Cutting edge: chromatin remodeling at the IL-4/IL-13 intergenic regulatory region for Th2-specific cytokine gene cluster

During the differentiation of naive Th cells into Th2 effector cells, the entire IL-4/IL-13 locus is remodeled into an accessible chromatin conformation. Here we show that ectopic expression and activation of Stat6 or GATA-3 in Th cells developing under Th1-polarizing conditions lead to the induction of chromatin remodeling not only at the flanking regions of the IL-4 and IL-13 genes but also at the IL-4/IL-13 intergenic regulatory region for the IL-4/IL-13/IL-5 gene cluster. Furthermore, we demonstrate that GATA-3 and another Th2-specific, inducible protein complex interact with the IL-4/IL-13 intergenic DNase I hypersensitive region specifically in Th2 cells.     

Short-term sequential analysis of sex hormones and helper T cells type 1 (Th1) and helper T cells type 2 (Th2) cytokines during and after multiple sclerosis relapse

Multiple sclerosis (MS) is an immune-mediated disease with a clear sex-bias that may be attributed to sex hormones, sex' linked genes or both. Here we sought to determine the evolution pattern of cortisol and sex hormones at MS relapse and 2-months later in 7 male patients with relapsing remitting MS, and whether there was a correlation with a specific Th1 and Th2 cytokine pattern. Our findings indicate the activation of the hypothalamic-pituitary-adrenal axis and the concomitant upregulation of pro- and anti-inflammatory cytokines during relapse. The further increase of sex hormones, in particular estradiol in our male MS patients suggest their possible implication in the physiopathology of the illness and a putative anti-inflammatory and neuroreparatory effect.     

Effect of T helper 1 (Th1)/Th2 cytokine on chemokine-induced dendritic cell functions

Effects of T helper 2 (Th2) and Th1 cytokines, interleukin (IL)-4, and interferon (IFN)-gamma, respectively, on chemokine-induced DC migration and endocytosis are not well understood. We investigated herein the effects of these cytokines on chemokine-induced functions of murine myeloid DCs. As expected, immature DCs markedly migrated to CCL3 but not CCL19, while mature DCs showed vigorous migration in response to CCL19 but not CCL3. Both IL-4 and IFN-gamma significantly decreased CCL3-induced migration of immature DCs. In contrast, these cytokines exerted no significant effects on CCL19-induced migration of mature DCs. Of note, both IL-4 and IFN-gamma markedly enhanced CCL3-induced endocytosis of immature DCs. The messenger RNA level of CCR5, a CCL3 receptor, in immature DCs was slightly increased by IL-4 or IFN-gamma treatment. These results demonstrate that these Th1/Th2 cytokines can act to either inhibit or enhance chemokine-mediated DC functions, and may play a role in increasing antigen uptake by immature DCs at inflammatory sites.     

Impaired GATA3-dependent chromatin remodeling and Th2 cell differentiation leading to attenuated allergic airway inflammation in aging mice

Age-related changes in lymphocytes are most prominent in the T cell compartment. There have been substantial numbers of reports on T cell function in aged mice and humans, such as on the production of Th1 and Th2 cytokines, but the results show considerable variation and contradictions. In the present study, we used 8- to 12-mo-old aging mice and a well-established in vitro Th1/Th2 cell differentiation culture system to identify molecular defects in Th1/Th2 cell differentiation that can be detected in the relatively early stages of aging. The capability to differentiate into Th2 cells is reduced in aging mouse CD4(+) T cells. Decreased activation of the ERK MAPK cascade upon TCR stimulation, but normal intracellular-free calcium ion concentration mobilization and normal IL-4-induced STAT6 activation were observed in aging mouse CD4(+) T cells. In addition, reduced expression of GATA3 was detected in developing Th2 cells. Chromatin remodeling of the Th2 cytokine gene locus was found to be impaired. Th2-dependent allergic airway inflammation was milder in aging mice compared with in young adult mice. These results suggest that the levels of Th2 cell differentiation and resulting Th2-dependent immune responses, including allergic airway inflammation, decline during aging through defects in the activation of the ERK MAPK cascade, expression of GATA3 protein and GATA3-dependent chromatin remodeling of the Th2 cytokine gene locus. In the present study, we provide the first evidence indicating that a chromatin-remodeling event in T cells is impaired by aging.     

Cathepsin-mediated Necrosis Controls the Adaptive Immune Response by Th2 (T helper type 2)-associated Adjuvants

Immunologic adjuvants are critical components of vaccines, but it remains unclear how prototypical adjuvants enhance the adaptive immune response. Recent studies have shown that necrotic cells could trigger an immune response. Although most adjuvants have been shown to be cytotoxic, this activity has traditionally been considered a side effect. We set out to test the role of adjuvant-mediated cell death in immunity and found that alum, the most commonly used adjuvant worldwide, triggers a novel form of cell death in myeloid leukocytes characterized by cathepsin-dependent lysosome-disruption. We demonstrated that direct lysosome-permeabilization with a soluble peptide, Leu-Leu-OMe, mimics the alum-like form of necrotic cell death in terms of cathepsin dependence and cell-type specificity. Using a combination of a haploid genetic screen and cathepsin-deficient cells, we identified specific cathepsins that control lysosome-mediated necrosis. We identified cathepsin C as critical for Leu-Leu-OMe-induced cell death, whereas cathepsins B and S were required for alum-mediated necrosis. Consistent with a role of necrotic cell death in adjuvant effects, Leu-Leu-OMe replicated an alum-like immune response in vivo, characterized by dendritic cell activation, granulocyte recruitment, and production of Th2-associated antibodies. Strikingly, cathepsin C deficiency not only blocked Leu-Leu-OMe-mediated necrosis but also impaired Leu-Leu-OMe-enhanced immunity. Together our findings suggest that necrotic cell death is a powerful mediator of a Th2-associated immune response.     

The Vibrio cholerae cytolysin promotes activation of mast cell (T helper 2) cytokine production

Many strains of Vibrio cholerae produce a cytolysin (VCC) that forms oligomeric transmembrane pores responsible for vacuolization of several cell types in culture. Here we suggest that VCC could contribute to the T helper 2 (Th2) response seen in the natural infection; acting through TLR2, VCC enhances mast cells secretion of IL-4, IL-6 and TNF-α by 330-, 290- and 550-fold respectively. Moreover, VCC-induced cytokine production is dependent on increased cytosolic Ca²⁺ and on the presence of the Src family kinases Lyn and Fyn, known to be required for FcεRI-dependent activation of mast cells. These findings strongly suggest that VCC has a pro-inflammatory activity promoting a Th2-type immune profile.     

Tumor-specific targeting of T helper type 1 (Th1) cells by anti-CD3 × anti-c-ErbB-2 bispecific antibody

T helper type1 (Th1) or type2 (Th2) cells were induced from naive Th cells obtained from ovalbumin-specific T cell receptor (TCR) transgenic mice. Th1 cells producing interferon γ (IFNγ) exhibited stronger antigen-specific cytotoxicity against ovalbumin-(323–339)-peptide-pulsed A20 tumor cells than did Th2 cells. To develop a general method for applying antigen-nonspecific Th1 cells to tumor immunotherapy, we examined the targeting of Th1 cells to tumor cells using a bispecific antibody (bsAb) consisting of anti-(mouse CD3) mAb and anti-(human c-ErbB-2) mAb. When ovalbumin-specific Th1 or Th2 cells were cocultured with c-erbB-2-positive transfectants (CMS7HE), neither type of cell showed significant cytotoxicity or cytokine production in response to tumor cells. However, addition of bsAb resulted in the triggering of both Th1 and Th2 cells. Th1 cells showed higher levels of bsAb-dependent cytotoxicity against CMS7HE tumor cells than did Th2 cells. The targeting of Th1 cells to CMS7HE tumor cells by bsAb also triggered the production of cytokines such as IFNγ, interleukin-2 and tumor necrosis factor α (TNFα). The released TNFα was demonstrated to be a critical cytolytic factor in bsAb-mediated cytotoxicity by Th1 cells. Finally, Th1 cells were demonstrated to show antitumor activity in vivo against human c-erbB-2-positive tumor cells implanted in nude mice. These results suggest that Th1 cells are useful effector cells for the application to adoptive tumor immunotherapy in conjunction with bsAb. Received: 22 April 1999 / Accepted: 2 July 1999     

(−)-Epigallocatechin-3-gallate induces up-regulation of Th1 and Th2 cytokine genes in Jurkat T cells

In the present study, we found that (−)-epigallocatechin-3-gallate (EGCG) significantly up-regulated the mRNA expression of the Th1/Th2 cytokines including IL-2, IFN-γ, IL-5 and IL-13 in Jurkat T cells. The EGCG-induced mRNA up-regulation of IL-2 and IL-5 was predominantly affected by the extracellular signal-regulated protein kinase (ERK) signalling, whereas IL-13 gene expression, the most responsive to the EGCG treatment, was dependent on neither ERK nor c-jun NH₂-terminal kinase (JNK) signalling. IFN-γ gene expression was partially mitigated by both inhibitors of the ERK and JNK pathways. Furthermore, catalase significantly attenuated the intracellular peroxide production, phosphorylation of ERK and JNK, and all cytokine gene expressions induced by EGCG. In addition, physiologically relevant concentrations of both EGCG and H₂O₂-induced up-regulation of IL-5 gene expression. Our findings provide biological evidence that EGCG induces Th1/Th2 cytokine mRNA expression via H₂O₂ production followed by activation of ERK or JNK in Jurkat T cells.     

Effect of suplatast tosilate (IPD-1151T) on cytokine production by allergen-specific human Th1 and Th2 cell lines.

Suplatast tosilate (IPD-1151T) is an antiallergic agent that suppresses airway eosinophil infiltration in asthma. We investigated the effects of IPD-1151T on proliferative response and cytokine production by human antigen-specific T cell lines. Purified protein derivatives (PPD)-specific T helper 1 (Th1) cell lines and Dermatophagoides farinae (Der f)-specific T helper 2 (Th2) cell lines were established from patients with asthma sensitized with house dust mite. Stimulation of PPD-specific and Der f-specific T cell lines with relevant antigens resulted in production of mostly interferon (IFN)-gamma and of interleukin (IL)-4 and IL-5, respectively. IPD-1151T did not inhibit the proliferative responses of either the Th1 or Th2 cell line to antigens. Although IPD-1151T did not inhibit IFN-gamma production by PPD-specific Th1 cell lines, it did inhibit IL-4 and IL-5 production by antigen-stimulated Der f-specific Th2 cell lines in a dose-dependent manner. IPD-1151T directly inhibited cytokine production by Der f-specific Th2 cell lines stimulated with immobilized anti-CD3 antibodies. Although IPD-1151T did not inhibit the clonal expansion of memory T cells among PBMCs into PPD-specific Th1 and Th2 cell lines, it did inhibit IL-4 and IL-5 production by Der f-specific Th2 cell lines but not IFN-gamma production by PPD-specific Th1 cell lines. These results suggest that IPD-1151T selectively inhibits Th2-type cytokine production.     

Differential sensitivity to Itk kinase signals for T helper 2 cytokine production and chemokine-mediated migration.

Allergic asthma is dependent on chemokine-mediated Th2 cell migration and Th2 cytokine secretion into the lungs. The inducible T cell tyrosine kinase Itk regulates the production of Th2 cytokines as well as migration in response to chemokine gradients. Mice lacking Itk are resistant to developing allergic asthma. However, the role of kinase activity of Itk in the development of this disease is unclear. In addition, whether distinct Itk-derived signals lead to T cell migration and secretion of Th2 cytokines is also unknown. Using transgenic mice specifically lacking Itk kinase activity, we show that active kinase signaling is required for control of Th2 responses and development of allergic asthma. Moreover, dominant suppression of kinase Itk activity led to normal Th2 responses, but significantly reduced chemokine-mediated migration, resulting in prevention of allergic asthma. These observations indicate that signals required for Th2 responses and migration are differentially sensitive to Itk activity. Manipulation of Itk's activity can thus provide a new strategy to treat allergic asthma by differentially affecting migration of T cells into the lungs, leaving Th2 responses intact.     

Type I interferon is required for T helper (Th) 2 induction by dendritic cells

Type 2 inflammation is a defining feature of infection with parasitic worms (helminths), as well as being responsible for widespread suffering in allergies. However, the precise mechanisms involved in T helper (Th) 2 polarization by dendritic cells (DCs) are currently unclear. We have identified a previously unrecognized role for type I IFN (IFN‐I) in enabling this process. An IFN‐I signature was evident in DCs responding to the helminth Schistosoma mansoni or the allergen house dust mite (HDM). Further, IFN‐I signaling was required for optimal DC phenotypic activation in response to helminth antigen (Ag), and efficient migration to, and localization with, T cells in the draining lymph node (dLN). Importantly, DCs generated from Ifnar1^?/? mice were incapable of initiating Th2 responses in vivo. These data demonstrate for the first time that the influence of IFN‐I is not limited to antiviral or bacterial settings but also has a central role to play in DC initiation of Th2 responses.     

Distinct gene expression profiles of human type 1 and type 2 T helper cells

Background

The development and activation of CD4⁺ helper T cell (Th) subsets with distinct patterns of unbalanced production of cytokines play an important part in infectious, allergic and autoimmune diseases. Human neonatal cord blood CD4⁺ Th cells can be polarized into type 1 or type 2-like effector cells in vitro by culturing them in the presence of interleukin (IL)-12 or IL-4, respectively. We have exploited this experimental system to identify marker genes that are differentially expressed by polarized Th1 and Th2 cells. An oligonucleotide microarray specifically designed to screen for inflammation-related candidate genes was used and the differential expression was further validated with a quantitative real-time RT-PCR method.

Results

In addition to the previously described marker genes of Th cells, we report subtle changes in the expression of several other genes that represent growth factors, receptors and other signaling molecules in polarized Th1 and Th2 cell subsets. Additionally, we describe a novel set of genes as Th1/Th2 differentiation markers for cells activated by anti-CD3 and anti-CD28 antibodies.

Conclusions

This study demonstrates the power of the targeted use of microarrays in combination with quantitative real-time RT-PCR in identifying and validating new marker genes for gene expression studies.