RESUMO
Influenza is one of the most prevalent viral diseases in humans. For some high-risk human populations, including the infant, the elderly, and the immunocompromised, who may not benefit from active immunization, passive immunotherapy with antibodies reactive with all influenza A strains may be an alternative. In this study, we characterized several fully human monoclonal antibodies (MAb) reactive with M2e, which were generated from transchromosomic mice engineered to produce fully human antibodies following immunization with a consensus-sequence M2e peptide. The MAbs showed strong binding to M2e peptide and to virus infected MDCK cells. One MAb recognizing the highly conserved N-terminal portion of consensus M2e displayed high binding to the majority of M2e variants from natural viral isolates, including highly pathogenic avian strains, which were recently reported to infect humans. Passive immunotherapy with this MAb in mice resulted in significant reduction in virus replication in the lung and protection from lethal infection when administered either prophylactically or therapeutically. These results suggest the potential of the anti-M2e human MAb with broad binding spectrum as a universal passive immunotherapeutic agent to infection by influenza A virus.
Assuntos
Anticorpos Monoclonais/administração & dosagem , Imunização Passiva , Vírus da Influenza A/imunologia , Influenza Humana/prevenção & controle , Proteínas da Matriz Viral/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/administração & dosagem , Especificidade de Anticorpos , Linhagem Celular , Cães , Feminino , Humanos , Influenza Humana/imunologia , Influenza Humana/terapia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Dados de Sequência MolecularRESUMO
NK cells are capable of responding quickly to infectious challenge and contribute to the early defense against a wide variety of pathogens. Although the innate NK cell response to murine CMV (MCMV) has been extensively characterized, its resolution and the fate of the activated NK cell population remains unexplored. Herein, we characterize both the expansion and contraction phases of the NK cell response to MCMV. We demonstrate that NK cell recruitment into the immune response to MCMV infection is restricted to the first 3 days of infection and as the peripheral NK cell compartment expands, NK cells undergo accelerated phenotypic maturation. During the resolution of the immune response, NK cell compartmental contraction is marked by the selective death of responding NK cells. Additionally, throughout the infection, a naive NK cell pool that remains responsive to additional stimuli is actively maintained. These findings illustrate the plasticity of the NK cell compartment in response to pathogens and underscore the homeostatic maintenance of the resting peripheral NK cell pool.
Assuntos
Infecções por Citomegalovirus/imunologia , Células Matadoras Naturais/fisiologia , Muromegalovirus , Animais , Antígeno CD11b/análise , Movimento Celular , Interferon gama/biossíntese , Lectinas Tipo C , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptores Imunológicos/análiseRESUMO
Changes in Valpha24+Vbeta11+ NKT cell number and function are associated with human autoimmune diseases and cancer. Restoration of this corresponding NKT cell population in mice or in vivo activation with alpha-galactosylceramide (KRN7000) can prevent or reduce tumor growth and autoimmunity. Although the therapeutic value of these natural killer T (NKT) cells in man remains to be determined, large numbers of functional antigen-specific NKT cells can be expanded in vitro. We show that Valpha24+Vbeta11+ human NKT cells are expanded by repeated stimulation with KRN7000, unfractionated donor peripheral blood mononuclear cells (PBMC), and recombinant human interleukin-2 (rhIL-2). NKT cells were expanded continuously for more than 2 months with a potential yield of >10(12) cells. The expanded NKT cells retained their CD4+ or CD4- phenotype after restimulation and were functional as shown by cytokine secretion, killing of antigen-pulsed target cells, and activation of NK cell cytotoxicity. This expansion method may be useful for proof-of-concept studies involving adoptive transfer of ex vivo-expanded NKT cells as a new therapeutic option for cancer and autoimmune diseases.
Assuntos
Adjuvantes Imunológicos/farmacologia , Galactosilceramidas/farmacologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Ativação Linfocitária/imunologia , Antígenos CD4/metabolismo , Antígenos CD8/metabolismo , Células Cultivadas , Citotoxicidade Imunológica , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Substâncias de Crescimento/análise , Substâncias de Crescimento/farmacologia , Humanos , Interleucina-2/análise , Interleucina-2/farmacologia , Células Jurkat , Receptores de Antígenos de Linfócitos T/imunologia , Proteínas Recombinantes/farmacologiaRESUMO
The killer cell lectin-like receptor G1 (KLRG1) is the mouse homologue of the rat mast cell function-associated Ag and contains a tyrosine-based inhibitory motif in its cytoplasmic domain. It has been demonstrated that KLRG1 is induced on activated NK cells and that KLRG1 can inhibit NK cell effector functions. In this study, we show that in naive C57BL/6 mice KLRG1 is expressed on a subset of CD44(high)CD62L(low) T cells. KLRG1 expression can be detected on a small number of V(alpha)14i NK T cells but not on CD8alphaalpha(+) intraepithelial T cells that are either TCRgammadelta(+) or TCRalphabeta(+). We also show that KLRG1 expression is dramatically induced on approximately 50% of the CD8(+) T cells during both a viral and a parasitic infection. Interestingly, during Toxoplasma gondii infection, KLRG1 is up-regulated on CD4(+) T cells. Although KLRG1 expression can be induced on both NK cells and T cells, the molecular mechanism leading to the induction of KLRG1 differs in these two subsets of cells. Indeed, the up-regulation of KLRG1 on NK cells can be driven in vivo by cytokines, whereas KLRG1 cannot be induced on CD8(+) T cells by cytokines. In addition, although induction of KLRG1 on T cells appears to require TCR engagement in vivo, TCR engagement is not sufficient for KLRG1 induction in vitro. Taken together, these data suggest that the expression and induction of KLRG1 on T cells are tightly regulated. This could have important biological consequences on T cell activation and homeostasis.
Assuntos
Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Receptores Imunológicos/biossíntese , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Doença Aguda , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Antígenos CD8/biossíntese , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/patologia , Linfócitos T CD8-Positivos/virologia , Divisão Celular/imunologia , Separação Celular , Citocinas/fisiologia , Infecções por Citomegalovirus/imunologia , Infecções por Citomegalovirus/metabolismo , Infecções por Citomegalovirus/patologia , Receptores de Hialuronatos/biossíntese , Memória Imunológica , Interferon gama/biossíntese , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Selectina L/biossíntese , Lectinas Tipo C/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T alfa-beta/biossíntese , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Toxoplasmose Animal/imunologia , Toxoplasmose Animal/metabolismoRESUMO
BACKGROUND & AIMS: Natural killer T (NKT) cells are a subset of lymphocytes incriminated in playing an important role in the modulation of the innate immune response and the development of autoimmunity. However, there have been only limited studies attempting to quantitate the number of NKT cells in autoimmune disease, particularly because of difficulties associated with definition of this subpopulation. METHODS: We used a human CD1d (hCD1d) tetramer produced by a baculovirus expressing recombinant CD1d protein complexed with alpha-galactosylceramide (alpha-GalCer) and quantitated hCD1d tetramer reactive cells in blood and liver from controls and patients with primary biliary cirrhosis (PBC). RESULTS: The majority of CD1d-alphaGalCer-restricted NKT cells were positive for TCR Valpha24 and Vbeta11. There was a distinct CD4- CD8+ population within the CD1d-alphaGalCer-restricted NKT cells in addition to the CD4- CD8- and CD4+ CD8- population. The frequency of CD1d-alphaGalCer-restricted NKT cells was similar between blood and liver in healthy individuals. In contrast, the frequency of CD1d-alphaGalCer-restricted NKT cells in the liver was significantly higher than in the blood of PBC patients. The frequency of CD1d-alpha-GalCer-restricted NKT cells in the liver was also significantly higher in PBC patients than in healthy individuals. CONCLUSIONS: The frequency and function of such cells should be studied not only in blood but also in the target organ of the autoimmune disease. Selective enrichment of CD1d-alphaGalCer-restricted NKT cells at the site of inflammation is observed in PBC, suggesting a role of these cells in the development of PBC.
Assuntos
Antígenos CD1/genética , Células Matadoras Naturais/imunologia , Cirrose Hepática Biliar/imunologia , Antígenos CD1d , Baculoviridae/genética , Citometria de Fluxo , Galactosilceramidas/imunologia , Humanos , Imunofenotipagem , Células Matadoras Naturais/química , Fígado/citologia , Fígado/imunologia , Linfonodos/citologia , Linfonodos/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/análise , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologiaRESUMO
The killer cell lectin-like receptor G1 (KLRG1) is the mouse homolog of the rat mast cell function-associated Ag and contains an immunoreceptor tyrosine-based inhibitory motif in its cytoplasmic domain. In this study we demonstrate that both pathogenic and nonpathogenic in vivo activation of NK cells induces the expression of KLRG1 on their cell surface. Upon infection with murine CMV, this induction peaks between days 5 and 7 with about 90% of the NK cells expressing KLRG1. On day 1.5 post-murine CMV infection of C57BL/6 mice, the main producers of IFN-gamma are the KLRG1-negative NK cells. This effect has been recapitulated in vitro as we show that engagement of KLRG1 on a transfected NK cell line inhibits both cytokine production and NK cell-mediated cytotoxicity. Taken together, these data illustrate the crucial role played by KLRG1 during the termination of mouse NK cell activation.