12. In addition, we found that polyclonal or clonal NK cell populations derived in the presence of IL-2 displayed an increased expression of NKRP1A after culture in IL-12. The IL-12-induced NKRP1A expression was time and dose dependent, reaching a maximum by 7 days of culture in the presence of 2 ng/ml IL-12 and it was inhibited by the addition of anti-IL-12 monoclonal antibody. The IL-12-dependent NKRP1A up-regulation was abrogated by the incubation of NK cells with actinomycin D, thus suggesting that IL-12 induces de novo transcription of NKRP1A mRNA. Functional analysis revealed that the engagement of the NKRP1A molecule in IL-12- but not in IL-2-cultured NK cells leads to a strong inhibition of the cytolytic activity induced by cross-linking of CD16 or p46, a recently described NK cell-specific triggering surface molecule. Our findings suggest that IL-12 up-regulates the expression of NKRP1A which, in turn, can regulate NK cell activation induced via different triggering pathways. This would imply that NKRP1A-mediated functions may be regulatd by the cytokine microenvironment that NK cells may encounter at inflammatory sites.

Antigens, Surface↗
PubMed1998

p49, a putative HLA class I-specific inhibitory NK receptor belonging to the immunoglobulin superfamily.

NK cells display several killer inhibitory receptors (KIR) specific for different alleles of MHC class I molecules. A family of KIR are represented by type I transmembrane proteins belonging to the immunoglobulin superfamily (Ig-SF). Besides cDNA encoding for these KIR, additional cDNA have been identified which encode for Ig-SF receptors with still undefined specificity. Here we analyze one of these cDNA, termed cl.15.212, which encodes a type I transmembrane protein characterized by two extracellular Ig-like domains and a 115-amino acid cytoplasmic tail containing a single immuno-receptor tyrosine-based inhibitory motif (ITIM) which is typical of KIR. cl.15.212 cDNA displays approximately 50 % sequence homology with other Ig-SF members. Different from the other KIR, cl.15.212 mRNA is expressed by all NK cells and by a fraction of KIR+ T cell clones. cl.15.212 cDNA codes for a membrane-bound receptor displaying an apparent molecular mass of 49 kDa, thus termed p49. To determine the specificity of the cl.15.212-encoded receptor, we generated soluble fusion proteins consisting of the ectodomain of p49 and the Fc portion of human IgG1. Soluble molecules bound efficiently to 221 cells transfected with HLA-G1, -A3, -B46 alleles and weakly to -B7 allele. On the other hand, they did not bind to 221 cells either untransfected or transfected with HLA-A2, -B51, -Cw3 or -Cw4. The binding specificity of soluble p49-Fc was confirmed by competition experiments using an anti-HLA class I-specific monoclonal antibody. Finally, different cDNA encoding for molecules homologous to cl.15.212 cDNA have been isolated, two of which lack the sequence encoding the transmembrane portion, thus suggesting they may encode soluble molecules.

Alleles↗
PubMed1998

p40/LAIR-1 regulates the differentiation of peripheral blood precursors to dendritic cells induced by granulocyte-monocyte colony-stimulating factor.

p40/LAIR-1, a member of the immunoglobulin superfamily, is a surface molecule broadly distributed among leukocytes which has been shown to down-regulate T and NK cell activation. In this study, we show that p40/LAIR-1 is highly expressed in CD14+ peripheral blood mononuclear cells (PBMC). When cultured in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF) for 10-14 days, CD14+ cells acquired morphologic and phenotypic features (i.e. loss of CD14 and expression of CD80bright and CD86bright) typical of dendritic cells (DC) and lost the expression of p40/LAIR-1. Engagement of p40/LAIR-1 (but not of CD58) by specific monoclonal antibodies prevented CD14+ PBMC differentiation into DC; when cultured in the presence of GM- CSF upon p40/LAIR-1 cross-linking, the resulting cells were CD14+CD80(dull)CD86(dull) and displayed a macrophage-like morphology. We have recently demonstrated that peripheral blood CD14+ cells co-expressing the CD34 progenitor marker represent the circulating precursors of CD83+ DC. Herein we show that cross-linking of p40/LAIR-1 prevented the maturation of CD14+CD34+ cells into CD83+ DC. This effect appears to be consequent to the impairment of GM-CSF receptor-mediated activation signaling. Indeed, triggering of GM-CSF receptors in both CD14+ and CD14+CD34+ cells led to increases in the intracellular free calcium concentrations which were inhibited by p40/LAIR-1 engagement. Taken together, these data suggest a possible regulating role played by p40/LAIR-1 in the process of differentiation from peripheral blood precursors into DC induced by GM-CSF.

Calcium↗
PubMed1998

The susceptibility to natural killer cell-mediated lysis of HLA class I-positive melanomas reflects the expression of insufficient amounts of different HLA class I alleles.

NK cells selectively lyse tumor cells which do not express one or more MHC class I alleles. The ability to discriminate between self normal or tumor cells is due to the expression of MHC class I-specific killer inhibitory receptors (KIR). In the present study we analyzed melanoma cell lines which were highly susceptible to NK cell-mediated lysis in spite of the expression of a complete set of HLA class I alleles. Quantitative analysis of the HLA class I expression using allele-specific monoclonal antibodies (mAb) revealed a down-regulation of all HLA class I molecules. Treatment of melanoma cells with IFN-gamma resulted in up-regulation of all HLA class I alleles that was paralleled by the acquisition of resistance to lysis. That resistance to lysis reflected the up-regulation of HLA class I molecules was revealed by the finding that mAb-mediated masking of either KIR or their HLA class I ligands completely restored the melanoma cell lysis. These results were obtained by the use of selected NK cell clones derived either from allogeneic or autologous donors. In addition, similar results were obtained using in vitro expanded autologous NK cell populations. Our data indicate that NK cells can lyse not only melanoma cells which have lost the expression of one or more HLA class I alleles but also cells expressing a decreased amount of class I molecules.

Alleles↗
PubMed1998

Regulation of KIR expression in human T cells: a safety mechanism that may impair protective T-cell responses.

Killer-cell inhibitory Receptors (KIRs) are a new family of major histocompatibility complex (MHC) class I-specific receptors. KIRs allow natural killer cells to identify and lyse self cells that do not express sufficient amounts of MHC class I molecules. Here, Maria Cristina Mingari and colleagues view the expression of KIRs by cytolytic T lymphocytes and their regulation by certain cytokines as a double-edged sword.

Gene Expression Regulation↗
PubMed1998

Cytokine-induced expression of killer inhibitory receptors in human T lymphocytes.

Killer inhibitory receptors (KIRs) represent a new family of HLA-class I-specific receptors. KIRs are involved in the function of Natural Killer cells and allow these cells to discriminate between normal cells and cells with impaired expression of HLA-class I molecules. KIRs are also expressed by a subset of cytolytic T lymphocytes in which they may exert an inhibitory effect on TCR-mediated function. Here we review recent data indicating that cytokines such as IL-15, may induce the de novo expression of CD94/NKG2A (a KIR which operationally detects the expression of various HLA-class I alleles). The expression of CD94/NKG2A has been documented not only in CD34+ precursors undergoing maturation towards NK cells, but also in mature T cells which respond in vitro to superantigens or allogeneic cells.

Animals↗
PubMed1997

p46, a novel natural killer cell-specific surface molecule that mediates cell activation.

Limited information is available on the surface molecules that are involved in natural killer (NK) cell triggering. In this study, we selected the BAB281 monoclonal antibody (mAb) on the basis of its ability to trigger NK-mediated target cell lysis. BAB281 identified a novel NK cell-specific surface molecule of 46 kD (p46) that is expressed by all resting or activated NK cells. Importantly, unlike the NK cell antigens identified so far, the expression of p46 was strictly confined to NK cells. Upon mAb-mediated cross-linking, p46 molecules induced strong cell triggering leading to [Ca2+]i increases, lymphokine production, and cytolytic activity both in resting NK cells and NK cell clones. The p46-mediated induction of Ca2+ increases or triggering of cytolytic activity was downregulated by the simultaneous engagement of inhibitory receptors including p58, p70, and CD94/NKG2A. Both the unique cellular distribution and functional capability of p46 molecules suggest a possible role in the mechanisms of non-major histocompatibility complex-restricted cytolysis mediated by human NK cells.

Amidohydrolases↗
PubMed1997

Expression of HLA class I-specific inhibitory natural killer cell receptors in HIV-specific cytolytic T lymphocytes: impairment of specific cytolytic functions.

Human T lymphocytes have been shown to express inhibitory natural killer cell receptors (NKR), which can down-regulate T cell antigen receptor-mediated T cell function, including cytolytic activity. In the present study, we demonstrate that CD3+NKR+ cells can be identified in HIV-infected patients. HIV-specific cytolytic activity was analyzed in five patients in whom autologous lymphoblastoid B cell lines could be derived as a source of autologous target cells. Phytohemagglutinin-activated T cell populations that had been cultured in interleukin 2 displayed HIV-specific cytotoxic T lymphocyte (CTL) activity against HIV env, gag, pol, and nef in 3 of 5 patients. Addition of anti-NKR mAb of IgM isotype could increase the specific CTL activity. Moreover, in one additional patient, HIV-specific CTL activity was undetectable; however, after addition of anti-NKR mAb such CTL activity appeared de novo. Similar results were obtained by analysis of CD3+NKR+ clones derived from two patients. These data provide direct evidence that CD3+NKR+ cells may include antigen (HIV)-specific CTLs and that mAb-mediated masking of inhibitory NKR may revert the down-regulation of CTL function.

Antibodies, Monoclonal↗
PubMed1997

Natural killer cell acceptance of H-2 mismatch bone marrow grafts in transgenic mice expressing HLA-Cw3 specific killer cell inhibitory receptor.

Natural killer (NK) cells express killer cell inhibitory receptors (KIRs) for major histocompatibility complex class I molecules. Engagement of these surface receptors inhibits NK cell cytotoxic programs. KIR can also be expressed on T cell subsets, and their engagement similarly results in inhibition of effector functions initiated by the CD3/T cell receptor complex. KIR genes belong to two distinct families: the immunoglobulin superfamily (IgSF KIRs) and dimeric C2 lectins (lectin-like KIRs). Whereas both IgSF (p58: CD158, p70, and p140) and lectin-like KIRs (CD94/NKG2A heterodimers) have been found in human, only lectin-like KIRs (all members of the Ly-49 family) have been described in the mouse. We have generated transgenic mice expressing an IgSF KIR, CD158b (p58.2), which recognizes HLA-Cw3. Our data show that CD158b is necessary and sufficient to confer specificity to NK cells, as well as to modulate T cell activation programs in vitro. In addition, we did not detect any adaptation of CD158b cell surface expression to that of HLA class I ligands in the CD158b x HLA-Cw3 double transgenic mice, in contrast to observations with Ly-49 in the mouse. Therefore, distinct strategies of selection/calibration appear to be used by IgSF and lectin-like KIRs. Finally, the transgenic expression of CD158b KIR prevents the in vivo rejection of H-2 mismatch bone marrow grafts, which express the cognate major histocompatibility class I HLA-Cw3 allele, demonstrating for the first time the in vivo implication of human IgSF KIRs in the negative regulation of NK cell function.

Animals↗
PubMed1997

NKRP1A molecule is involved in transendothelial migration of CD4+ human T lymphocytes.

Among human CD4+ T lymphocytes, 5-20% express the C-type lectin molecule NKRP1A. Interestingly, CD4+ NKRP1A+ T lymphocytes express high levels of beta 1 and beta 2 integrins, thus representing a T lymphocyte subset that can possibly adhere and migrate through vascular endothelium. Indeed, resting CD4+ NKRP1A+ lymphocytes, differently from the CD4+ NKRP1A- subset, migrated across endothelial cell monolayers in a Transwell chamber system. This transendothelial migration was strongly reduced after pre-treatment with an anti-NKRP1A monoclonal antibody (mAb). In addition, the NKRP1A negative Jurkatt CD4+ T-cell line that had been stably transfected with NKRP1A cDNA, migrated more rapidly and efficiently than untransfected Jurkatt cells. Finally, mAb-mediated cross-linking of NKRP1A molecule in CD4+ T lymphocytes induced the upregulation of the LFA1 Mg2+ binding site as well as beta 1 and beta 2 integrin chains. Altogether, these findings indicate that NKRP1A molecule is involved in transendothelial migration of resting CD4+ T lymphocytes.

Antigens, Surface↗
PubMed1997

Reconstituted killer cell inhibitory receptors for major histocompatibility complex class I molecules control mast cell activation induced via immunoreceptor tyrosine-based activation motifs.

Natural killer and T cells express at their surface, members of a multigenic family of killer cell inhibitory receptors (KIR) for major histocompatibility complex Class I molecules. KIR engagement leads to the inhibition of natural killer and T cell activation programs. We investigated here the functional reconstitution of KIR in a non-lymphoid cell type. Using stable transfection in the RBL-2H3 mast cell line, we demonstrated that (i) KIR can inhibit signals induced by FcepsilonRIgamma or CD3zeta polypeptides that bear immunoreceptor tyrosine-based activation motifs; (ii) two distinct immunoreceptor tyrosine-based inhibition motifs-bearing receptors, i.e. KIR and FcgammaRIIB, use distinct inhibitory pathways since KIR engagement inhibits the intracellular Ca2+ release from endoplasmic reticulum stores, in contrast to FcgammaRIIB, which only inhibits extracellular Ca2+ entry; (iii) KIR require co-ligation with an immunoreceptor tyrosine-based activation motif-dependent receptor to mediate their inhibitory function. This latter finding is central to the mechanism by which KIR selectively inhibit only the activatory receptors in close vicinity. Taken together our observations also contribute to define and extend the family of immunoreceptor tyrosine-based inhibition motif-bearing receptors involved in the negative control of cell activation.

Animals↗
PubMed1997

The CD94 and NKG2-A C-type lectins covalently assemble to form a natural killer cell inhibitory receptor for HLA class I molecules.

CD94, a type II membrane protein containing a C-type lectin domain, has been shown to be involved in natural killer (NK) cell-mediated recognition of different HLA allotypes. The inhibitory form of the CD94 receptor has recently been identified by the specific monoclonal antibody (mAb) Z199. Herein, we demonstrate that the inhibitory receptor is in fact a complex formed by the covalent association of CD94 with the NKG2-A molecule (Mr approximately 43 kDa), another member of the C-type lectin superfamily, and that Z199 mAb specifically recognize NKG2-A molecules. Although the NKG2-A-encoding cDNA has been known for several years, the corresponding protein and its possible function remained undefined. Moreover, we show that the NKG2-B protein, an alternatively spliced product of the NKG2-A gene, can also assemble with CD94. Remarkably, both NKG2-A and NKG2-B proteins contain cytoplasmic immunoreceptor tyrosine-based inhibitory motifs (ITIM). This may provide the molecular basis of the inhibitory function mediated by the CD94/NKG2-A receptor complexes.

Antigens, CD↗