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Biomedical subjects

G P Linette

Publications and source records attributed to G P Linette.

At least 19 recordsLinked to original sources
PubMed2000

HLA-A2.1/K(b) transgenic murine dendritic cells transduced with an adenovirus encoding human gp100 process the same A2.1-restricted peptide epitopes as human antigen-presenting cells and elicit A2.1-restricted peptide-specific CTL.

HLA-A2.1/K(b) transgenic mice (A2.1/K(b) mice) were used to investigate the processing of human gp100 melanoma antigen by murine antigen presenting cells (APC). Bone marrow-derived dendritic cells (DC) from A2.1/K(b) mice were transduced with adenovirus encoding human gp100 (Ad2/hugp100v2). The Ad2/hugp100v2-transduced DC express human gp100, as documented by immunoperoxidase staining. Flow cytometric analysis demonstrates that Ad vector transduction does not downregulate expression of several markers, including MHC class I. We show that Ad2/hugp100v2-transduced DC are recognized by peptide-specific, A2.1-restricted CTL, suggesting correct processing and presentation of the hugp100 antigen by murine DC. To assess dominance among the various A2.1-restricted epitopes encoded by hugp100, A2.1/K(b) transgenic mice were immunized with Ad2/hugp100v2-transduced DC. Resulting effector cytotoxic T lymphocytes (CTL) were assayed for peptide specificity using a panel of six synthetic peptides known to encode A2.1-restricted epitopes of human gp100 (denoted G154, G177, G209, G280, G457, G476). CTL obtained from Ad2/hugp100v2-transduced DC immunized A2.1/K(b) mouse lysed target cells presenting five of the six epitopes, supporting the observation that murine cells correctly process the hugp100 antigen. The immunogenicity of individual gp100 epitopes correlates with their binding affinity to A2.1. CTL generated from A2.1/K(b) mice immunized with Ad2/hugp100v2-transduced DC also specifically recognize A2.1(+)/gp100(+) human melanoma cells. These data suggest that murine APC process and present the same set of HLA-restricted peptides, similar to human APC. HLA transgenic mice serve as a useful model system to study class I-restricted epitopes of human tumor-associated antigens.

Adenoviridae↗
PubMed2000

In vitro priming with adenovirus/gp100 antigen-transduced dendritic cells reveals the epitope specificity of HLA-A*0201-restricted CD8+ T cells in patients with melanoma.

Replication-deficient recombinant adenovirus (Ad) encoding human gp100 or MART-1 melanoma Ag was used to transduce human dendritic cells (DC) ex vivo as a model system for cancer vaccine therapy. A second generation E1/E4 region deleted Ad which harbors the CMV immediate-early promoter/enhancer and a unique E4-ORF6/pIX chimeric gene was employed as the backbone vector. We demonstrate that human monocyte-derived DC are permissive to Ad infection at multiplicity of infection between 100 and 500 and occurs independent of the coxsackie Ad receptor. Fluorescent-labeled Ad was used to assess the kinetics and distribution of viral vector within DC. Ad-transduced DC show peak transgene expression at 24-48 h and expression remains detectable for at least 7 days. DC transduced with replication-deficient Ad do not exhibit any unusual phenotypic characteristics or cytopathic effects. DC transduced with Ad2/gp100v2 can elicit tumor-specific CTL in vitro from patients bearing gp100+ metastatic melanoma. Using a panel of gp100-derived synthetic peptides, we show that Ad2/gp100v2-transduced DC elicit Ag-specific CTL that recognize only the G209 and G280 epitopes, both of which display relatively short half-lives ( approximately 7-8 h) on the surface of HLA-A*0201+ cells. Thus, patients with metastatic melanoma are not tolerant to gp100 Ag based on the detection of CD8+ T cells specific for multiple HLA-A*0201-restricted, gp100-derived epitopes.

Adenoviruses, Human↗
PubMed1996

Cross talk between cell death and cell cycle progression: BCL-2 regulates NFAT-mediated activation.

BCL-2-deficient T cells demonstrate accelerated cell cycle progression and increased apoptosis following activation. Increasing the levels of BCL-2 retarded the G0-->S transition, sustained the levels of cyclin-dependent kinase inhibitor p27Kip1, and repressed postactivation death. Proximal signal transduction events and immediate early gene transcription were unaffected. However, the transcription and synthesis of interleukin 2 and other delayed early cytokines were markedly attenuated by BCL-2. In contrast, a cysteine protease inhibitor that also blocks apoptosis had no substantial affect upon cytokine production. InterleUkin 2 expression requires several transcription factors of which nuclear translocation of NFAT (nuclear factor of activated T cells) and NFAT-mediated transactivation were impaired by BCL-2. Thus, select genetic aberrations in the apoptotic pathway reveal a cell autonomous coregulation of activation.

Apoptosis↗
PubMed1995

Bcl-XL and Bcl-2 repress a common pathway of cell death.

The effect of Bcl-xL upon the developmental death of T cells was assessed by generating transgenic mice that expressed Bcl-xL within all thymocyte subsets. Bcl-xL protected thymocytes from a variety of apoptotic stimuli, including gamma irradiation, glucocorticoids, and anti-CD3 treatment. Bcl-xL altered thymocyte maturation, resulting in increased numbers of CD3int/hi and CD4-8+ thymocytes. Overall, the phenotype of Bcl-xL transgenics was essentially indistinguishable from a Bcl-2 transgenic model. Overexpression of Bcl-xL or Bcl-2 resulted in the down-regulation of the other molecule, providing further evidence of their reciprocal regulation. In a genetic test of redundancy, the Bcl-xL transgene rescued mature T cells in Bcl-2 null mice. Immunoprecipitation indicated that Bcl-xL, like Bcl-2, heterodimerized with the death-promoting molecule Bax in thymocytes. This in vivo model argues that Bcl-xL, like Bcl-2, functions in a common pathway to repress cell death.

Animals↗
PubMed1995

Peripheral T-cell lymphoma in lckpr-bcl-2 transgenic mice.

t(14;18) is the most common translocation in human lymphoid malignancy and results in bcl-2 overexpression. Bcl-2 blocks apoptosis and constitutes the initial member of a new category of oncogenes, ie, regulators of cell death. Bcl-2-Ig transgenic mice develop follicular hyperplasia and progress to malignant B-cell lymphoma. To assess the oncogenic potential of bcl-2 in the T-cell lineage, a cohort of 68 lckpr-bcl-2 transgenic mice and 56 control littermates were monitored for signs of malignancy over a 24-month period. Eighteen (26%) lckpr-bcl-2 mice developed diffuse, predominantly large-cell lymphomas at a mean age of 18 months. In contrast, only one nontransgenic control mouse developed lymphoma. CD3 surface expression and clonal T-cell receptor beta rearrangements support the T-lineage classification of these neoplasms. lckpr-bcl-2-enforced lymphomas are predominantly CD4+CD8-, consistent with a mature peripheral T-cell phenotype. These data provide support for the thesis that violation of homeostasis through the repression of cell death can be a primary mechanism of tumorigenesis in multiple lineages.

Animals↗
PubMed1995

Unimpaired thymic and peripheral T cell death in mice lacking the nuclear receptor NGFI-B (Nur77).

T cell hybridomas require the immediate-early gene NGFI-B (nur77) for T cell receptor (TCR)-mediated apoptosis, a model for negative selection of self-reactive T cells. TCR-mediated death was examined in mice bearing an NGFI-B loss-of-function mutation, either by administration of antibodies to CD3 (anti-CD3) or in two well-characterized transgenic models expressing self-reactive TCRs. Both the extent and the rate of thymocyte death were unimpaired. Anti-CD3-induced death was normal in CD4+ peripheral T cells, in which death is mediated predominantly by the Fas signaling pathway. Thus, no unique requirement for NGFI-B is observed for thymic or peripheral T cell death.

Animals↗
PubMed1995

Reactive oxygen species and the regulation of cell death by the Bcl-2 gene family.

The maintenance of homeostasis in normal tissues reflects a balance between cell proliferation and cell death. Bcl-2 inaugurated a new category of oncogenes, regulators of cell death. The Bcl-2 gene was identified at the chromosomal breakpoint of t(14;18) bearing B cell lymphomas. Bcl-2 proved unique by blocking programmed cell death rather than promoting proliferation. In adults, Bcl-2 is topographically restricted to progenitor cells and longlived cells but is much more widespread in the developing embryo. Transgenic mice that overexpress Bcl-2 demonstrate extended cell survival, and progress to high grade lymphomas. Bcl-2 has been localized to mitochondria, endoplasmic reticulum and nuclear membranes, also the sites of reactive oxygen species generation. Bcl-2 does not appear to influence the generation of oxygen free radicals but does prevent oxidative damage to cellular constituents including lipid membranes. Bcl-2 deficient mice complete embryonic development but undergo fulminant lymphoid apoptosis of thymus and spleen. Moreover, they demonstrate two unexpected pathologies resulting from cell death, polycystic kidney disease and hair hypopigmentation. The latter is a potential oxidant injury from the melanin biosynthetic pathway. A family of Bcl-2 related genes is emerging that includes Bax, a conserved homolog that heterodimerizes in vivo with Bcl-2 and promotes cell death. The ratio of family members, such as Bcl-2/Bax, determines the survival or death of cells following an apoptotic stimulus.

Amino Acid Sequence↗
PubMed1994

Differentiation and cell death: lessons from the immune system.

The immune system provides a unique vantage point from which to view the interrelationship between differentiation and cell death, as apoptosis is a prominent feature of B- and T-lymphocyte development. Two common themes emerge from recent experimental observations. First, survival signals are crucial during the differentiation process. The upregulation of Bcl-2 during positive selection suggests that this molecule serves as a survival signal to maintain lymphoid homeostasis. Second, if cell death is repressed, cellular differentiation can occur in the absence of inductive signals.

Apoptosis↗
PubMed1994

Bcl-2 is upregulated at the CD4+ CD8+ stage during positive selection and promotes thymocyte differentiation at several control points.

In vivo thymocyte maturation models were used to investigate the differentiation role of Bcl-2. In alpha/beta T cell receptor (TCR) class II-restricted transgenic mice, Bcl-2 was upregulated at the CD4+ CD8+ stage during positive selection. The lckpr-bcl2 transgene was bred onto MHC classes I-I- and II-I-, MHC-I-, and alpha/beta TCR backgrounds to determine whether Bcl-2 promoted thymocyte maturation in the absence of coreceptor-MHC interaction. Bcl-2 rescued CD8+ thymocytes in class I-I- and alpha/beta TCR in mice; however, they were not exported to the periphery. Bcl-2 had no effect on CD4 lineage maturation in class II-I- mice. No single-positive thymocytes accumulate in MHC-I- mice despite overexpressed Bcl-2. Thus, Bcl-2 enables selection of certain TCRs on class II molecules and their differentiation along the CD8 pathway; however, Bcl-2 did not substitute for positive selection. In RAG-1-I- mice, Bcl-2 promoted differentiation to the CD4+ CD8+ stage. Bcl-2 can promote thymocyte maturation at several control points.

Animals↗
PubMed1991

Resistance in murine schistosomiasis is contingent on activated IL-2 receptor-bearing L3T4+ lymphocytes, negatively regulated by Lyt-2+ cells, and uninfluenced by the presence of IL-4.

These studies assess the roles of subpopulations of T lymphocytes in inducing and modulating resistance to Schistosoma mansoni. C57BL/6 mice were depleted in vivo of L3T4+, Lyt-1+, Lyt-2+, IL-2R+ cells, or IL-4 by administration of appropriate mAb. Resistance and various correlative parameters of the immune response were studied in normal, depleted, and congenitally athymic mice. Depletion of T lymphocytes by anti-L3T4 or anti-IL-2R mAb reduced the development and expression of resistance, IgG2a and IgE antibody formation, and delayed type hypersensitivity reactivity against schistosome Ag. Depletion with anti-IL-4 antibody led to profound suppression of IgE-eosinophil-mediated antibody-dependent cell-mediated cytotoxicity and passive cutaneous anaphylaxis responses against the parasite and no effect on IgG2a antibody, Ag-mediated blast transformation, or resistance. Depletion of Lyt-2+ cells produced augmented development and expression of resistance and an increase in the immunological parameters of anti-schistosome reactivity. These studies suggest that protective immunity to S. mansoni in mice, induced by irradiated cercariae, is dependent on L3T4+, IL-2R+ lymphocytes and negatively regulated by Lyt-2+ cells. IL-4 does not appear to be essential for the development of resistance but is essential for the IgE response to the parasite.

Animals↗
PubMed1990

The regulation of resistance to Schistosoma mansoni by auto-anti-idiotypic immunity. III. An analysis of effects on epitopic recognition, idiotypic expression, and anti-idiotypic reactivity at the clonal level.

Auto-anti-idiotypic mechanisms can regulate the protective immune response against Schistosoma mansoni. Anti-idiotypic responses were stimulated by immunization of mice either with nonspecifically induced lymphoblasts, produced with Con A, or with Ag-induced lymphoblasts bearing specific idiotypic receptors. The effect of the induced anti-idiotypic response upon clonotypic cellular reactivity was assessed in vitro through the suppression of antigen-mediated blast transformation by cloned T cells and in vivo by suppression of resistance to S. mansoni and delayed-type hypersensitivity responses against specific Ag. Differential regulation of humoral immune responses was studied at the levels of specific epitopic recognition, the expression of specific Id, and the production of anti-idiotypic responses directed against mAb bearing specific Id. Anti-idiotypic sensitization resulted in variable (10 to 90%) suppression of the immune response to discrete antigenic epitopes, the expression of specific idiotypic phenotypes, and anti-idiotypic, antiparatopic responses against T cell clonotypes and antibody idiotypic phenotypes. In vitro admixture and in vivo challenge studies resulted in consonant differential suppression. Thus idiotypic regulation can mold the fine specificities of the protective immune response to S. mansoni at the clonal level and may provide an approach to optimize the expression and assessment of resistance.

Animals↗
PubMed1990

The regulation of resistance to Schistosoma mansoni by auto-anti-idiotypic immunity. II. Global qualitative and quantitative regulation.

These studies explore the suppression of resistance to schistosomiasis mansoni through interactions of autologous immune functions derived from an induced anti-idiotypic response. This anti-clonotypic response is induced by immunization with syngeneic L3T4+ receptor-bearing lymphoblasts and for the sake of description is termed "auto-anti-idiotypic". It is antigenically restricted and cannot be induced by allogeneic cells. Anti-idiotypic immunization profoundly suppressed the development of protective immunity after exposure to irradiated cercariae and altered a wide variety of functional humoral and cellular immune responses to the parasite. In addition to quantitative suppressive effects, the anti-idiotypic network also regulated qualitative aspects of the immune response by increasing the heterogeneity and reducing the functional binding avidity of antibody for Ag. These effects also were reflected in analogous alterations in cellular reactivity, using the criteria of the Ag mediated blast transformation and delayed type hypersensitivity. Thus idiotypic regulation can mold the specificity and sensitivity of the immune response to Schistosoma mansoni by affecting quantitative and qualitative responses. Manipulation of idiotypic recognition provides an approach to optimize the expression of protective resistance to schistosomiasis.

Animals↗
PubMed1988

The regulation of resistance to Schistosoma mansoni by auto-anti-idiotypic immunity.

These studies explore auto-anti-idiotypic mechanisms as potential regulators of the protective immune response against Schistosoma mansoni. Anti-idiotypic responses were stimulated by immunization of mice with lymphoblasts, bearing specific idiotypic receptors. These receptors were produced in vitro by stimulation of Ag-reactive T cells by soluble cercarial immunogen, keyhole limpet hemocyanin, or Con A. The animals were then exposed to irradiated cercariae, keyhole limpet hemocyanin, or SRBC. The results indicate that the soluble cercarial immunogen lymphoblast recipient mice demonstrated reduction in a number of parameters of their immune response to schistosome Ag, including resistance to challenge by parasites. These changes were immunologically specific. Anti-idiotypic antibodies and anti-clonotypic T cell reactivity was demonstrated in the lymphoblast immunized mice. The suppression of reactivity in LBM was mediated by Lyt-1-, L3T-4-, and Lyt-2+ lymphocytes. These studies suggest that idiotypically dependent pathways might be important for the regulation of resistance to schistosomiasis.

Animals↗
PubMed1988

HIV-1-infected T cells show a selective signaling defect after perturbation of CD3/antigen receptor.

The binding of antigen or monoclonal antibody to the T cell receptor for antigen or the closely associated CD3 complex causes increases in the concentration of intracellular ionized calcium and subsequent cell proliferation. By measuring second messenger production in primary cultures of human immunodeficiency virus (HIV-1)--infected T cells stimulated with monoclonal antibodies specific for either CD3 or CD2, a specific impairment of membrane signaling was revealed. The HIV-1--infected T cells were unable to mobilize Ca2+ after stimulation with anti-CD3, whereas CD2-induced calcium mobilization remained intact. Furthermore, the HIV-1--infected cells proliferated poorly after CD3 stimulation, although the cells retained normal DNA synthesis in response to interleukin-2 stimulation. These results show that the signals initiated by CD2 and CD3 can be regulated independently within the same T cell; uncoupling of signal transduction after antigen-specific stimulation provides a biochemical mechanism to explain, in part, the profound immunodeficiency of patients with HIV-1 infection.

Acquired Immunodeficiency Syndrome↗
PubMed1987

The immune response to Schistosoma mansoni infections in inbred rats. VI. Regulation by T cell subpopulations.

These studies assess the roles of subpopulations of T lymphocytes in inducing and modulating resistance to Schistosoma mansoni. CDF rats were depleted of RT 7.1+ (anti-Pan-T), W3/25+ (anti-T helper/inducer), or OX8+ (anti-T suppressor) cells by the in vivo administration of monoclonal antibodies (mAb). The development of parasites and immunity to challenge by S. mansoni were compared with results in undepleted normal and congenitally athymic rats. Discrete subpopulations of T lymphocytes were adoptively transferred to ascertain effects upon parasite development and the protective immune response. In vitro studies, involving utilizing cocultivation of cell subpopulations +/- cyclosporin A, were utilized to dissect mechanisms. Depletion of T lymphocytes by anti-RT7.1 mAb and anti-W3/25 mAb resulted in augmented initial worm development, suboptimal resistance, and decreased antibody and delayed-type hypersensitive reactivity directed against schistosome antigens. Depletion with OX8 mAb produced opposite effects. The adoptive transfer of T cell subpopulations produced concordant results with T cell regulation expressed B cell-dependent effector mechanisms. The coadoptive transfer of cells resulted in the suppression of resistance afforded by the W3/25+ cells by OX8+ cells, which could be augmented in vitro by cyclosporin A. Thus, protective immunity to S. mansoni in rats is regulated by discrete subpopulations of T lymphocytes. The findings suggest the possibility of selective immune regulation of resistance based on the manipulation of specific T cell subpopulation.

Animals↗
PubMed1987

In vivo T cell depletion regulates resistance and morbidity in murine schistosomiasis.

These studies assessed the roles of subpopulations of T lymphocytes in inducing and modulating resistance to schistosomiasis and thereby influencing subsequent morbidity. C57BL/6 mice were depleted in vivo of Lyt-1+, Lyt-2+, and L3T4+ cells by the daily administration of monoclonal antibodies. The development of protective immunity, induced by exposure to irradiated Schistosoma mansoni cercariae as expressed in depleted animals, was compared to that demonstrated in undepleted, normal, and congenitally athymic C57BL/6 mice. The development of morbidity was determined by spleen weight, portal pressure and reticuloendothelial system activity. The results indicated that depletion of specific subpopulations of T lymphocytes minimally affected the primary development of parasites; however, depletion strongly influenced the development of resistance to the parasite and subsequent morbidity due to infection. Depletion of T lymphocytes by anti-Lyt-1+ or anti-L3T4+ antibody decreased the development of resistance, antibody and delayed-type hypersensitivity directed against schistosome antigens. Morbidity due to disease was increased. Depletion of Lyt-2+ cells produced opposite changes with augmented resistance and reduced morbidity. Congenitally athymic mice developed minimal resistance and morbidity. Moreover, resistance was inversely related to the morbidity shown by a given animal. These studies indicate that the development of protective immunity to S. mansoni cercariae is regulated by discrete subpopulations of T lymphocytes. The feasibility of decreasing morbidity by increasing specific immunologically mediated resistance is suggested.

Animals↗