Eddy A. Wierenga

Possible mechanism involving T-lymphocyte response to non-structural protein 3 in viral clearance in acute hepatitis C virus infection

In acute hepatitis C virus (HCV) infection only 20-50% of patients spontaneously clear the virus. To characterise the immune reaction during that phase we studied the response of peripheral blood mononuclear cells (PBMC) to the recombinant HCV proteins core, non-structural protein 3 (NS3), NS4, and NS5 in 14 patients with acute hepatitis C. All eight patients with self-limited disease compared with two of six with evolving chronic infection showed an NS3- specific PBMC response (p = 0.015). Of 65 patients with established chronic hepatitis C, five showed a PBMC response to NS3. NS3-specific CD4 T-cell clones from patients with self-limited infection predominantly produced interferon-gamma and may thus support cytotoxic effector mechanisms important for viral clearance.

Microbial Compounds Selectively Induce Th1 Cell-Promoting or Th2 Cell-Promoting Dendritic Cells In Vitro with Diverse Th Cell-Polarizing Signals

Upon microbial infection, specific Th1 or Th2 responses develop depending on the type of microbe. Here, we demonstrate that different microbial compounds polarize the maturation of human myeloid dendritic cells (DCs) into stably committed Th1 cell-promoting (DC1) or Th2 cell-promoting (DC2) effector DCs that polarize Th cells via different mechanisms. Protein extract derived from the helminth Schistosoma mansoni induced the development of DC2s that promote the development of Th2 cells via the enhanced expression of OX40 ligand. Likewise, toxin from the extracellular bacterium Vibrio cholerae induced development of DC2s as well, however, via an OX40 ligand-independent, still unknown mechanism. In contrast, toxin from the intracellular bacterium Bordetella pertussis induced the development of DC1s with enhanced IL-12 production, which promotes a Th1 cell development. Poly(I:C) (dsRNA, mimic for virus) induced the development of extremely potent Th1-inducing DC1, surprisingly, without an enhanced IL-12 production. The obtained DC1s and DC2s are genuine effector cells that stably express Th cell-polarizing factors and are unresponsive to further modulation. The data suggest that the molecular basis of Th1/Th2 polarization via DCs is unexpectedly diverse and is adapted to the nature of the microbial compounds.

Functional subsets of allergen-reactive human CD4+ T cells

After a period of resistance, the concept of human helper T (TH)-cell subsets is gaining currency. This is the result of analyses from a number of laboratories on the cytokine profiles of T-cell clones prepared from chronically-infected and hypersensitive individuals. Here, Martien Kapsenberg and colleagues summarize these studies and speculate on the significance of skewed TH1 and TH2 responses.

IL-4 Is a Mediator of IL-12p70 Induction by Human Th2 Cells: Reversal of Polarized Th2 Phenotype by Dendritic Cells

IL-12 is a key inducer of Th1-associated inflammatory responses, protective against intracellular infections and cancer, but also involved in autoimmune tissue destruction. We report that human Th2 cells interacting with monocyte-derived dendritic cells (DC) effectively induce bioactive IL-12p70 and revert to Th0/Th1 phenotype. In contrast, the interaction with B cells preserves polarized Th2 phenotype. The induction of IL-12p70 in Th2 cell-DC cocultures is prevented by IL-4-neutralizing mAb, indicating that IL-4 acts as a Th2 cell-specific cofactor of IL-12p70 induction. Like IFN-γ, IL-4 strongly enhances the production of bioactive IL-12p70 heterodimer in CD40 ligand-stimulated DC and macrophages and synergizes with IFN-γ at low concentrations of both cytokines. However, in contrast to IFN-γ, IL-4 inhibits the CD40 ligand-induced production of inactive IL-12p40 and the production of either form of IL-12 induced by LPS, which may explain the view of IL-4 as an IL-12 inhibitor. The presently described ability of IL-4 to act as a cofactor of Th cell-mediated IL-12p70 induction may allow Th2 cells to support cell-mediated immunity in chronic inflammatory states, including cancer, autoimmunity, and atopic dermatitis.

Glatiramer Acetate (Copolymer-1, Copaxone) Promotes Th2 Cell Development and Increased IL-10 Production Through Modulation of Dendritic Cells

Glatiramer acetate (GA; copolymer-1, Copaxone) suppresses the induction of experimental autoimmune encephalomyelitis and reduces the relapse frequency in relapsing-remitting multiple sclerosis. Although it has become clear that GA induces protective degenerate Th2/IL-10 responses, its precise mode of action remains elusive. Because the cytokine profile of Th cells is often regulated by dendritic cells (DC), we studied the modulatory effects of GA on the T cell regulatory function of human DC. This study shows the novel selective inhibitory effect of GA on the production of DC-derived inflammatory mediators without affecting DC maturation or DC immunostimulatory potential. DC exposed to GA have an impaired capacity to secrete the major Th1 polarizing factor IL-12p70 in response to LPS and CD40 ligand triggering. DC exposed to GA induce effector IL-4-secreting Th2 cells and enhanced levels of the anti-inflammatory cytokine IL-10. The anti-inflammatory effect of GA is mediated via DC as GA does not affect the polarization patterns of naive Th cells activated in an APC-free system. Together, these results reveal that APC are essential for the GA-mediated shift in the Th cell profiles and indicate that DC are a prime target for the immunomodulatory effects of GA.

Commensal Gram-negative bacteria prime human dendritic cells for enhanced IL-23 and IL-27 expression and enhanced Th1 development

Dendritic cells (DC) are the main orchestrators of specific immune responses. Depending on microbial information they encounter in peripheral tissues, they promote the development of Th1, Th2 or unpolarized Th cell responses. In this study we have investigated the immunomodulatory effect of non‐pathogenic intestinal Gram‐negative (Escherichia coli, Bacteroides vulgatus,Veillonella parvula, Pseudomonas aeruginosa) and Gram‐positive (Bifidobacterium adolescentis, Enteroccocus faecalis, Lactobacillus plantarum and Staphylococcus aureus) bacteria on human monocyte‐derived DC (moDC). None of the Gram‐positive bacteria (GpB) primed for Th1 or Th2 development. In contrast, despite the low levels of IL‐12 they induce, all Gram‐negative bacteria (GnB) primed moDC for enhanced Th1 cell development, which was dependent on IL‐12 and an additional unidentified cofactor. Strikingly, GnB‐matured moDC expressed elevated levels of p19 and p28 mRNA, the critical subunits of IL‐23 and IL‐27, respectively, suggesting that the IL‐12 family members may jointly be responsible for their Th1‐driving capacity. Purified majorcell wall components of either GnB or GpB did not yield Th cell profiles identical to those obtained with whole bacteria, and could not explain the induction of the IL‐12 family members nor Th1 priming by GnB. Importantly, this study gives indications that the expression of the different IL‐12 family members is dictated by different priming conditions of immature DC.

Intercellular Adhesion Molecule-1/LFA-1 Ligation Favors Human Th1 Development

Th cell polarization toward Th1 or Th2 cells is strongly driven by exogenous cytokines, in particular IL-12 or IL-4, if present during activation by Ag-presenting dendritic cells (DC). However, additional Th cell polarizing mechanisms are induced by the ligation of cell surface molecules on DC and naive Th cells. In the present study, the role of LFA-1/ICAM-1 ligation in human Th cell polarization was investigated. Triggering of LFA-1 on anti-CD3/CD28 stimulated naive Th cells with immobilized Fc-ICAM-1, in the absence of DC and exogenous cytokines, induced a marked shift toward Th1 cell development, accompanied by a dose-dependent decrease in GATA-3 expression and a dose-dependent increase in T-bet expression. Th1 polarization by LFA-1 ligation could be demonstrated only under low cytokine conditions, as it was largely overruled by IL-12 or IL-4. This IL-12-independent Th1-driving mechanism appears to be operated by certain subsets of effector DC. Maturation of DC by poly(I:C), a synthetic dsRNA, used as an in vitro model for viral infections, leads to the generation of Th1-driving effector DC (DC1), which express elevated levels of ICAM-1 but produce only low levels of IL-12p70. Blocking the ICAM-1/LFA-1 interaction in cocultures of these DC with naive Th cells attenuated their Th1-driving capacity. The molecular mechanism by which LFA-1 signaling supports Th1 differentiation is blocked by specific inhibitors of extracellular signal-regulated kinase phosphorylation. The present data indicate the existence of an IL-12-independent, extracellular signal-regulated kinase-mediated mechanism, through which high ICAM-1-expressing DC1 can drive Th1 polarization. This mechanism may be operational during viral infections.

IL-12-induced reversal of human Th2 cells is accompanied by full restoration of IL-12 responsiveness and loss of GATA-3 expression.

IL‐12 is a potent inducer of IFN‐γ production and drives the development of Th1 cells. Human polarized Th2 cells do not express the signaling β2‐subunit of the IL‐12R and, therefore, do not signal in response to IL‐12. The question was raised as to what extent the loss of the IL‐12Rβ2 chain in Th2 cells has bearing on the stability of the human Th2 phenotype. In the present report, we show that restimulation of human fully polarized Th2 cells in the presence of IL‐12 primes for a shift towards Th0/Th1 phenotypes, accompanied by suppression of GATA‐3 expression and induction of T‐bet expression. These reversed cells are further characterized by a marked IL‐12Rβ2 chain expression and fully restored IL‐12‐inducible STAT4 activation. The IL‐12‐induced phenotypic shift proved to be stable as a subsequent restimulation in the presence of IL‐4 and in the absence of IL‐12 could not undo the accomplished changes. Identical results were obtained with cells from atopic patients, both with polyclonal Th2 cell lines and allergen‐specific Th2 cell clones. These findings suggest the possibility of restoring IL‐12 responsiveness in established Th2 cells of atopic patients by stimulation in the presence of IL‐12, and that IL‐12‐promoting immunotherapy can be beneficial for Th2‐mediated immune disorders, targeting both naive and memory effector T cells.

CD26/DPPIV signal transduction function, but not proteolytic activity, is directly related to its expression level on human Th1 and Th2 cell lines as detected with living cell cytochemistry.

CD26/DPPIV is a cell surface glycoprotein that functions both in signal transduction and as a proteolytic enzyme, dipeptidyl peptidase IV (DPPIV). To investigate how two separate functions of one molecule are regulated, we analyzed CD26 protein expression and DPPIV enzyme activity on living human T-helper 1 (Th1) and Th2 cells that express different levels of CD26/DPPIV. DPPIV activity was specifically determined with the synthetic fluorogenic substrate ala-pro-cresyl violet and CD26 protein expression was demonstrated with an FITC-conjugated CD26-specific antibody. Fluorescence of liberated cresyl violet (red) and FITC (green) was detected simultaneously on living T-cells using flow cytometry and spectrofluorometry. Th1 cells expressed three- to sixfold more CD26 protein than Th2 cells. The signal transduction function of the CD26/DPPIV complex, tested by measuring its co-stimulatory potential for proliferation, was directly related to the amount of CD26 protein at the cell surface. However, DPPIV activity was similar in both cell populations at physiological substrate concentrations because of differences in Km and Vmax values of DPPIV on Th1 and Th2 cells. Western blotting and zymography of Th1 and Th2 whole-cell lysates demonstrated similar patterns. This study shows that two functions of one molecule can be controlled differentially.

The role of antigen-presenting cells in the regulation of allergen-specific T cell responses

Allergic reactions in atopic patients follow from a generalized enhanced polarization of Th cells, predominantly imposed by factors derived from antigen-presenting cells from a pathogen-stressed tissue; these sample information not only on antigen structures but also on the nature of the stress. Antigen-presenting cells of atopic individuals show aberrant characteristics which, through a highly interactive communication network, play an active role in aberrant Th-cell polarization. This generalized bias may follow from intrinsic abnormalities of antigen-presenting cells and also from a low degree of cross-regulation by micro-organisms.