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Biomedical subjects

A Moretta

Publications and source records attributed to A Moretta.

At least 37 records · Page 2Linked to original sources

Human natural killer cell function and receptors.

Recent years have witnessed major progress in our understanding of the molecular mechanisms regulating natural killer cell (NK cell) function. These advances stem primarily from the discovery of a number of receptors specific for major histocompatibility complex (MHC) class I and, more recently, of the activating receptors and coreceptors responsible for natural cytotoxicity. Important studies performed over the past year have allowed us to define the evolution of the MHC-specific inhibitory receptors by comparative analysis in different species. The roles of the 'activating natural cytotoxicity receptors', NKG2D and certain coreceptors in the lysis of different tumors have been defined in detail. The mechanism by which the 2B4 coreceptor renders patients with X-linked lymphoproliferative disease unable to control Epstein-Barr virus has been elucidated. Inhibitory receptors identified in NK cells may also be expressed by normal and leukemic myeloid cells, in which they can block cell proliferation and survival. It has also become clear that viruses such as cytomegalovirus have evolved strategies to interfere with NK-cell function to protect themselves from NK-mediated attack.

Cytotoxicity, Immunologic↗

Human natural killer cell receptors and co-receptors.

In the absence of sufficient signaling by their HLA class I-specific inhibitory receptors, human natural killer (NK) cells become activated and display potent cytotoxicity against cells that are either HLA class I negative or deficient. This indicates that the NK receptors responsible for the induction of cytotoxicity recognize ligands on target cells different from HLA class I molecules. On this basis, the process of NK-cell triggering can be considered as a mainly non-MHC-restricted mechanism. The recent identification of a group of NK-specific triggering surface molecules has allowed a first series of pioneering studies on the functional/molecular characteristics of such receptors. The first three members of a receptor family that has been termed natural cytotoxicity receptors (NCR) are represented by NKp46, NKp44 and NKp30. These receptors are strictly confined to NK cells, and their engagement induces a strong activation of NK-mediated cytolysis. A direct correlation exists between the surface density of NCR and the ability of NK cells to kill various target cells. Importantly, mAb-mediated blocking of these receptors has been shown to suppress cytotoxicity against most NK-susceptible target cells. However, the process of NK-cell triggering during target cell lysis may also depend on the concerted action of NCR and other triggering receptors, such as NKG2D, or surface molecules, including 2B4 and NKp80, that appear to function as co-receptors rather than as true receptors. Notably, a dysfunction of 2B4 has been associated with a severe form of immunodeficiency termed X-linked lymphoproliferative disease. Future studies will clarify whether also the altered expression and/or function of other NK-triggering molecules may represent a possible cause of immunological disorders.

Cytotoxicity, Immunologic↗

Human alloantigen-specific anergic cells induced by a combination of CTLA4-Ig and CsA maintain anti-leukemia and anti-viral cytotoxic responses.

The success of allogeneic hematopoietic stem cell transplantation from HLA-disparate donors depends on the development of new strategies for graft-versus-host disease prevention able to target specifically donor antihost alloreactivity, while preserving GVL and antiviral immune surveillance. Recent experimental and clinical work has shown the feasibility of an approach based on induction of anergy to host alloantigens through blockade of B7/CD28 costimulatory signal in donor T cells, but data on the impact of this strategy on the recovery of the immune system are still lacking. We devised an ex vivo method for induction of host alloantigen-specific unresponsiveness based on treatment with the B7/CD28 blocking agent CTLA4-Ig associated with CsA. We then proceeded to assess the maintenance of an effective immune response towards viral pathogens and tumor cells after CTLA4-Ig/CsA treatment, by measuring the frequency of CTL precursors directed against CMV- and EBV-infected targets, and against autologous leukemic blasts. We demonstrated that (1) CTLA4-Ig and CsA can act synergistically in inducing a state of unresponsiveness to alloantigens; (2) the number of leukemia-reactive, EBV-specific and CMV-specific CTLp is not impaired by CTLA4-Ig/CsA treatment. Our data provide the first direct in vitro evidence that it is possible to preserve antiviral and antileukemia effector cells after blockade of CD28/B7 interaction during MLR.

Abatacept↗

Cellular and molecular pathogenesis of X-linked lymphoproliferative disease.

In the past few years important advances have been made in the understanding of the molecular mechanisms leading to X-linked lymphoproliferative disease (XLP). It has been possible to identify the gene defective in XLP and to demonstrate that, in normal individuals, it encodes the Src homology 2 domain-containing protein 1 A (SH2D1A) that plays a crucial role in signaling via a number of surface molecules expressed by cells of the immune system. At present a variety of mutations have been identified that result in lack of defective SH2D1A molecules. Importantly, it has recently been demonstrated that lack of SH2D1A affects the function of surface molecules that are involved in the mechanism of natural killer-mediated recognition and killing of Epstein-Barr virus-infected B cells.

Animals↗

Activating receptors and coreceptors involved in human natural killer cell-mediated cytolysis.

Natural killer cells can discriminate between normal cells and cells that do not express adequate amounts of major histocompatibility complex (MHC) class I molecules. The discovery, both in mouse and in human, of MHC-specific inhibitory receptors clarified the molecular basis of this important NK cell function. However, the triggering receptors responsible for positive NK cell stimulation remained elusive until recently. Some of these receptors have now been identified in humans, thus shedding some light on the molecular mechanisms involved in NK cell activation during the process of natural cytotoxicity. Three novel, NK-specific, triggering surface molecules (NKp46, NKp30, and NKp44) have been identified. They represent the first members of a novel emerging group of receptors collectively termed natural cytotoxicity receptors (NCR). Monoclonal antibodies (mAbs) to NCR block to differing extents the NK-mediated lysis of various tumors. Moreover, lysis of certain tumors can be virtually abrogated by the simultaneous masking of the three NCRs. There is a coordinated surface expression of the three NCRs, their surface density varying in different individuals and also in the NK cells isolated from a given individual. A direct correlation exists between the surface density of NCR and the ability of NK cells to kill various tumors. NKp46 is the only NCR involved in human NK-mediated killing of murine target cells. Accordingly, a homologue of NKp46 has been detected in mouse. Molecular cloning of NCR revealed novel members of the Ig superfamily displaying a low degree of similarity to each other and to known human molecules. NCRs are coupled to different signal transducing adaptor proteins, including CD3 zeta, Fc epsilon RI gamma, and KARAP/DAP12. Another triggering NK receptor is NKG2D. It appears to play either a complementary or a synergistic role with NCRs. Thus, the triggering of NK cells in the process of tumor cell lysis may often depend on the concerted action of NCR and NKG2D. In some instances, however, it may uniquely depend upon the activity of NCR or NKG2D only. Strict NKG2D-dependency can be appreciated using clones that, in spite of their NCR(dull) phenotype, efficiently lyse certain epithelial tumors or leukemic cell lines. Other triggering surface molecules including 2B4 and the novel NKp80 appear to function as coreceptors rather than as true receptors. Indeed, they can induce natural cytotoxicity only when co-engaged with a triggering receptor. While an altered expression or function of NCR or NKG2D is being explored as a possible cause of immunological disorders, 2B4 dysfunction has already been associated with a severe form of immunodeficiency. Indeed, in patients with the X-linked lymphoproliferative disease, the inability to control Epstein-Barr virus infections may be consequent to a major dysfunction of 2B4 that exerts inhibitory instead of activating functions.

Adaptor Proteins, Signal Transducing↗

X-linked lymphoproliferative disease. 2B4 molecules displaying inhibitory rather than activating function are responsible for the inability of natural killer cells to kill Epstein-Barr virus-infected cells.

2B4 is a surface molecule involved in activation of the natural killer (NK) cell-mediated cytotoxicity. It binds a protein termed Src homology 2 domain-containing protein (SH2D1A) or signaling lymphocyte activation molecule (SLAM)-associated protein (SAP), which in turn has been proposed to function as a regulator of the 2B4-associated signal transduction pathway. In this study, we analyzed patients with X-linked lymphoproliferative disease (XLP), a severe inherited immunodeficiency characterized by critical mutations in the SH2D1A gene and by the inability to control Epstein-Barr virus (EBV) infections. We show that, in these patients, 2B4 not only fails to transduce triggering signals, but also mediates a sharp inhibition of the NK-mediated cytolysis. Other receptors involved in NK cell triggering, including CD16, NKp46, NKp44, and NKp30, displayed a normal functional capability. However, their activating function was inhibited upon engagement of 2B4 molecules. CD48, the natural ligand of 2B4, is highly expressed on the surface of EBV(+) B cell lines. Remarkably, NK cells from XLP patients could not kill EBV(+) B cell lines. This failure was found to be the consequence of inhibitory signals generated by the interaction between 2B4 and CD48, as the antibody-mediated disruption of the 2B4-CD48 interaction restored lysis of EBV(+) target cells lacking human histocompatibility leukocyte antigen (HLA) class I molecules. In the case of autologous or allogeneic (HLA class I(+)) EBV(+) lymphoblastoid cell lines, restoration of lysis was achieved only by the simultaneous disruption of 2B4-CD48 and NK receptor-HLA class I interactions. Molecular analysis revealed that 2B4 molecules isolated from either XLP or normal NK cells were identical. As expected, in XLP-NK cells, 2B4 did not associate with SH2D1A, whereas similar to 2B4 molecules isolated from normal NK cells, it did associate with Src homology 2 domain-containing phosphatase 1.

Antigens, CD↗

Involvement of natural cytotoxicity receptors in human natural killer cell-mediated lysis of neuroblastoma and glioblastoma cell lines.

The surface receptors involved in natural killer (NK) cell triggering during the process of target cell lysis have been at least in part identified. These are members of a novel family of receptors that has been termed natural cytotoxicity receptors (NCR). The first three members of this emerging group of receptors are the NKp46, NKp44 and NKp30 molecules that all belong to the immunoglobulin superfamily. Blocking of these receptors inhibits NK-mediated cytotoxicity against a wide variety of tumor target cells. In the present study, we show that these NCR are also involved in NK-mediated killing of tumor cells of neural origin. Glioblastoma and neuroblastoma target cells were efficiently killed by all NK clones analyzed since little protection from NK lysis was mediated by HLA class I molecules. Blocking of one or another NCR inhibited cytotoxicity; however, optimal inhibition was only observed when the three receptors were blocked simultaneously. A sharp difference in cytotoxicity against neural tumors was demonstrated between NCR(bright) and NCR(dull) NK clones, further supporting the notion that NCR play a critical role in the induction of cytotoxicity against tumor target cells of different histotype. Finally, our data also indicate that CD16 does not function as a triggering receptor involved in lysis of neural tumors since no difference in cytotoxicity could be substantiated between CD16(+) and CD16(-) NK clones and no correlation could be detected between the NCR(bright)/NCR(dull) phenotype and CD16 expression.

Antibodies, Monoclonal↗

Killer inhibitory receptor (CD158b) modulates the lytic activity of tumor-specific T lymphocytes infiltrating renal cell carcinomas.

In this study, we showed that renal tumors contain substantial subsets of CD8(+) p58(+) T cells. From 1 of these tumors, T cells were amplified in mixed lymphocytes-tumor cell cultures and p58(+) T cells were selected immunologically. After expansion, phenotypic and functional features of p58(+) and p58(-) T cells were examined. The p58(+) T cells expressed p58.2 receptor and corresponded to CD3(+), CD8(+), T-cell receptor (TCR) alpha/beta(+) T cells that were CD56(+) and CD28(-). Functionally, p58(+) T cells showed a low level of lytic activity against autologous tumor cells that was dramatically and specifically increased by anti-p58.2 monoclonal antibody. On the other hand, p58(-) CD8(+) T cells did not lyse autologous tumor cells and had non-major histocompatibility complex-restricted cytotoxicity against K562 and Daudi cells. A p58(+) cytotoxic T lymphocyte (CTL) clone (4C7) with the same characteristics as the p58(+) T-cell line was derived. This CTL clone did not lyse autologous normal B cells but lysed several HLA-A1(+) renal tumor cell lines. Analysis of TCR repertoire diversity showed that the p58(+) T-cell line contained 3 TCR rearrangements, whereas the TCR repertoire of p58(-) T cells was polyclonal. Interestingly, TCR transcripts of p58(+) T cells and of CTL clone 4C7 were detected as prominent ex vivo in tumor cells but not in peripheral blood mononuclear cells, suggesting that these cells are antigen specific and amplified at the tumor site. (Blood. 2000;95:2883-2889)

Antigens, CD↗

2B4 functions as a co-receptor in human NK cell activation.

Natural cytotoxicity receptors (NKp46, NKp44 and NKp300) play a predominant role in human NK cell triggering during natural cytotoxicity. Human 2B4 also induced NK cell activation in redirected killing assays using anti-2B4 monoclonal antibodies (mAb) and murine targets. Since this effect was confined to a fraction of NK cells, this suggested a functional heterogeneity of 2B4 molecules. Here we show that activation via 2B4 in redirected killing against murine targets is strictly dependent upon the engagement of NKp46 by murine ligand (s) on target cells. Thus, NK cell clones expressing high surface density of NKp46 (NKp46bright) were triggered by anti-2B4 mAb, whereas NKp46dull clones were not although they expressed a comparable surface density of 2B4. mAb-mediated modulation of NKp46 molecules in NKp46bright clones had no effect on the expression of 2B4 while it rendered cells unresponsive to anti-2B4 mAb. Finally, anti-2B4 mAb could induce NK cell triggering in NKp46dull clones provided that suboptimal doses of anti-NKp44 or anti-CD16 mAb were added to the redirected killing assay. These results indicate that differences in responses do not reflect a functional heterogeneity of 2B4 but rather depend on the co-engagement of triggering receptors.

Animals↗

Neonatal invariant Valpha24+ NKT lymphocytes are activated memory cells.

NKT cells are a small subset of T lymphocytes which express an invariant V(alpha24JalphaQ TCR and recognize glycolipids presented by CD1d. In adults, NKT cells have a memory phenotype, frequently associated with oligoclonal expansion, express NK cell markers, and produce TO cytokines upon primary stimulation. Because of these features, NKT cells are regarded as lymphocytes of innate immunity. We investigated NKT cells from cord blood to see how these cells appear in the absence of exogenous stimuli. We found that NKT cells are present at comparable frequencies in cord blood and adult peripheral blood mononuclear cells and in both cases display a memory (CD45RO+CD62L-) phenotype. However, neonatal NKT cells differ from their adult counterparts by the following characteristics: (1) they express markers of activation, such as CD25; (2) they are polyclonal; (3) they do not produce cytokines in response to primary stimulation. Together, our data show that human NKT cells arise in the newborn with an activated memory phenotype, probably due to recognition of an endogenous ligand(s). The absence of oligoclonal expansion and primary effector functions also suggest that neonatal NKT cells, despite their activated memory phenotype, require a further priming/differentiation event to behave as fully functional cells of innate immunity.

Adult↗

Identification and molecular characterization of a natural mutant of the p50.2/KIR2DS2 activating NK receptor that fails to mediate NK cell triggering.

P50/KIR2DS molecules represent the activating form of the HLA-C-specific inhibitory NK receptors. They are characterized, in the transmembrane portion, by a charged amino acid that is involved in coupling with signal-transducing adaptor polypeptides. In this study we identified a novel p50.2/KIR2DS2 surface molecule, isolated from NK cell clones derived from an otherwise normal donor, that was unable to transduce activating signals. Sequence analysis of the cDNA encoding this molecule revealed six non-conservative codon mutations in the exon coding for the putative transmembrane portion. Notably, one of such mutations involved the charged residue lysine thought to be important for the association with signal-transducing polypeptides. Indeed, co-transfection experiments revealed that this naturally occurring p50.2/KIR2DS2 mutant, termed Mp50.2, displayed a sharply reduced ability to associate with DAP12 polypeptides. These data provide the first in vivo demonstration of the crucial role played by the transmembrane region of p50.2 receptor molecules in the functional association with DAP12 adaptor molecules and in the process of activation of NK-mediated cytotoxicity.

Amino Acid Sequence↗

Analysis of the molecular mechanism involved in 2B4-mediated NK cell activation: evidence that human 2B4 is physically and functionally associated with the linker for activation of T cells.

While 2B4 is a well-known surface receptor involved in NK cell triggering and induction of cytotoxicity against CD48-positive target cells, little is known about the downstream events which lead to NK cell activation. In this study we show that, in normal human NK cells, 2B4 constitutively associates with the linker for activation of T cells (LAT). Antibody-mediated engagement of 2B4 resulted in tyrosine phosphorylation not only of 2B4 but also of the associated LAT molecules. Moreover, tyrosine phosphorylation of LAT led to the recruitment of intracytoplasmic signaling molecules including phospholipase Cgamma and Grb2. These data support the concept that 2B4 may mediate NK cell triggering via a LAT-dependent signaling pathway.

Antigens, CD↗

Surface receptors delivering opposite signals regulate the function of human NK cells.

The effector function of NK cells is regulated by a number of positive and negative signals. The receptors involved in the negative regulation of NK cells are known since several years: they are represented by different families of MHC-specific receptors characterized by ITIM sequences in their cytoplasmic tail. In contrast, the surface molecules responsible for NK cell activation in the process of natural cytotoxicity and tumor cell lysis have remained elusive until recently. Three distinct NK-specific molecules termed 'natural cytotoxicity receptors' (NCR) have recently been identified and cloned. They play a complementary role in mediating NK cell activation in the interaction and lysis of most tumor cells. They are associated with different ITAM-containing adaptor proteins mediating signal transduction. Molecular cloning revealed novel members of the Ig superfamily displaying a limited homology with known human receptors.

Cytotoxicity, Immunologic↗

Selective expansion of intraepithelial lymphocytes expressing the HLA-E-specific natural killer receptor CD94 in celiac disease.

BACKGROUND & AIMS: Celiac disease is a gluten-induced enteropathy characterized by the presence of gliadin-specific CD4(+) T cells in the lamina propria and by a prominent intraepithelial T-cell infiltration of unknown mechanism. The aim of this study was to characterize the subset(s) of intraepithelial lymphocytes (IELs) expanding during active celiac disease to provide insights into the mechanisms involved in their expansion. METHODS: Flow-cytometric analysis of isolated IELs and/or immunohistochemical staining of frozen sections were performed in 51 celiac patients and 50 controls with a panel of monoclonal antibodies against T-cell and natural killer (NK) receptors. In addition, in vitro studies were performed to identify candidate stimuli for NK receptor expression. RESULTS: In normal intestine, different proportions of IELs, which were mainly T cells, expressed the NK receptors CD94/NKG2, NKR-P1A, KIR2D/3D, NKp46, Pen5, or CD56. During the active phase of celiac disease, the frequency of CD94(+) IELs, which were mostly alphabeta T cells, was conspicuously increased over controls. In contrast, the expression of other NK markers was not modified. Furthermore, expression of CD94 could be selectively induced in vitro by T-cell receptor activation and/or interleukin 15, a cytokine produced by intestinal epithelial cells. CONCLUSIONS: The gut epithelium favors the development of T cells that express NK receptors. In active celiac disease, there is a specific and selective increase of IELs expressing CD94, the HLA-E-specific NK receptor that may be related to T-cell receptor activation and/or interleukin 15 secretion.

Adult↗

Human natural killer cell activating receptors.

Natural killer (NK) cells were poorly characterized until 10 years ago and few molecules expressed on their cell surface were known. Now the situation has changed dramatically, since a plethora of receptors characterized by opposite functions have been functionally and molecularly defined. NK cells express clonally distributed inhibitory receptors specific for different groups of HLA class I alleles, thus protecting normal cells from NK-mediated lysis. On the contrary, various activating receptors are involved in triggering of NK-mediated natural cytotoxicity. Their engagement induces human NK cells to kill target cells that are either HLA class I-negative or -deficient. Here a brief description of the activating receptors and coreceptor and of their ligand(s) is given.

Cytotoxicity, Immunologic↗

Natural cytotoxicity receptors that trigger human NK-cell-mediated cytolysis.

Natural killer (NK) cells can detect whether cells have undergone tumour transformation or viral infection. The discovery of specific inhibitory receptors for major histocompatibility complex class I molecules clarified the basis of this discrimination. However, the receptors responsible for NK-cell triggering in the process of natural cytotoxicity remained elusive until recently. Here, Alessandro Moretta and colleagues describe the identification and characterization of several such receptors.

Animals↗