The contribution of T helper (Th) and T cytotoxic (Tc) type 1 lymphocytes in the expression of allergic contact dermatitis to haptens has been amply documented. Conversely, the existence of T cell-based regulatory mechanisms has been poorly investigated. Here, we examined the properties of a subset of nickel-specific CD4+ T cells displaying the cytokine profile (IL-10 , IL-5 , IFN-gamma+/-, IL-4+/-) of T regulatory cells 1 (Tr1) and with the potential to down-modulate immune responses to nickel. Tr1 clones were isolated from skin challenged with NiSO4 and peripheral blood of nickel-allergic patients, and from the blood of healthy individuals. Tr1 clones expressed CD25, CD28, CD30, CD26, and the IL-12 receptor beta2 chain upon activation, whereas the lymphocyte activation antigen-3 was present on 50% of the clones. Monocytes precultured with Tr1 cells in the presence of nickel, or treated with Tr1-derived supernatant, exhibited a markedly diminished capacity to stimulate nickel-specific Th1 responses. Tr1 supernatants also blocked the differentiation of dendritic cells (DC) from monocytes, as well as DC maturation and IL-12 production induced by lipopolysaccharide. As a consequence, the ability of DC to stimulate nickel-specific Th1 and Tc1 responses was greatly impaired. These inhibitory effects were completely prevented by IL-10, but not IL-5, neutralization. In aggregate, the results indicate that Tr1 cells can potently regulate the expression of Th1-mediated allergic diseases via release of IL-10.

The present study demonstrates that the quality of the virus‐specific CD8+ T cell responses, as detected by both enzyme‐linked immunospot assay and specific MHC‐peptide tetramers, changed in relation to the different disease activity in chronically hepatitis C virus‐infected patients. Indeed, both the serum alanine transaminase and the hepatic flogosis levels were related directly to the frequencies of peripheral memory effector CD8+ T cells producing IFN‐γ (Tc1), but inversely to the frequencies of those producing both IL‐4 and IL‐10 (Tc2). Longitudinal studies highlighted that Tc1 or Tc2 responses fluctuate in relation to the different phases of the disease in the same individual. Furthermore, the Tc1 or Tc2 phenotype correlates with tetramer‐positive cells expressing either CXCR3 or CCR3, promoting differential tissue localization of these cells and the maintenance of T cell homeostasis. Finally, studies at the level of liver‐infiltrating lymphocytes indicated that they produced both IFN‐γ and IL‐4 with an evident bias towards the Tc1‐like phenotype. Our studies suggest that the progressive fluctuation of Tc1 and Tc2 responses may play a fundamental role in maintaining a long‐lasting low‐level liver inflammation, and may constitute the basis for new therapeutic strategies of immune regulation.

Phage display selection strategies rely on the physical link between the displayed heterologous protein ligand and the DNA encoding it. Thus, genes expressing a ligand with a specific binding affinity can be selected rapidly. To improve the specificity and sensitivity of this technology for potential use in identifying ligands to a specific antibody present in a complex mixture, we incorporated a DNA selection step along with the phage display technology. Ligands for hepatitis C virus (HCV) antibodies present in serum were identified by panning a phage-displayed random peptide library against pools of serum HCV antibodies. An additional DNA hybridization screening step using single-stranded DNA isolated from one of the pools increased the specificity and sensitivity, resulting in the selection of an HCV antibody ligand with diagnostic potential.

Bacteria are being actively investigated as vaccine carriers for inducing or boosting protective immune responses. In this study, human monocyte-derived dendritic cells (DCs) and normal B cells were compared for their capacity to present the C fragment of tetanus toxin (TTFC), expressed on the surface of recombinant Streptococcus gordonii, to specific CD4 ؉ T lymphocytes. DCs were more efficient than B cells at presenting soluble TTFC and remarkably more capable of presenting bacterium-associated TTFC both in terms of the amount of antigen required to obtain a given T-cell response and on a per-cell basis. This difference was associated with a much lower capacity of B cells to endocytose soluble TTFC and phagocytose recombinant S. Recent efforts in developing more efficacious vaccine strategies have pointed to the use of bacteria as vectors of heterologous antigens. Bacteria can be easily manipulated at the genetic level and can be engineered to express the gene product in different forms (11,21,26). Alternatively, attenuated bacteria can serve as carriers for delivering antigen-encoding DNA to antigen-presenting cells (APCs) (6,17,30). These approaches have been used for inducing protective immune responses against viral and tumor antigens in mouse models (6,17,26). In particular, recombinant strains of Streptococcus gordonii, which is a commensal bacterium of the human oral cavity, induce both local and systemic antibody responses, as well as a T-cell response, to viral antigens in both mice and macaques (7,15,16). However, the APCs and the mechanisms involved in these bacterium-based immunostimulating systems have been only marginally investigated.Dendritic cells (DCs) and B lymphocytes show important differences in their APC functions (1, 19). They differ in the capacity to endocytose antigens, to cluster with T cells, to provide proper costimulation, and to secrete regulatory cytokines (2,5,9,13,28). A number of studies have indeed demonstrated that DCs are much more potent APCs than are antigen-specific and non-antigen-specific B lymphocytes in the activation of both naive and memory T cells (2,4,9,14,20,27). Most experiments, however, has been performed using soluble antigens given in the form of native proteins or immunogenic peptides, and very few studies have compared the capacity of DCs and B cells to present particulate or bacterium-associated antigens (8,23,29).Mouse DCs present a major histocompatibility complex (MHC) class I-restricted heterologous antigen, expressed on the surface of recombinant S. gordonii, to T lymphocytes at high efficiency, and S. gordonii induces neobiosynthesis and membrane stabilization of MHC class I and class II molecules (25). Moreover, human DCs fed with recombinant bacteria stimulated the specific CD4 ϩ -T-cell response with much higher efficiency than did DCs pulsed with soluble antigen. Finally, S. gordonii provided a potent stimulus for inducing DC maturation and release of chemokines active on T cells (3). In the present study, DCs and B cells were compared for th...

Disease-specific epitope discovery from random peptide libraries displayed on phage using sera from patients involves a number of screening steps with many immune and non-immune sera. To rapidly identify mimotopes of the human hepatitis C virus (HCV) core protein, we have used an anti-core human monoclonal antibody (mAb; B12.F8) as a probe in screening phage that were affinity-selected using a serum from an HCV infected patient. Three different positive phage were isolated displaying low or no homology with the natural antigen, but which still efficiently bound to the antigen binding site of the B12.F8 antibody. Testing the reactivity of these phage with forty-five sera from HCV infected patients showed that antibodies recognizing them are present in more than 80% of this population. These antibodies showed distinct fine specificity, as they bound the selected phage in a mutually exclusive fashion. Co-expression of two mimotopes in the same cells led to chimeric particles which were recognized by antibodies of different specificity. These data provide novel information on the potential use of the phage display technology for the characterization of antibody specificity as well as disease diagnosis and prevention.