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Gennaro De Libero

Publications and source records attributed to Gennaro De Libero.

At least 19 recordsLinked to original sources
PubMed2006

Mechanisms of lipid-antigen generation and presentation to T cells.

The presentation of lipid antigens by CD1 molecules follows precise rules imposed by the biochemical nature of lipids. The structures of CD1-lipid complexes are elucidating how T-cell receptors interact with hydrophobic antigens. The mechanism of lipid uptake and the pathways followed by lipids embedded in the cell membrane contribute to the efficient presentation of exogenous and self-lipids. Lipid presentation is further regulated by the trafficking route of CD1 proteins and their precise membrane localization within endosomal vesicles. Moreover, the generation of immunogenic lipids might require adequate processing, which occurs in the presence of lipid-binding proteins, including CD1e. Here, we review recent experimental evidence that has revealed new protagonists involved in generating immunogenic lipids and has indicated unexpected biological mechanisms contributing to immune recognition.

Animals↗
PubMed2006

How T lymphocytes recognize lipid antigens.

Recognition of lipid antigens by T lymphocytes is well established. Lipids are recognized by T cells when presented in association with CD1 antigen-presenting molecules. Both microbial and self lipids stimulate specific T lymphocytes, thus participating in immune reactions during infections and autoimmune diseases. The immune system uses a variety of strategies to solubilise lipid antigens, to facilitate their internalization, processing, and loading on CD1 molecules. Recent studies in the field of lipid antigen presentation have revealed new mechanisms which allow the immune system to sense lipids as stimulatory antigens.

Animals↗
PubMed2006

Endogenous phosphatidylcholine and a long spacer ligand stabilize the lipid-binding groove of CD1b.

CD1 proteins present lipid antigens to T cells. The antigens are acquired in the endosomal compartments. This raises the question of how the large hydrophobic CD1 pockets are preserved between the moment of biosynthesis in the endoplasmic reticulum and arrival to the endosomes. To address this issue, the natural ligands associated with a soluble form of human CD1b have been investigated. Using isoelectric focusing, native mass spectrometry and resolving the crystal structure at 1.8 A resolution, we found that human CD1b is simultaneously associated with endogenous phosphatidylcholine (PC) and a 41-44 carbon atoms-long spacer molecule. The two lipids appear to work in concert to stabilize the CD1b groove, their combined size slightly exceeding the maximal groove capacity. We propose that the spacer serves to prevent binding of ligands with long lipid tails, whereas short-chain lipids might still displace the PC, which is exposed at the groove entrance. The data presented herein explain how the CD1b groove is preserved, and provide a rationale for the in vivo antigen-binding properties of CD1b.

Antigens, CD1↗
PubMed2006

Synthesis of sulfated galactocerebrosides from an orthogonal beta-D-galactosylceramide scaffold for the study of CD1-antigen interactions.

CD1a protein binds sulfatide (3-O-sulfo-beta-D-galactosylceramide) to form an antigen complex that interacts with T cell receptors and activates T cells. To assess the role of the position of the sulfate in T cell activation, the synthesis of three beta-D-galactosylceramides, variously bearing a sulfate at position 2, 4, or 6 of galactose, has been planned and carried out. The compounds were synthesized by an orthogonal sulfation strategy from a common beta-D-galactosylceramide scaffold, which was in turn obtained through an efficient glycosylation reaction between a fully orthogonally protected galactosyl imidate and 3-O-benzoylazidosphingosine. Immunological evaluation of the three sulfated compounds in CD1a-mediated T cell activation, in comparison with natural sulfatide, provided evidence of the influence of the sulfate position in the recognition event between the antigen, the CD1 protein and the T cell receptor.

Antigens, CD1↗
PubMed2006

Recognition of pollen-derived phosphatidyl-ethanolamine by human CD1d-restricted gamma delta T cells.

BACKGROUND: Evidences from mice and human beings indicate that gammadelta T cells could be relevant in recognition of stress-induced self and/or yet unidentified inhaled foreign antigens. Their specificity differs from classic MHC-restricted alphabeta T cells and involves the immunoglobulin-like structure of the gammadelta T-cell receptor with the recognition of small organic molecules, alkylamines, and self lipid compounds presented by CD1+ dendritic cells. OBJECTIVE: Because CD1 receptors are mainly devoted to lipid antigen presentation, we sought to determine whether exogenous pollen membrane lipids may act as allergens for CD1-restricted gammadelta T cells. METHODS: Peripheral blood and nasal mucosa-associated gammadelta T cells were cloned from normal controls and cypress-sensitive subjects and tested for their antigen specificity and CD1-restriction with phospholipids extracted from tree pollen grains, as well with other natural or synthetic compounds. Phospholipid reactivity of cloned gammadelta T cells was measured by mean of proliferative response and cytokine release as well as by testing their helper activity on IgE production in vitro and in vivo. RESULTS: Cloned gammadelta T lymphocytes from subjects with allergy, but not normal controls, were found to recognize pollen-derived phosphatidyl-ethanolamine (PE) in a CD1d-restricted fashion. Only 16:0/18:2 and 18:2/18:2 PE were stimulatory, whereas no response was recorded for disaturated PE, phosphatidylcholine, neutral lipids, or protein extract. Proliferating clones secreted both T(H)1-type and T(H)2-type cytokines and drove IgE production in vitro and in vivo. CONCLUSION: CD1d-restricted gammadelta T cells specific for phospholipids can represent a key mucosal regulatory subset for the control of early host reactivity against tree pollens. CLINICAL IMPLICATIONS: By knowing how lipid allergen constituents interact with mucosal immune system, we can expand our possibilities in diagnostic and therapeutic interventions.

Antigen Presentation↗
PubMed2006

Synthesis and biological evaluation of alpha-galactosylceramide (KRN7000) and isoglobotrihexosylceramide (iGb3).

Glycoceramides can activate NKT cells by binding with CD1d to produce IFN-gamma, IL-4, and other cytokines. An efficient synthetic pathway for alpha-galactosylceramide (KRN7000) was established by coupling a protected galactose donor to a properly protected ceramide. During the investigation, it was discovered that when the ceramide was protected with benzyl groups, only beta-galactosylceramide was produced from the glycosylation reaction. In contrast, the ceramide with benzoyl protecting groups produced alpha-galactosylceramide. Isoglobotrihexosylceramide (iGb3) was prepared by glycosylation of Galalpha1-3Galbeta1-4Glc donor with 2-azido-sphingosine in high yield. Biological assays on the synthetic KRN7000 and iGb3 were performed using human and murine iNKT cell clones or hybridomas.

Animals↗
PubMed2006

Functional CD1a is stabilized by exogenous lipids.

Self-glycosphingolipids bind to surface CD1 molecules and are readily displaced by other CD1 ligands. This capacity to exchange antigens at the cell surface is not common to other antigen-presenting molecules and its physiological importance is unclear. Here we show that a large pool of cell-surface CD1a, but not CD1b molecules, is stabilized by exogenous lipids present in serum. Under serum deprivation CD1a molecules are altered and functionally inactive, as they are unable to present lipid antigens to T cells. Glycosphingolipids and phospholipids bind to, and restore functionality to CD1a without the contribution of newly synthesized and recycling CD1a molecules. The dependence of CD1a stability on exogenous lipids is not related to its intracellular traffic and rather to its antigen-binding pockets. These results indicate a functional dichotomy between CD1a and CD1b molecules and provide new information on how the lipid antigenic repertoire is immunologically sampled.

Antigens, CD1↗
PubMed2005

Serum lipoproteins: Trojan horses of the immune response?

T cells recognizing lipid antigens presented by CD1 molecules have an important role in the immune response. Several lipid antigens for CD1-restricted T cells have been identified, as have some rules of CD1 loading and CD1-restricted presentation. Little is known, however, about the delivery of lipid antigens from either extracellular compartments or CD1-negative cells to CD1-expressing antigen-presenting cells (APCs). A recent study provides evidence for a role for apolipoprotein E in binding lipid antigens and delivering them to APCs.

Animals↗
PubMed2005

Assistance of microbial glycolipid antigen processing by CD1e.

Complexes between CD1 molecules and self or microbial glycolipids represent important immunogenic ligands for specific subsets of T cells. However, the function of one of the CD1 family members, CD1e, has yet to be determined. Here, we show that the mycobacterial antigens hexamannosylated phosphatidyl-myo-inositols (PIM6) stimulate CD1b-restricted T cells only after partial digestion of the oligomannose moiety by lysosomal alpha-mannosidase and that soluble CD1e is required for this processing. Furthermore, recombinant CD1e was able to bind glycolipids and assist in the digestion of PIM6. We propose that, through this form of glycolipid editing, CD1e helps expand the repertoire of glycolipidic T cell antigens to optimize antimicrobial immune responses.

Acylation↗
PubMed2005

Human CD1-restricted T cell recognition of lipids from pollens.

Plant pollens are an important source of environmental antigens that stimulate allergic responses. In addition to acting as vehicles for foreign protein antigens, they contain lipids that incorporate saturated and unsaturated fatty acids, which are necessary in the reproduction of higher plants. The CD1 family of nonpolymorphic major histocompatibility complex-related molecules is highly conserved in mammals, and has been shown to present microbial and self lipids to T cells. Here, we provide evidence that pollen lipids may be recognized as antigens by human T cells through a CD1-dependent pathway. Among phospholipids extracted from cypress grains, phosphatidyl-choline and phosphatidyl-ethanolamine were able to stimulate the proliferation of T cells from cypress-sensitive subjects. Recognition of phospholipids involved multiple cell types, mostly CD4(+) T cell receptor for antigen (TCR)alphabeta(+), some CD4(-)CD8(-) TCRgammadelta(+), but rarely Valpha24i(+) natural killer-T cells, and required CD1a(+) and CD1d(+) antigen presenting cell. The responding T cells secreted both interleukin (IL)-4 and interferon-gamma, in some cases IL-10 and transforming growth factor-beta, and could provide help for immunoglobulin E (IgE) production. Responses to pollen phospholipids were maximally evident in blood samples obtained from allergic subjects during pollinating season, uniformly absent in Mycobacterium tuberculosis-exposed health care workers, but occasionally seen in nonallergic subjects. Finally, allergic, but not normal subjects, displayed circulating specific IgE and cutaneous weal and flare reactions to phospholipids.

Adult↗
PubMed2005

Ligands for natural killer cell-activating receptors are expressed upon the maturation of normal myelomonocytic cells but at low levels in acute myeloid leukemias.

Natural killer (NK) cell-mediated cytolytic activity against tumors requires the engagement of activating NK receptors by the tumor-associated ligands. Here, we have studied the role of NKG2D and natural cytotoxicity receptors (NCRs) in the recognition of human leukemia. To detect as-yet-unknown cell-surface molecules recognized by NCRs, we developed soluble forms of NKp30, NKp44, and NKp46 as staining reagents binding the putative cognate ligands. Analysis of UL16-binding protein-1 (ULBP1), ULBP2, and ULBP3 ligands for NKG2D and of potential ligands for NKp30, NKp44, and NKp46 in healthy hematopoietic cells demonstrated the ligand-negative phenotype of bone marrow-derived CD34(+) progenitor cells and the acquisition of cell-surface ligands during the course of myeloid differentiation. In acute myeloid leukemia (AML), leukemic blasts from approximately 80% of patients expressed very low levels of ULBPs and NCR-specific ligands. Treatment with differentiation-promoting myeloid growth factors, together with interferon-gamma, upregulated cell-surface levels of ULBP1 and putative NCR ligands on AML blasts, conferring an increased sensitivity to NK cell-mediated lysis. We conclude that the ligand-negative/low phenotype in AML is a consequence of cell maturation arrest on malignant transformation and that defective expression of ligands for the activating NKG2D and NCR receptors may compromise leukemia recognition by NK cells.

Acute Disease↗
PubMed2005

Bacterial infections promote T cell recognition of self-glycolipids.

Recognition of self is essential for repertoire selection, immune regulation, and autoimmunity and may be a consequence of infection. Self-induced recognition may represent the escape mechanism adopted by pathogens but may also incite autoimmune diseases. Here, we show that bacterial infection may promote activation of T cells reactive to self-glycosphingolipids (self-GSL). CD1+ antigen-presenting cells (APCs) infected with bacteria (Escherichia coli, Bacillus subtilis, Staphylococcus aureus, or Mycobacterium bovis-Bacillus Calmette Guerín [BCG]) or treated with the bacterial components lipopolysaccharide, lipoteichoic acid, or Pam3CysSerLys4 (P3CSK4) lipopeptide acquire the capacity to stimulate self-GSL-specific T cells to cytokine release. Immediately after infection, APCs increase the endogenous GSL synthesis and stimulate GSL-specific T cells in a CD1- and T cell receptor (TCR)-dependent manner. This stimulation may contribute to inflammatory responses during bacterial infections and may predispose individuals to autoimmune diseases.

Animals↗
PubMed2005

Recognition of lipid antigens by T cells.

Recent studies have shown that the recognition of lipid antigens by the immune system is important for defence against infection and other diseases, and that lipid-specific responses occur at higher frequencies than previously suspected. Thanks to several recent advances in this field, we now have a better appreciation of the molecular and cellular requirements of T-cell stimulation by lipids. These findings have raised new questions about the mechanisms of lipid presentation, the priming and clonal expansion of lipid-specific T cells, and their differentiation into memory cells. A greater understanding of lipid-specific T cells and the molecular mechanisms of lipid immunogenicity should facilitate the development of lipid-based vaccines.

Animals↗
PubMed2004

The role of innate immunity in autoimmunity.

During the 2004 International Congress of Immunology in Montreal, a panel of experts gathered for an "Ideashop" discussion on the potential role of innate immunity in autoimmunity and the ways in which this might be targeted in future therapies.

Animals↗
PubMed2004

Diacylated sulfoglycolipids are novel mycobacterial antigens stimulating CD1-restricted T cells during infection with Mycobacterium tuberculosis.

Mycobacterial lipids comprise a heterogeneous group of molecules capable of inducing T cell responses in humans. To identify novel antigenic lipids and increase our understanding of lipid-mediated immune responses, we established a panel of T cell clones with different lipid specificities. Using this approach we characterized a novel lipid antigen belonging to the group of diacylated sulfoglycolipids purified from Mycobacterium tuberculosis. The structure of this sulfoglycolipid was identified as 2-palmitoyl or 2-stearoyl-3-hydroxyphthioceranoyl-2'-sulfate-alpha-alpha'-D-trehalose (Ac2SGL). Its immunogenicity is dependent on the presence of the sulfate group and of the two fatty acids. Ac2SGL is mainly presented by CD1b molecules after internalization in a cellular compartment with low pH. Ac2SGL-specific T cells release interferon gamma, efficiently recognize M. tuberculosis-infected cells, and kill intracellular bacteria. The presence of Ac2SGL-responsive T cells in vivo is strictly dependent on previous contact with M. tuberculosis, but independent from the development of clinically overt disease. These properties identify Ac2SGL as a promising candidate to be tested in novel vaccines against tuberculosis.

Antigen Presentation↗