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F Sinigaglia

Publications and source records attributed to F Sinigaglia.

At least 55 records · Page 3Linked to original sources
PubMed1998

The viral chemokine macrophage inflammatory protein-II is a selective Th2 chemoattractant.

Kaposi's sarcoma (KS) lesions are characterized by a prominent leukocyte infiltrate composed of mononuclear phagocytes and T cells. KS-associated CD4(+) and CD8(+) cells showed predominantly a type II cytokine profile. The CC chemokine viral macrophage inflammatory protein-II (vMIP-II) encoded by the KS-associated herpes virus 8 was a selective chemoattractant for T helper 2 (Th2 cells) and for monocytes, whereas it was inactive on other leukocytes, including Th1 cells, dendritic cells, and natural killer (NK) cells. vMIP-II was an agonist for CCR8, a chemokine receptor selectively expressed on CD4(+) and CD8(+) cells with a type II cytokine profile. Hence, vMIP-II has agonist activity for a chemokine receptor (CCR8), which is preferentially expressed on polarized Th2 cells. The capacity of vMIP-II to attract type II T cells selectively is likely to be a component of the virus strategy to subvert the host immune response.

Cell Movement↗
PubMed1998

Selective up-regulation of chemokine receptors CCR4 and CCR8 upon activation of polarized human type 2 Th cells.

Polarized Th1 and Th2 cells differentially express adhesion molecules and chemokine receptors, endowing these cells with distinct tissue homing capabilities. Here we report that, in contrast to other chemokine receptors, the expression of CCR4 and CCR8 on Th2 cells is transiently increased following TCR and CD28 engagement. IL-4 is not required for this activation-induced up-regulation of CCR4 and CCR8. In accordance with receptor expression, the response of Th2 cells to I-309 (CCR8 ligand) and thymus- and activation-regulated chemokine (CCR4 and CCR8 ligand) is enhanced upon activation. Moreover, activated Th1 cells up-regulate CCR4 expression and functional responsiveness to thymus- and activation-regulated chemokine. Analysis of polarized subsets of CD8+ T cells reveals a similar pattern of chemokine receptor expression and modulation of responsiveness. Taken together, these findings suggest that an up-regulation of CCR4 and CCR8 following Ag encounter may contribute to the proper positioning of activated T cells within sites of antigenic challenge and/or specialized areas of lymphoid tissues.

Calcium↗
PubMed1998

The chemokine receptor CCR8 is preferentially expressed in Th2 but not Th1 cells.

In this paper we report on the cloning and characterization of mouse CCR8. Like its human homologue, it is predominantly expressed in the thymus. In the periphery, murine CCR8 mRNA was found most abundantly expressed in activated Th2-polarized cells and in NK1.1+ CD4+ T cells. Human CCR8 is also preferentially expressed in human Th2-polarized cells and clones. This pattern of expression suggests that CCR8 is part of a Th2-specific gene expression program. The CCR8 ligands I-309 and its mouse homologue T cell activation gene 3 (TCA-3) are potent chemoattractants for Th2-polarized cells. Taken together, these observations strongly suggest that CCR8 plays a role in the control of Th2 responses, and may represent a potential target for treatment of allergic diseases.

Amino Acid Sequence↗
PubMed1998

Relationship between peptide selectivities of human transporters associated with antigen processing and HLA class I molecules.

Efficiency of presentation of a peptide epitope by a MHC class I molecule depends on two parameters: its binding to the MHC molecule and its generation by intracellular Ag processing. In contrast to the former parameter, the mechanisms underlying peptide selection in Ag processing are poorly understood. Peptide translocation by the TAP transporter is required for presentation of most epitopes and may modulate peptide supply to MHC class I molecules. To study the role of human TAP for peptide presentation by individual HLA class I molecules, we generated artificial neural networks capable of predicting the affinity of TAP for random sequence 9-mer peptides. Using neural network-based predictions of TAP affinity, we found that peptides eluted from three different HLA class I molecules had higher TAP affinities than control peptides with equal binding affinities for the same HLA class I molecules, suggesting that human TAP may contribute to epitope selection. In simulated TAP binding experiments with 408 HLA class I binding peptides, HLA class I molecules differed significantly with respect to TAP affinities of their ligands. As a result, some class I molecules, especially HLA-B27, may be particularly efficient in presentation of cytosolic peptides with low concentrations, while most class I molecules may predominantly present abundant cytosolic peptides.

ATP-Binding Cassette Transporters↗
PubMed1998

Cytoskeleton-dependent inhibition of the ADP-ribosyl cyclase activity of CD38 in thrombin-stimulated platelets.

Stimulation of human platelets with thrombin caused a 42% inhibition of the ADP-ribosyl cyclase activity of membrane CD38. This effect was mediated by the activation of the platelet thrombin receptor rather than by proteolysis of CD38, and was not due to a different distribution of the synthesised nucleotide or to a reduced accessibility of CD38 to the substrate. The inhibitory effect of thrombin required actin polymerisation and was not observed when interaction of CD38 with the cytoskeleton was prevented by cytochalasin D. Finally, we analysed whether cADPR could play a role as a Ca2+-mobilising agent in human platelets. Using saponin-permeabilised cells, we found that unlike IP3, cADPR did not induce any release of Ca2+ from intracellular stores. These results indicate that the enzymatic activity of membrane CD38 can be modulated by platelet activation, and that the function of this glycoprotein is probably not related to Ca2+ mobilisation.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1998

Thrombin induces the association of cyclic ADP-ribose-synthesizing CD38 with the platelet cytoskeleton.

The effect of platelet stimulation on the subcellular localization of CD38, a membrane glycoprotein that catalyses the synthesis of cyclic ADP-ribose from beta-NAD+ was investigated. Treatment of human platelets with thrombin caused the association of about 40% of the total ADP-ribosyl cyclase activity with the cytoskeleton, through the translocation of the CD38 molecule from the Triton X-100-soluble to the insoluble fraction. The interaction of CD38 with the cytoskeleton was a specific and reversible process, mediated by the binding to the actin-rich filaments and was inhibited by treatment of platelets with cytochalasin D. This event was regulated by integrin alphaIIb beta3 and platelet aggregation as it was prevented by the inhibition of fibrinogen binding and was not observed in platelets from a patient affected by Glanzmann thrombasthenia. These results demonstrate that the subcellular localization of CD38 can be influenced by platelet stimulation with physiological agonists, and that membrane CD38 can interact with intracellular proteins.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1998

Differential expression of chemokine receptors and chemotactic responsiveness of type 1 T helper cells (Th1s) and Th2s.

T helper cells type 1 (Th1s) that produce interferon-gamma predominantly mediate cellular immune responses and are involved in the development of chronic inflammatory conditions, whereas Th2s which produce large amounts of IL-4 and IL-5 upregulate IgE production and are prominent in the pathogenesis of allergic diseases. The precise factors determining whether Th1- or Th2-mediated immune responses preferentially occur at a peripheral site of antigen exposure are largely unknown. Chemokines, a superfamily of polypeptide mediators, are a key component of the leukocyte recruitment process. Here we report that among four CXC (CXCR1-4) and five CC (CCR1-5) chemokine receptors analyzed, CXCR3 and CCR5 are preferentially expressed in human Th1s. In contrast, Th2s preferentially express CCR4 and, to a lesser extent, CCR3. In agreement with the differential chemokine receptor expression, Th1s and Th2s selectively migrate in response to the corresponding chemokines. The differential expression of chemokine receptors may dictate, to a large extent, the migration and tissue homing of Th1s and Th2s. It may also determine different susceptibility of Th1s and Th2s to human immunodeficiency virus strains using different fusion coreceptors.

Cell Movement↗
PubMed1998

Inhibition of IL-12 production by 1,25-dihydroxyvitamin D3. Involvement of NF-kappaB downregulation in transcriptional repression of the p40 gene.

Interleukin 12 (IL-12), produced by myelomonocytic cells, plays a pivotal role in the development of T helper 1 (Th1) cells, which are involved in the pathogenesis of chronic inflammatory autoimmune disorders. 1,25-Dihydroxyvitamin D3 [1,25(OH)2D3] inhibits IL-12 production by activated macrophages and dendritic cells, thus providing a novel interpretation to its immunosuppressive properties. 1,25(OH)2D3 significantly inhibits mRNA expression for both IL-12 p35 and p40 subunits acting at the transcriptional level. The effect of 1,25(OH)2D3 on p40 promoter activation was analyzed by cotransfecting monocytic RAW264.7 cells with p40 promoter/reporter constructs and expression vectors for vitamin D3 receptor (VDR) and/or retinoid X receptor (RXRalpha). We observed transcriptional repression of the p40 gene by 1,25(OH)2D3, which required coexpression of VDR with RXR and an intact VDR DNA-binding domain. The repressive effect maps to a region in the p40 promoter containing a binding site for NF-kappaB (p40-kappaB). Deletion of the p40-kappaB site abrogates part of the inhibitory effect on the p40 promoter, confirming the functional relevance of this site. Activation of monocytic THP-1 cells in the presence of 1,25(OH)2D3 results in reduced binding to the p40-kappaB site. Thus, 1,25(OH)2D3 may negatively regulate IL-12 production by downregulation of NF-kappaB activation and binding to the p40-kappaB sequence.

Animals↗
PubMed1998

Decreased IL-12 production and Th1 cell development by acetyl salicylic acid-mediated inhibition of NF-kappaB.

IL-12 is a 75-kDa heterodimeric cytokine composed of two covalently linked p35 and p40 chains. This pro-inflammatory cytokine plays a prominent role in the development of Th1 cell-mediated immune responses. Th1 cell-mediated immune responses have been implicated in the pathogenesis of chronic inflammatory autoimmune diseases. Thus, IL-12 appears to be a critical factor in the generation and maintenance of chronic inflammatory conditions. In this study, we investigated the effects of a commonly prescribed anti-inflammatory drug, acetyl salicylic acid (ASA), on IL-12 production and Th1 cell development. ASA was found to inhibit secretion of the IL-12 heterodimer as well as p40 monomer by human monocytic cells. This was associated with the down-regulation of IL-12p40 mRNA expression. Analysis of the regulation of the p40 gene promoter revealed that ASA inhibited NF-kappaB activation and binding to the p40-kappaB site in the p40 promoter, leading to transcriptional repression of the p40 gene. Addition of ASA to an in vitro T helper cell differentiation system, at concentrations compatible with plasma levels reached during anti-inflammatory therapy, resulted in reduced development of Th1 cells. These results suggest that the inhibition of NF-kappaB activation by ASA leads to down-regulation of IL-12 production and inhibition of Th1 cell development.

Animals↗
PubMed1998

Interleukin-12 and its free p40 subunit regulate immune recognition of endometrial cells: potential role in endometriosis.

An alteration of immune recognition and killing of misplaced endometrial cells, refluxed with menstrual debris in ectopic sites, has been claimed to be responsible for the initiation and progression of endometriosis. In particular, current evidence emphasizes the role of natural killer (NK) cells as potential effectors of peritoneal immune surveillance. Interleukin-12 (IL-12), a heterodimeric cytokine composed of p40 and p35 chains, has potent regulatory effects on NK cell growth and function. The purpose of this study was to evaluate whether this cytokine may also have a role in the specific cytolytic NK cell response toward endometrial antigens. To this aim, concentrations of IL-12 and its free p40 subunit were determined in peritoneal fluid of 33 patients with endometriosis and 40 women without laparoscopic evidence of the disease. Similar concentrations of IL-12, but significantly higher levels of free p40, were present in peritoneal fluid of patients with endometriosis compared to those in women without the disease. We also observed that the IL-12 plus free p40/IL-12 ratio increased with the severity of the disease. Moreover, we investigated whether incubation of NK cells with heterodimeric IL-12 and/or p40 has any effect on NK cell-mediated lysis of endometrial cells. NK cells pretreated with heterodimeric IL-12 exhibited an enhanced cytotoxic response toward endometrial targets. This IL-12-induced cytotoxicity could be abrogated by the p40 subunit in a specific and dose-dependent manner. The p40 inhibitory effect was mediated by down-regulation of IL-12 high affinity binding sites on NK cells, as we observed inhibition of surface IL-12 receptor beta1-chain expression, a decrease in IL-12-binding capacity, and inhibition of phosphorylation of STAT4 (signal transducer and activator of transcription) protein. These data suggest that the excess of p40 present in peritoneal fluid of patients with endometriosis may be related to the NK cell defect associated with the disease. Moreover, IL-12 could be a potential specific agent able to correct the p40-induced defect in vivo.

Ascitic Fluid↗
PubMed1997

A novel approach to human anti-HLA mABs production: use of phage display libraries.

The production of human monoclonal antibodies was previously limited to very laborious and time-consuming processes involving EBV-transformation and/or hybridoma generation. Due to the development of molecular cloning techniques, it is now possible to produce human monoclonal antibody fragments quickly by panning phage display libraries against predefined antigenic specificities. Therefore, we tested this technology for producing human single chain Fv fragments (scFvs) against HLA-DR1 purified molecules immobilized on solid phase. Enrichment of DR1-specific phages was measured through five selection rounds of a synthetic library and revealed a 100-fold amplification. Soluble antibody fragments were then expressed and 7 out of 48 clones were found to secrete scFvs which specifically bind to DR1 molecules in ELISA. Further analysis revealed binding of the scFvs also to DR3 but not to DR5 or DR7 molecules correlating with the presence of particular polymorphic aminoacid residues in the DR beta chain. Western blot analysis indicated that the 7 scFvs react with the DR1 alpha/beta-dimer but not with free alpha- or beta- chains. This study shows that the innovative approach of phage display libraries can efficiently provide scFv fragments as useful reagents for the identification and dissection of HLA polymorphic epitopes.

Antibodies, Monoclonal↗
PubMed1997

Beta2-agonists prevent Th1 development by selective inhibition of interleukin 12.

Interleukin 12 (IL-12) plays a central role in the immune system by skewing the immune response towards T helper 1 (Th1) type responses which are characterized by high interferon-gamma and low IL-4 production. In this report we present evidence that beta2-agonists inhibit IL-12 production by both human monocytes in response to lipopolysaccharide (LPS) and dendritic cells stimulated via CD40. Inhibition of IL-12 production is selective, as other cytokines produced by monocytes are unaffected. IL-12 inhibition is dependent on beta2-adrenoceptor stimulation and correlates with increased levels of intracellular cAMP. In conjunction with their ability to suppress IL-12 production, when beta2-agonists are added at priming of neonatal T lymphocytes, they inhibit the development of Th1-type cells, while promoting T helper 2 (Th2) cell differentiation. Further, the in vivo administration of a therapeutic dose of salbutamol results in the selective inhibition of IL-12 production by whole blood lymphocytes stimulated in vitro with LPS. These findings provide new insight into the immunological consequences of the clinical use of beta2-agonists and may suggest new approaches for the treatment of Th1-mediated diseases.

Adrenergic beta-2 Receptor Agonists↗
PubMed1997

Binding of malaria T cell epitopes to DR and DQ molecules in vitro correlates with immunogenicity in vivo: identification of a universal T cell epitope in the Plasmodium falciparum circumsporozoite protein.

The efficacy of a malaria peptide vaccine would be enhanced by the inclusion of a parasite-derived universal T cell epitope to ensure that all vaccinees develop parasite-specific cellular and humoral immunity. Two circumsporozoite (CS) protein T cell epitopes, previously identified by CD4+ T cell clones derived from Plasmodium falciparum sporozoite-immunized volunteers, were studied to determine their HLA class II binding potential. One epitope, located in amino acid (aa) 326-345 of the P. falciparum (NF54 strain) CS protein, was "universal" in that it could bind to multiple DR and DQ molecules in vitro. In contrast, the second epitope, T1, which is located in the CS repeat region, was recognized by T cells in the context of DQ6 (DQB1*0603) and did not bind with high affinity to any of the class II molecules tested in the peptide binding assays. The in vitro patterns of peptide/HLA interactions correlated with immunogenicity in vivo. A multiple antigen peptide (MAP) containing the aa 326-345 epitope elicited responses in eight inbred strains (H-2(a,b,d,k,p,q,r,s)), while the T1 MAP was recognized by only a single haplotype, H-2b. The combination of the universal aa 326-345 T cell epitope and the T1 repeat in a di-epitope MAP overcame the genetic restriction to the P. falciparum CS repeat region and elicited antisporozoite Ab responses in all of the MAP-immunized mice. Synthetic peptide malaria vaccines containing the aa 326-345 universal T cell epitope would be expected to elicit parasite-specific immune responses in both sporozoite-primed and naive individuals of diverse genetic backgrounds.

Animals↗
PubMed1997

Different modes of peptide interaction enable HLA-DQ and HLA-DR molecules to bind diverse peptide repertoires.

The role of HLA-DQ molecules in Ag presentation has, thus far, remained elusive. Here we report that two DQ allotypes, DQ7 (DQA1*0501/B1*0301) and DQ9 (DQA1*0201/B1*0303), are capable of binding peptide repertoires in complementarity with DR molecules. The results reflect fundamental differences in the binding modes of these two HLA class II isotypes, in that DQ7 and DQ9 but not DR molecules appear to have the capacity to bind peptide structures without type 1-like anchor residues. Consistent with this is our observation that none of the amino acid side chains of the class II-associated invariant chain peptides (CLIP) are required for association with DQ7 and DQ9, even though many of them are essential for CLIP-DR interaction. Together, these data reveal a functional complementarity of HLA-DR and -DQ molecules in Ag presentation.

Amino Acid Sequence↗
PubMed1997

Selective expression of an interleukin-12 receptor component by human T helper 1 cells.

Interleukin-12 (IL-12), a heterodimeric cytokine produced by activated monocytes and dendritic cells, plays a crucial role in regulating interferon (IFN)-gamma production and in the generation of IFN-gamma-producing T helper 1 (Th1) cells. Here we show that the IL-12 receptor (IL-12R) beta 2 subunit, a recently cloned binding and signal transducing component of the IL-12R, is expressed on human Th1 but not Th2 clones and is induced during differentiation of human naive cells along the Th1 but not the Th2 pathway. IL-12 and type I but not type II interferons induce expression of the IL-12R beta 2 chain during in vitro T cell differentiation after antigen receptor triggering. The selective expression and regulation of the IL-12R beta 2 subunit may help to understand the basis of Th1/Th2 differentiation and may provide therapeutic options for altering the Th1/Th2 balance in several immuno-pathological conditions such as autoimmune diseases and allergies.

Cell Differentiation↗