RESUMEN
BACKGROUND: Cancer immunotherapy with monoclonal antibodies and chimeric antigen receptor (CAR) T cell therapies can benefit from selection of new targets with high levels of tumor specificity and from early assessments of efficacy and safety to derisk potential therapies. METHODS: Employing mass spectrometry, bioinformatics, immuno-mass spectrometry and CRISPR/Cas9 we identified the target of the tumor-specific SF-25 antibody. We engineered IgE and CAR T cell immunotherapies derived from the SF-25 clone and evaluated potential for cancer therapy. RESULTS: We identified the target of the SF-25 clone as the tumor-associated antigen SLC3A2, a cell surface protein with key roles in cancer metabolism. We generated IgE monoclonal antibody, and CAR T cell immunotherapies each recognizing SLC3A2. In concordance with preclinical and, more recently, clinical findings with the first-in-class IgE antibody MOv18 (recognizing the tumor-associated antigen Folate Receptor alpha), SF-25 IgE potentiated Fc-mediated effector functions against cancer cells in vitro and restricted human tumor xenograft growth in mice engrafted with human effector cells. The antibody did not trigger basophil activation in cancer patient blood ex vivo, suggesting failure to induce type I hypersensitivity, and supporting safe therapeutic administration. SLC3A2-specific CAR T cells demonstrated cytotoxicity against tumor cells, stimulated interferon-γ and interleukin-2 production in vitro. In vivo SLC3A2-specific CAR T cells significantly increased overall survival and reduced growth of subcutaneous PC3-LN3-luciferase xenografts. No weight loss, manifestations of cytokine release syndrome or graft-versus-host disease, were detected. CONCLUSIONS: These findings identify efficacious and potentially safe tumor-targeting of SLC3A2 with novel immune-activating antibody and genetically modified cell therapies.
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Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Inmunoglobulina E/metabolismo , Inmunoterapia/métodos , Receptores Quiméricos de Antígenos/inmunología , Animales , Humanos , RatonesRESUMEN
CD98, which is required for the rapid proliferation of both normal and cancer cells, and MET, the hepatocyte growth factor receptor, are potential targets for therapeutic antitumor Abs. In this study, we report that the antiproliferative activity of a prototype anti-CD98 Ab, UM7F8, is due to Ab-induced membrane-associated ring CH (MARCH) E3 ubiquitin ligase-mediated ubiquitination and downregulation of cell surface CD98. MARCH1-mediated ubiquitination of CD98 is required for UM7F8's capacity to reduce CD98 surface expression and its capacity to inhibit the proliferation of murine T cells. Similarly, CD98 ubiquitination is required for UM7F8's capacity to block the colony-forming ability of murine leukemia-initiating cells. To test the potential generality of the paradigm that MARCH E3 ligases can mediate the antiproliferative response to antitumor Abs, we examined the potential effects of MARCH proteins on responses to emibetuzumab, an anti-MET Ab currently in clinical trials for various cancers. We report that MET surface expression is reduced by MARCH1, 4, or 8-mediated ubiquitination and that emibetuzumab-induced MET ubiquitination contributes to its capacity to downregulate MET and inhibit human tumor cell proliferation. Thus, MARCH E3 ligases can act as cofactors for antitumor Abs that target cell surface proteins, suggesting that the MARCH protein repertoire of cells is a determinant of their response to such Abs.
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Antígenos de Neoplasias/metabolismo , Antineoplásicos Inmunológicos/farmacología , Cadena Pesada de la Proteína-1 Reguladora de Fusión/metabolismo , Neoplasias/tratamiento farmacológico , Ubiquitina-Proteína Ligasas/metabolismo , Animales , Anticuerpos Monoclonales Humanizados/farmacología , Anticuerpos Monoclonales Humanizados/uso terapéutico , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/inmunología , Antineoplásicos Inmunológicos/uso terapéutico , Proliferación Celular/efectos de los fármacos , Cadena Pesada de la Proteína-1 Reguladora de Fusión/antagonistas & inhibidores , Cadena Pesada de la Proteína-1 Reguladora de Fusión/genética , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Técnicas de Inactivación de Genes , Células HeLa , Humanos , Células Jurkat , Ratones , Ratones Noqueados , Neoplasias/inmunología , Neoplasias/patología , Proteolisis , Proteínas Proto-Oncogénicas c-met/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-met/genética , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Ubiquitina-Proteína Ligasas/genética , Ubiquitinación/inmunologíaRESUMEN
Chlamydia trachomatis infection of the human fallopian tubes can lead to damaging inflammation and scarring, ultimately resulting in infertility. To study the human cellular responses to chlamydial infection, researchers have frequently used transformed cell lines that can have limited translational relevance. We developed a primary human fallopian tube epithelial cell model based on a method previously established for culture of primary human bronchial epithelial cells. After protease digestion and physical dissociation of excised fallopian tubes, epithelial cell precursors were expanded in growth factor-containing medium. Expanded cells were cryopreserved to generate a biobank of cells from multiple donors and cultured at an air-liquid interface. Culture conditions stimulated cellular differentiation into polarized mucin-secreting and multiciliated cells, recapitulating the architecture of human fallopian tube epithelium. The polarized and differentiated cells were infected with a clinical isolate of C. trachomatis, and inclusions containing chlamydial developmental forms were visualized by fluorescence and electron microscopy. Apical secretions from infected cells contained increased amounts of proteins associated with chlamydial growth and replication, including transferrin receptor protein 1, the amino acid transporters SLC3A2 and SLC1A5, and the T-cell chemoattractants CXCL10, CXCL11, and RANTES. Flow cytometry revealed that chlamydial infection induced cell surface expression of T-cell homing and activation proteins, including ICAM-1, VCAM-1, HLA class I and II, and interferon gamma receptor. This human fallopian tube epithelial cell culture model is an important tool with translational potential for studying cellular responses to Chlamydia and other sexually transmitted pathogens.
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Células Epiteliales/inmunología , Regulación de la Expresión Génica/inmunología , Interacciones Microbiota-Huesped/inmunología , Linfocitos T/inmunología , Adulto , Sistema de Transporte de Aminoácidos ASC/genética , Sistema de Transporte de Aminoácidos ASC/inmunología , Antígenos CD/genética , Antígenos CD/inmunología , Biomarcadores/metabolismo , Quimiocina CCL5/genética , Quimiocina CCL5/inmunología , Quimiocina CXCL10/genética , Quimiocina CXCL10/inmunología , Quimiocina CXCL11/genética , Quimiocina CXCL11/inmunología , Infecciones por Chlamydia/genética , Infecciones por Chlamydia/inmunología , Infecciones por Chlamydia/microbiología , Chlamydia trachomatis/crecimiento & desarrollo , Chlamydia trachomatis/inmunología , Células Epiteliales/microbiología , Trompas Uterinas/citología , Trompas Uterinas/cirugía , Femenino , Cadena Pesada de la Proteína-1 Reguladora de Fusión/genética , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Antígenos de Histocompatibilidad Clase I/genética , Antígenos de Histocompatibilidad Clase I/inmunología , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/inmunología , Interacciones Microbiota-Huesped/genética , Humanos , Molécula 1 de Adhesión Intercelular/genética , Molécula 1 de Adhesión Intercelular/inmunología , Antígenos de Histocompatibilidad Menor/genética , Antígenos de Histocompatibilidad Menor/inmunología , Modelos Biológicos , Cultivo Primario de Células , Receptores de Interferón/genética , Receptores de Interferón/inmunología , Receptores de Transferrina/genética , Receptores de Transferrina/inmunología , Salpingectomía , Linfocitos T/microbiología , Molécula 1 de Adhesión Celular Vascular/genética , Molécula 1 de Adhesión Celular Vascular/inmunología , Receptor de Interferón gammaRESUMEN
OBJECTIVE: CD98 regulates integrin signaling and is critical for tumor cell proliferation. It is also expressed on endothelial cells (EC), but its role in angiogenesis is unclear. APPROACH AND RESULTS: We used specific genetic targeting and antibody blockade approaches to examine the function of CD98 in EC proliferation, blood vessel growth, and tumor angiogenesis. It is upregulated on angiogenic ECs, and EC-specific deletion of CD98 in mice inhibited tumor growth, retinal angiogenesis, and EC proliferation. Reconstitution with CD98 mutants showed that integrin and CD98 interaction is necessary for EC survival and growth. Moreover, anti-CD98 treatment inhibited vessel formation and reversed EC-assisted tumor growth. CONCLUSIONS: Our findings demonstrate a requirement for CD98 in EC growth and suggest that CD98-specific reagents could have a dual anticancer effect: directly by inhibiting tumor cell proliferation and indirectly by preventing tumor angiogenesis.
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Carcinoma Pulmonar de Lewis/metabolismo , Cadena Pesada de la Proteína-1 Reguladora de Fusión/metabolismo , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Melanoma Experimental/metabolismo , Neovascularización Patológica , Neovascularización Fisiológica , Neovascularización Retiniana/metabolismo , Inhibidores de la Angiogénesis/farmacología , Animales , Anticuerpos/farmacología , Carcinoma Pulmonar de Lewis/irrigación sanguínea , Carcinoma Pulmonar de Lewis/tratamiento farmacológico , Carcinoma Pulmonar de Lewis/genética , Línea Celular Tumoral , Proliferación Celular , Femenino , Cadena Pesada de la Proteína-1 Reguladora de Fusión/genética , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Genotipo , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/inmunología , Células Endoteliales de la Vena Umbilical Humana/trasplante , Humanos , Integrinas/metabolismo , Masculino , Melanoma Experimental/irrigación sanguínea , Melanoma Experimental/tratamiento farmacológico , Melanoma Experimental/genética , Ratones Noqueados , Neovascularización Fisiológica/efectos de los fármacos , Fenotipo , Fosforilación , Neovascularización Retiniana/tratamiento farmacológico , Neovascularización Retiniana/genética , Neovascularización Retiniana/patología , Transducción de Señal , Factores de Tiempo , Carga Tumoral , Factor A de Crecimiento Endotelial Vascular/farmacología , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Graft-versus-host disease (GvHD) is a major barrier to the broader use of allogenic hematopoietic stem cell transplantation for non-malignant clinical applications. A murine model of C57BL/6 to B6D2F1 acute GvHD was employed with T lymphocytes harboring a deletion of the CD98 heavy chain (CD98hc(-/-)) as donor cells. The CD98hc(-/-) resulted in lower responses to alloantigen stimulation in a mixed leukocyte reaction assay, and prevented the mortality associated with disease progression. The percentage of donor CD8 T lymphocytes was significantly decreased, while the percentage of Foxp3-positive regulatory T cells (Tregs) in recipients was increased by CD98hc(-/-). Decreased expression of FAS, FASL, ICOS, ICOSL, PD-1 and PD-L1 by donor CD8 T cells, and mRNA expression of cytotoxic T cell-related cytokines in the recipients were shown in those with CD98hc(-/-). Fewer infiltrated cells are found in the lungs, liver, tongue and skin of recipients with CD98hc(-/-) compared with the wild type recipients. Taken together, our data indicate that T cell-specific deletion of CD98hc can contribute to the prevention of GvHD development due to the attenuation of lymphocyte migration and by increasing the generation of Treg cells. These findings are expected to make it possible to develop novel approaches for the prevention of GvHD.
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Movimiento Celular/inmunología , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Enfermedad Injerto contra Huésped/inmunología , Trasplante de Células Madre Hematopoyéticas , Transducción de Señal/inmunología , Linfocitos T Reguladores/inmunología , Aloinjertos , Animales , Movimiento Celular/genética , Femenino , Cadena Pesada de la Proteína-1 Reguladora de Fusión/genética , Enfermedad Injerto contra Huésped/genética , Enfermedad Injerto contra Huésped/patología , Ratones , Transducción de Señal/genética , Linfocitos T Reguladores/patologíaRESUMEN
The blood-brain barrier (BBB) poses a major challenge for developing effective antibody therapies for neurological diseases. Using transcriptomic and proteomic profiling, we searched for proteins in mouse brain endothelial cells (BECs) that could potentially be exploited to transport antibodies across the BBB. Due to their limited protein abundance, neither antibodies against literature-identified targets nor BBB-enriched proteins identified by microarray facilitated significant antibody brain uptake. Using proteomic analysis of isolated mouse BECs, we identified multiple highly expressed proteins, including basigin, Glut1, and CD98hc. Antibodies to each of these targets were significantly enriched in the brain after administration in vivo. In particular, antibodies against CD98hc showed robust accumulation in brain after systemic dosing, and a significant pharmacodynamic response as measured by brain Aß reduction. The discovery of CD98hc as a robust receptor-mediated transcytosis pathway for antibody delivery to the brain expands the current approaches available for enhancing brain uptake of therapeutic antibodies.
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Anticuerpos/uso terapéutico , Transporte Biológico/fisiología , Barrera Hematoencefálica/metabolismo , Encéfalo/metabolismo , Receptores de Transferrina/metabolismo , Animales , Anticuerpos/inmunología , Células Endoteliales/metabolismo , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Ratones , Proteómica/métodos , Transcitosis/fisiologíaRESUMEN
Upon their recognition of antigens presented by the MHC, T cell proliferation is vital for clonal expansion and the acquisition of effector functions, which are essential for mounting adaptive immune responses. The CD98 heavy chain (CD98hc, Slc3a2) plays a crucial role in the proliferation of both CD4+ and CD8+ T cells, although it is unclear if CD98hc directly regulates the T cell effector functions that are not linked with T cell proliferation in vivo. Here, we demonstrate that CD98hc is required for both CD4+ T cell proliferation and Th1 functional differentiation. T cell-specific deletion of CD98hc did not affect T cell development in the thymus. CD98hc-deficient CD4+ T cells proliferated in vivo more slowly as compared with control T cells. C57BL/6 mice lacking CD98hc in their CD4+ T cells could not control Leishmania major infections due to lowered IFN-γ production, even with massive CD4+ T cell proliferation. CD98hc-deficient CD4+ T cells exhibited lower IFN-γ production compared with wild-type T cells, even when comparing IFN-γ expression in cells that underwent the same number of cell divisions. Therefore, these data indicate that CD98hc is required for CD4+ T cell expansion and functional Th1 differentiation in vivo, and suggest that CD98hc might be a good target for treating Th1-mediated immune disorders.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Proliferación Celular/fisiología , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Interferón gamma/inmunología , Animales , Diferenciación Celular/inmunología , Activación de Linfocitos/inmunología , Ratones , Ratones Endogámicos C57BL , Células TH1/inmunologíaRESUMEN
CD4(+) T cell activation is controlled by signaling through the T cell receptor in addition to various co-receptors, and is also affected by their interactions with effector and regulatory T cells in the microenvironment. Inflammatory bowel diseases (IBD) are caused by the persistent activation and expansion of auto-aggressive CD4(+) T cells that attack intestinal epithelial cells. However, the molecular basis for the persistent activation of CD4(+) T cells in IBD remains unclear. In this study, we investigated how the CD98 heavy chain (CD98hc, Slc3a2) affected the development of colitis in an experimental animal model. Transferring CD98hc-deficient CD4(+)CD25(-) T cells into Rag2(-/-) mice did not cause colitis accompanied by increasing Foxp3(+) inducible regulatory T cells. By comparison, CD98hc-deficient naturally occurring regulatory T cells (nTregs) had a decreased capability to suppress colitis induced by CD4(+)CD25(-) T cells, although CD98hc-deficient mice did not have a defect in the development of nTregs. Blocking CD98hc with an anti-CD98 blocking antibody prevented the development of colitis. Our results indicate that CD98hc regulates the expansion of autoimmune CD4(+) T cells in addition to controlling nTregs functions, which suggests the CD98hc as an important target molecule for establishing strategies for treating colitis.
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Linfocitos T CD4-Positivos/inmunología , Colitis/inmunología , Colitis/patología , Colon/patología , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Animales , Enfermedades Autoinmunes/inmunología , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD4-Positivos/patología , Colitis/genética , Colon/inmunología , Colon/metabolismo , Proteínas de Unión al ADN/genética , Cadena Pesada de la Proteína-1 Reguladora de Fusión/genética , Eliminación de Gen , Subunidad alfa del Receptor de Interleucina-2/genética , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BLRESUMEN
CD98hc is a type II transmembrane protein that covalently links to one of several L-type amino acid transporters. CD98hc was first identified as a lymphocyte activation marker. In this study, we examined the role that CD98hc plays in the functions of macrophages using tissue specific knock-out miceCD98hc (CD98hc(flox/-)LysM-cre mice). When isolated peritoneal macrophages were incubated for 48 h, the macrophages obtained from the knock-out mice showed round-shaped morphologies, while almost all of the cells obtained from the control mice were spindle-shaped. The macrophage functions such as the antigen-presenting, phagocytic, and fusion activities, have been reported to decrease in CD98hc-deficient peritoneal macrophages. In addition, when the CD98hc deficient macrophages were stimulated with either IFN-γ/LPS or IL-4, the production of NO(2) or arginase-I decreased in comparison to that observed in the control macrophages. These findings show that the CD98hc molecules play an important role in the activation and functions of macrophages.
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Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Macrófagos Peritoneales/inmunología , Animales , Células Cultivadas , Cadena Pesada de la Proteína-1 Reguladora de Fusión/genética , Ratones , Ratones Noqueados , FagocitosisRESUMEN
BACKGROUND: Little is known about the CD98 heavy chain (CD98hc) in the T lymphocyte-mediated immune response to alloantigen. METHODS: We used an in vitro mixed leukocyte reaction assay and a cardiac transplantation model to evaluate the mechanisms of CD98hc in regulating alloimmune responses. RESULTS: A T cell-specific deficiency of CD98hc resulted in lower responses to alloantigen stimulation in a mixed leukocyte reaction assay, and CD98hc-deficient mice accepted full major histocompatibility complex-mismatched cardiac allografts. Consistent with graft survival, the infiltration of the graft by immune cells in CD98hc-deficient mice was significantly lower than that in wild-type mice. A chemotaxis assay revealed the migration of CD98hc-deficient lymphocytes to decrease in the presence of CCL5 compared with wild-type cells. Moreover, the proportion of CD4/Foxp3-positive cells and Foxp3 messenger RNA increased significantly in CD98hc-deficient recipients, consistent with the down-regulation of mammalian target of rapamycin and PS6 kinase; and allograft permanent acceptance was shortened by the depletion of antibody-induced regulatory T cells. Finally, neutralizing antibody against CD98hc prolonged the cardiac allograft survival. CONCLUSIONS: Taken together, our data indicate that T cell-specific deficiency in CD98hc can contribute to cardiac allograft permanent acceptance correlating with the attenuation of lymphocyte migration and by increasing the generation of regulatory T cells. These findings are expected to make it possible to develop novel approaches for treating allograft rejection and promoting transplantation tolerance.
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Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Rechazo de Injerto/prevención & control , Supervivencia de Injerto/inmunología , Trasplante de Corazón/inmunología , Linfocitos T/inmunología , Animales , Anticuerpos Monoclonales/uso terapéutico , Secuencia de Bases , Cadena Pesada de la Proteína-1 Reguladora de Fusión/genética , Cadena Pesada de la Proteína-1 Reguladora de Fusión/metabolismo , Rechazo de Injerto/inmunología , Trasplante de Corazón/mortalidad , Isoantígenos/inmunología , Estimación de Kaplan-Meier , Leucocitos/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Eliminación de Secuencia , Linfocitos T/metabolismo , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Trasplante Homólogo/inmunologíaRESUMEN
The interplay of CD4(+) and CD8(+) T cells targeting autoantigens is responsible for the progression of a number of autoimmune diseases, including type 1 diabetes mellitus (T1D). Understanding the molecular mechanisms that regulate T cell activation is crucial for designing effective therapies for autoimmune diseases. We probed a panel of Abs with T cell-modulating activity and identified a mAb specific for the H chain of CD98 (CD98hc) that was able to suppress T cell proliferation. The anti-CD98hc mAb also inhibited Ag-specific proliferation and the acquisition of effector function by CD4(+) and CD8(+) T cells in vitro and in vivo. Injection of the anti-CD98hc mAb completely prevented the onset of cyclophosphamide-induced diabetes in NOD mice. Treatment of diabetic NOD mice with anti-CD98hc reversed the diabetic state to normal levels, coincident with decreased proliferation of CD4(+) T cells. Furthermore, treatment of diabetic NOD mice with CD98hc small interfering RNA resolved T1D. These data indicate that strategies targeting CD98hc might have clinical application for treating T1D and other T cell-mediated autoimmune diseases.
Asunto(s)
Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 1/prevención & control , Cadena Pesada de la Proteína-1 Reguladora de Fusión/fisiología , Animales , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales/uso terapéutico , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD4-Positivos/patología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/patología , Ciclofosfamida/administración & dosificación , Diabetes Mellitus Tipo 1/inducido químicamente , Femenino , Cadena Pesada de la Proteína-1 Reguladora de Fusión/genética , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Inhibidores de Crecimiento/genética , Inhibidores de Crecimiento/metabolismo , Inhibidores de Crecimiento/fisiología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones SCID , Ratones Transgénicos , Ratas , Ratas WistarRESUMEN
The proliferation of antigen-specific lymphocytes and resulting clonal expansion are essential for adaptive immunity. We report here that B cell-specific deletion of the heavy chain of CD98 (CD98hc) resulted in lower antibody responses due to total suppression of B cell proliferation and subsequent plasma cell formation. Deletion of CD98hc did not impair early B cell activation but did inhibit later activation of the mitogen-activated protein kinase Erk1/2 and downregulation of the cell cycle inhibitor p27. Reconstitution of CD98hc-deficient B cells with CD98hc mutants showed that the integrin-binding domain of CD98hc was required for B cell proliferation but that the amino acid-transport function of CD98hc was dispensable for this. Thus, CD98hc supports integrin-dependent rapid proliferation of B cells. We propose that the advantage of adaptive immunity favored the appearance of CD98hc in vertebrates.
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Formación de Anticuerpos , Linfocitos B/inmunología , Proliferación Celular , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Animales , Linfocitos B/citología , Transporte Biológico Activo , Diferenciación Celular/fisiología , Células Cultivadas , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Integrinas/metabolismo , Activación de Linfocitos , Ratones , Ratones Transgénicos , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Células Plasmáticas/citología , Células Plasmáticas/inmunología , Unión ProteicaRESUMEN
Because glutathione scavenges reactive oxygen species (ROS) and also donates electrons to antioxidative systems, it may compensate for the oxidative stress caused by SOD1 deficiency. The cystine/glutamate transporter, which consists of two proteins, xCT and 4F2hc, has been designated system x (c) (-) . This transporter system plays a role in the maintenance of glutathione levels in mammalian cells. In the present study, we created SOD1 (-/-); xCT (-/-) double-knockout mice by intercrossing xCT-knockout and SOD1-knockout animals. We determined if the double-knockout mice express the phenotypic characteristics unique to SOD1 (-/-) mice-increased oxidative stress and the production of autoantibodies against erythrocytes. We also compared the phenotype of the double-knockout mice with those of the single-knockout and wild-type mice. Although two major antioxidative systems were found to be defective in the SOD1 (-/-); xCT (-/-) mice, relative to the SOD1 (-/-) mice, no functional deficits were observed. Based on these results, it appears that defects in system x (c) (-) do not exacerbate the phenotypic consequences of SOD1 deficiency in postnatal mice under ordinary breeding conditions.
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Sistema de Transporte de Aminoácidos y+/metabolismo , Estrés Oxidativo/fisiología , Especies Reactivas de Oxígeno/metabolismo , Superóxido Dismutasa/metabolismo , Sistema de Transporte de Aminoácidos y+/genética , Sistema de Transporte de Aminoácidos y+/inmunología , Animales , Autoanticuerpos/inmunología , Autoanticuerpos/metabolismo , Transporte Biológico/fisiología , Cruzamientos Genéticos , Eritrocitos/inmunología , Cadena Pesada de la Proteína-1 Reguladora de Fusión/genética , Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Cadena Pesada de la Proteína-1 Reguladora de Fusión/metabolismo , Ratones , Ratones Noqueados , Fenotipo , Especies Reactivas de Oxígeno/inmunología , Superóxido Dismutasa/genética , Superóxido Dismutasa/inmunología , Superóxido Dismutasa-1RESUMEN
CD98 is a widely expressed cell surface heterodimeric protein of 125 kDa. Its expression is upregulated during lymphocyte activation induced by mitogen, superantigen, conventional antigen, and a combination of phorbol myristate acetate (PMA) and ionomycin. However, the role of CD98 in the immune system is not so well understood. The role of CD98 in murine T lymphocyte proliferation was investigated, especially in correlation with the interleukin 2 (IL-2)/interleukin 2 receptor (IL-2R) system. Monoclonal antibody (mAb) directed against murine CD98 heavy chain (mCD98HC) suppressed the proliferation of lymphocytes stimulated with concanavalin A (Con A). Anti-mCD98HC mAb did not suppress the expression of IL-2Ralpha. Anti-IL-2Ralpha mAb, which suppressed DNA synthesis, did not inhibit the expression of CD98HC. Murine IL-2 (recombinant), which induced considerable DNA synthesis by lymphocytes stimulated with a sub-optimal dose of Con A, did not induce CD98HC expression in lymphocytes. In addition, anti-mCD98HC mAb did not inhibit the production of IL-2 by lymphocyte stimulated with Con A. Taken together with these findings, it was speculated that the CD98 system is independent of the IL-2/IL-2R system in murine T lymphocyte activation.
Asunto(s)
Cadena Pesada de la Proteína-1 Reguladora de Fusión/inmunología , Interleucina-2/inmunología , Activación de Linfocitos , Receptores de Interleucina-2/inmunología , Linfocitos T/inmunología , Animales , Anticuerpos Monoclonales/metabolismo , Concanavalina A/farmacología , Masculino , Ratones , Ratones Endogámicos C57BL , Linfocitos T/efectos de los fármacosRESUMEN
Spleen cells from a (BALB/c x C57BL/6)F1 mouse immunized with CBA/J spleen cells were fused with the myeloma cell line NS-1. One of the six established hybrid cell lines continuously secreted antibody that recognized a new antigenic specificity, tentatively called "Ly-10.1". This newly found antigen is expressed on thymocytes, on splenic T and B cells, on bone-marrow cells, and on the cells derived from brain, kidney and liver. It is also expressed on a continuous cell line, 416B, with stem-cell characteristics. The unique tissue distribution and, furthermore, a distinct strain distribution pattern distinguishes Ly-10.1 from any known murine lymphocyte alloantigen. On the basis of reactivity with cells of the C57BL/6-Lyt-1a congenic strain, one gene governing Ly-10 expression is assigned to the Lyt-1 region of chromosome 19.