(1/8548) Crystal structure of an MHC class I presented glycopeptide that generates carbohydrate-specific CTL.
T cell receptor (TCR) recognition of nonpeptidic and modified peptide antigens has been recently uncovered but is still poorly understood. Immunization with an H-2Kb-restricted glycopeptide RGY8-6H-Gal2 generates a population of cytotoxic T cells that express both alpha/beta TCR, specific for glycopeptide, and gamma/delta TCR, specific for the disaccharide, even on glycolipids. The crystal structure of Kb/RGY8-6H-Gal2 now demonstrates that the peptide and H-2Kb structures are unaffected by the peptide glycosylation, but the central region of the putative TCR binding site is dominated by the extensive exposure of the tethered carbohydrate. These features of the Kb/RGY8-6H-Gal2 structure are consistent with the individual ligand binding preferences identified for the alpha/beta and gamma/delta TCRs and thus explain the generation of a carbohydrate-specific T cell response. (+info)
(2/8548) Crystal structures of two H-2Db/glycopeptide complexes suggest a molecular basis for CTL cross-reactivity.
Two synthetic O-GlcNAc-bearing peptides that elicit H-2Db-restricted glycopeptide-specific cytotoxic T cells (CTL) have been shown to display nonreciprocal patterns of cross-reactivity. Here, we present the crystal structures of the H-2Db glycopeptide complexes to 2.85 A resolution or better. In both cases, the glycan is solvent exposed and available for direct recognition by the T cell receptor (TCR). We have modeled the complex formed between the MHC-glycopeptide complexes and their respective TCRs, showing that a single saccharide residue can be accommodated in the standard TCR-MHC geometry. The models also reveal a possible molecular basis for the observed cross-reactivity patterns of the CTL clones, which appear to be influenced by the length of the CDR3 loop and the nature of the immunizing ligand. (+info)
(3/8548) Control of metastasis by Asn-linked, beta1-6 branched oligosaccharides in mouse mammary cancer cells.
Studies in cell lines and malignant human tissues have shown that increased cell-surface Asn-linked beta1-6(GlcNAcbeta1-6Man) branching is associated with increased tumorigenic and metastatic properties. In this study, three mouse mammary cancer cell lines were transfected with an expression vector containing the mouse cDNA for N-acetylglucosaminyltransferase V (GlcNAcT-V EC 22.214.171.124), the glycosyltransferase responsible for initiating beta1-6 branching on Asn-linked carbohydrates. The cell lines were screened for increased cytotoxicity to L-PHA, a lectin specific for beta1-6 branching structures. Cell lines exhibiting increased L-PHA cytotoxicity expressed increased levels of beta1-6 branching structures. Northern blots detected the presence of GlcNAcT-V transcribed from the expression vector in the L-PHA sensitive cell lines. After injection into the tail veins of mice, transfected cell lines with increased beta1-6 branching on the cell surface formed elevated levels of lung tumors relative to control transfected cell lines (P < 0.002). Western blots of membrane proteins from GlcNAcT-V transfected and control cells probed with the lectins DSA and WGA did not show an increase in polyN-acetyllactosamine and sialic acid content in the transfected cell lines. These results demonstrate that a specific increase in beta1-6 branching due to an elevation in GlcNAcT-V expression increases metastatic potential. (+info)
(4/8548) Giardia induces proliferation and interferon gamma production by intestinal lymphocytes.
BACKGROUND: Murine intraepithelial lymphocytes kill Giardia lambia; responses of human intestinal lymphocytes to this parasite are unknown. AIMS: To examine giardia induced proliferation, interferon gamma production, migration, and cytotoxicity by lymphocytes from the human intestine and peripheral blood. METHODS: Giardia were added to intraepithelial lymphocytes, lamina propria lymphocytes, and peripheral blood lymphocytes, obtained from jejunal mucosa and blood of otherwise healthy patients undergoing gastric bypass surgery for morbid obesity. Proliferation was measured by 3H-thymidine incorporation; frequency of proliferation precursors, by limiting dilution analysis; interferon gamma production, by ELISA; cytotoxicity, by 51Cr release of radiolabelled giardia and by release of serine esterases by effector lymphocytes that mediate cytotoxicity. RESULTS: The CD4+ T lymphocytes from intestine and blood proliferated in response to giardia. The stimulus by the parasite was mitogenic rather than antigenic due to the fact that the peak response was on day 3 rather than day 6, and the large number of precursors was in the range of that for mitogens. CD4+ T lymphocytes from both sites produced interferon gamma in response to giardia. Lymphocytes did not migrate towards or kill the parasite. CONCLUSIONS: Giardia induced the same degree of proliferation and interferon gamma production by CD4+ T lymphocytes in intestine and blood, but did not trigger cytotoxicity or migration. (+info)
(5/8548) Enhanced tumor growth and invasiveness in vivo by a carboxyl-terminal fragment of alpha1-proteinase inhibitor generated by matrix metalloproteinases: a possible modulatory role in natural killer cytotoxicity.
Matrix metalloproteinases (MMPs) are believed to contribute to the complex process of cancer progression. They also exhibit an alpha1-proteinase inhibitor (alphaPI)-degrading activity generating a carboxyl-terminal fragment of approximately 5 kd (alphaPI-C). This study reports that overexpression of alphaPI-C in S2-020, a cloned subline derived from the human pancreas adenocarcinoma cell line SUIT-2, potentiates the growth capability of the cells in nude mice. After stable transfection of a vector containing a chimeric cDNA encoding a signal peptide sequence of tissue inhibitor of metalloproteinase-1 followed by cDNA for alphaPI-C into S2-020 cells, three clones that stably secrete alphaPI-C were obtained. The ectopic expression of alphaPI-C did not alter in vitro cellular growth. However, subcutaneous injection of the alphaPI-C-secreting clones resulted in tumors that were 1.5 to 3-fold larger than those of control clones with an increased tendency to invasiveness and lymph node metastasis. These effects could be a result of modulation of natural killer (NK) cell-mediated control of tumor growth in nude mice, as the growth advantage of alphaPI-C-secreting clones was not observed in NK-depleted mice, and alphaPI-C-secreting clones showed decreased NK sensitivity in vitro. In addition, production of alphaPI and generation of the cleaved form of alphaPI by MMP were observed in various human tumor cell lines and in a highly metastatic subline of SUIT-2 in vitro. These results provide experimental evidence that the alphaPI-degrading activity of MMPs may play a role in tumor progression not only via the inactivation of alphaPI but also via the generation of alphaPI-C. (+info)
(6/8548) Suppression of angiogenesis, tumorigenicity, and metastasis by human prostate cancer cells engineered to produce interferon-beta.
We determined whether the IFN-beta gene can be used to suppress angiogenesis, tumor growth, and metastasis of human prostate cancer cells growing in the prostate of nude mice. Highly metastatic PC-3M human prostate cancer cells were engineered to constitutively produce murine IFN-beta subsequent to infection with a retroviral vector containing murine IFN-beta cDNA. Parental (PC-3M-P), control vector-transduced (PC-3M-Neo), and IFN-beta-transduced (PC-3M-IFN-beta) cells were injected into the prostate (orthotopic) or subcutis (ectopic) of nude mice. PC-3M-P and PC-3M-Neo cells produced rapidly growing tumors and regional lymph node metastases, whereas PC-3M-IFN-beta cells did not. PC-3M-IFN-beta cells also suppressed the tumorigenicity of bystander nontransduced prostate cancer cells. PC-3M-IFN-beta cells produced small tumors (3-5 mm in diameter) in nude mice treated with anti-asialo GM1 antibodies and in severe combined immunodeficient/Beige mice. Immunohistochemical staining revealed that PC-3M-IFN-beta tumors were homogeneously infiltrated by macrophages, whereas control tumors contained fewer macrophages at their periphery. Most tumor cells in the control tumors were stained positive by an antibody to proliferative cell nuclear antigen; very few were positively stained by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling. In sharp contrast, PC-3M-IFN-beta tumors contained fewer proliferative cell nuclear antigen-positive cells and many terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling-positive cells. Staining with antibody against CD31 showed that control tumors contained more blood vessels than PC-3M-IFN-beta tumors. PC-3M-IFN-beta cells were more sensitive to lysis mediated by natural killer cells in vitro or to cytostasis mediated by macrophages than control transduced cells. Conditioned medium from PC-3M-IFN-beta cells augmented splenic cell-mediated cytolysis to control tumor cells, which could be neutralized by antibody against IFN-beta. Collectively, the data suggest that the suppression of tumorigenicity and metastasis of PC-3M-IFN-beta cells is due to inhibition of angiogenesis and activation of host effector cells. (+info)
(7/8548) Rapid death of adoptively transferred T cells in acquired immunodeficiency syndrome.
Human immunodeficiency virus (HIV)-specific cytotoxic T lymphocytes (CTL) probably play the major role in controlling HIV replication. However, the value of adoptive transfer of HIV-specific CTL expanded in vitro to HIV+ patients has been limited: this contrasts with the success of CTL therapy in treating or preventing Epstein-Barr virus and cytomegalovirus disease after bone marrow transplantation (BMT). We investigated the fate of expanded HIV-specific CTL clones in vivo following adoptive transfer to a patient with acquired immunodeficiency syndrome (AIDS). Two autologous CTL clones specific for HIV Gag and Pol were expanded to large numbers (>10(9)) in vitro and infused into an HIV-infected patient whose viral load was rising despite antiretroviral therapy. The fate of one clone was monitored by staining peripheral blood mononuclear cells (PBMCs) with T-cell receptor-specific tetrameric major histocompatibility complex (MHC)-peptide complexes. Although the CTL transfer was well tolerated, there were no significant changes in CD4 and CD8 lymphocyte counts and virus load. By tracking an infused clone using soluble MHC-peptide complexes, we show that cells bearing the Gag-specific T-cell receptors were rapidly eliminated within hours of infusion through apoptosis. Thus, the failure of adoptively transferred HIV-specific CTL to reduce virus load in AIDS may be due to rapid apoptosis of the infused cells, triggered by a number of potential mechanisms. Further trials of adoptive transfer of CTL should take into account the susceptibility of infused cells to in vivo apoptosis. (+info)
(8/8548) Contribution of natural killer cells to inhibition of angiogenesis by interleukin-12.
Interleukin-12 (IL-12) inhibits angiogenesis in vivo by inducing interferon-gamma (IFN-gamma) and other downstream mediators. Here, we report that neutralization of natural killer (NK) cell function with antibodies to either asialo GM1 or NK 1.1 reversed IL-12 inhibition of basic fibroblast growth factor (bFGF)-induced angiogenesis in athymic mice. By immunohistochemistry, those sites where bFGF-induced neovascularization was inhibited by IL-12 displayed accumulation of NK cells and the presence of IP-10-positive cells. Based on expression of the cytolytic mediators perforin and granzyme B, the NK cells were locally activated. Experimental Burkitt lymphomas treated locally with IL-12 displayed tumor tissue necrosis, vascular damage, and NK-cell infiltration surrounding small vessels. After activation in vitro with IL-12, NK cells from nude mice became strongly cytotoxic for primary cultures of syngeneic aortic endothelial cells. Cytotoxicity was neutralized by antibodies to IFN-gamma. These results document that NK cells are required mediators of angiogenesis inhibition by IL-12, and provide evidence that NK-cell cytotoxicity of endothelial cells is a potential mechanism by which IL-12 can suppress neovascularization. (+info)