Immunophenotyping of Circulating T Helper Cells Argues for Multiple Functions and Plasticity of T Cells In Vivo in Humans - Possible Role in Asthma

BACKGROUND: The immune process driving eosinophilic and non-eosinophilic asthma is likely driven by different subsets of T helper (Th) cells. Recently, in vitro studies and animal studies suggest that Th cell subsets displays plasticity by changing their transcription factor or by expressing multiple transcription factors. Our aim was to determine whether individuals with asthma and elevated circulating eosinophils express signs of different regulatory immune mechanisms compared with asthmatics with low blood eosinophils and non-asthmatic control subjects. In addition, determine the relationship between eosinophilia and circulating Th cell subsets. METHODOLOGY/PRINCIPAL FINDINGS: Participants were selected from a random epidemiological cohort, the West Sweden Asthma Study. Immunophenotypes of fresh peripheral blood cells obtained from stable asthmatics, with and without elevated eosinophilic inflammation (EOS high and EOS low respectively) and control subjects, were determined by flow cytometry. No differences in the number of Th1 (T-bet), Th2 (GATA-3), Th17 (RORγt) or Treg (FOXP3) cells were observed between the groups when analysing each subset separately. However, in all groups, each of the Th subsets showed expression of additional canonical transcription factors T-bet, GATA-3, RORγt and FOXP3. Furthermore, by in vitro stimulation with anti-CD3/anti-CD28 there was a significant increase of single expressing GATA-3(+) and co-expressing T-bet(+)GATA-3(+) cells in the EOS high asthmatics in comparison with control subjects. In addition, T-bet(-)GATA-3(+)RORγt(+)FOXP3(+) were decreased in comparison to the EOS low asthmatics. Finally, in a group of control subjects we found that the majority of proliferating Th cells (CD4(+)CD25(+)Ki67(+)) expressed three or four transcription factors. CONCLUSIONS: The ability of human Th cells to express several regulatory transcription factors suggests that these cells may display plasticity in vivo

Tissue Specific Deletion of Inhibitor of Kappa B Kinase 2 with OX40-Cre Reveals the Unanticipated Expression from the OX40 Locus in Skin Epidermis

NF-κB signalling plays an essential role in T cell activation and generation of regulatory and memory populations in vivo. In the present study, we aimed to investigate the role of NF-κB signalling in post-activation T cells using tissue specific ablation of inhibitor of kappa-B kinase 2 expression, an important component of the inhibitor of kappa-B kinase complex in canonical NF-κB signalling. The OX40 antigen is expressed on activated T cells. Therefore, we used previously described mouse strain expressing Cre recombinase from the endogenous OX40 locus. Ablation of IKK2 expression using OX40Cre mice resulted in the development of an inflammatory response in the skin epidermis causing wide spread skin lesions. The inflammatory response was characterised by extensive leukocytic infiltrate in skin tissue, hyperplasia of draining lymph nodes and widespread activation in the T cell compartment. Surprisingly, disease development did not depend on T cells but was rather associated with an unanticipated expression of Cre in skin epidermis, and activation of the T cell compartment did not require Ikbk2 deletion in T cells. Employment of Cre reporter strains revealed extensive Cre activity in skin epidermis. Therefore, development of skin lesions was rather more likely explained by deletion of Ikbk2 in skin keratinocytes in OX40Cre mice

The Role of T cell PPAR γ in mice with experimental inflammatory bowel disease

<p>Abstract</p> <p>Background</p> <p>Peroxisome proliferator-activated receptor γ (PPAR γ) is a nuclear receptor whose activation has been shown to modulate macrophage and T cell-mediated inflammation. The objective of this study was to investigate the mechanisms by which the deletion of PPAR γ in T cells modulates immune cell distribution and colonic gene expression and the severity of experimental IBD.</p> <p>Methods</p> <p>PPAR γ flfl; CD4 Cre⁺(CD4cre) or Cre- (WT) mice were challenged with 2.5% dextran sodium sulfate in their drinking water for 0, 2, or 7 days. Mice were scored on disease severity both clinically and histopathologically. Flow cytometry was used to assess lymphocyte and macrophage populations in the blood, spleen, and mesenteric lymph nodes (MLN). Global gene expression in colonic mucosa was profiled using Affymetrix microarrays.</p> <p>Results</p> <p>The deficiency of PPAR γ in T cells accelerated the onset of disease and body weight loss. Examination of colon histopathology revealed significantly greater epithelial erosion, leukocyte infiltration, and mucosal thickening in the CD4cre mice on day 7. CD4cre mice had more CD8⁺T cells than WT mice and fewer CD4⁺FoxP3⁺regulatory T cells (Treg) and IL10⁺CD4⁺T cells in blood and MLN, respectively. Transcriptomic profiling revealed around 3000 genes being transcriptionally altered as a result of DSS challenge in CD4cre mice. These included up-regulated mRNA expression of adhesion molecules, proinflammatory cytokines interleukin-6 (IL-6) and IL-1β, and suppressor of cytokine signaling 3 (SOCS-3) on day 7. Gene set enrichment analysis (GSEA) showed that the ribosome and Krebs cycle pathways were downregulated while the apoptosis pathway was upregulated in colons of mice lacking PPAR γ in T cells.</p> <p>Conclusions</p> <p>The expression of PPAR γ in T cells is involved in preventing gut inflammation by regulating colonic expression of adhesion molecules and inflammatory mediators at later stages of disease while favoring the recruitment of Treg to the mucosal inductive sites.</p

Effects of 15-Deoxy-Δ12,14-Prostaglandin J2 (15d-PGJ2) and Rosiglitazone on Human Vδ2+ T Cells

BACKGROUND:Thiazolidinediones (TZD) class of drugs, and 15-deoxy-D12,14-prostaglandin J2 (15d-PGJ2) are immune regulators predicted to modulate human autoimmune disease. Their effects on gammadelta T cells, which are involved in animal model and human and animal autoimmune diseases, are unknown. METHODOLOGY/PRINCIPAL FINDINGS:We characterized the activity of rosiglitazone (from the TZD class of drugs) and 15d-PGJ2 in human Vdelta2 T cells. We found that 15d-PGJ2 and rosiglitazone had different effects on Vdelta2 T cell functions. Both 15d-PGJ2 and rosiglitazone suppressed Vdelta2 T cell proliferation in response to IPP and IL2. However, only 15d-PGJ2 suppressed functional responses including cytokine production, degranulation and cytotoxicity against tumor cells. The mechanism for 15d-PGJ2 effects on Vdelta2 T cells acts through inhibiting Erk activation. In contrast, rosiglitazone did not affect Erk activation but the IL2 signaling pathway, which accounts for rosiglitazone suppression of IL2-dependent, Vdelta2 T cell proliferation without affecting TCR-dependent functions. Rosiglitazone and 15d-PGJ2 are designed to be peroxisome proliferator-activated receptor gamma (PPARgamma) ligands and PPARgamma was expressed in Vdelta2 T cell. Surprisingly, when PPARgamma levels were lowered by specific siRNA, 15d-PGJ2 and rosiglitazone were still active, suggesting their target of action induces cellular proteins other than PPARgamma. CONCLUSIONS/SIGNIFICANCE:The current findings expand our understanding of how the immune system is regulated by rosiglitazone and 15d-PGJ2 and will be important to evaluate these compounds as therapeutic agents in human autoimmune disease

Inhibition of Effector Function but Not T Cell Activation and Increase in FoxP3 Expression in T Cells Differentiated in the Presence of PP14

Background: T-helper polarization of naïve T cells is determined by a complex mechanism that involves many factors, eventually leading to activation of Th1, Th2, or Th17 responses or alternatively the generation of regulatory T cells. Placental Protein 14 (PP14) is a 28 kDa glycoprotein highly secreted in early pregnancy that is able to desensitize T cell receptor (TCR) signaling and modulate T cell activation. Methodology/Principal Findings: Prolonged antigen-specific stimulation of T cells in the presence of PP14 resulted in an impaired secretion of IFN-c, IL-5 and IL-17 upon restimulation, although the cells proliferated and expressed activation markers. Furthermore, the generation of regulatory CD4 + CD25 high Foxp3 + T cells was induced in the presence of PP14, in both antigen-specific as well as polyclonal stimulation. In accordance with previous reports, we found that the induction of FoxP3 expression by PP14 is accompanied by down regulation of the PI3K-mTOR signaling pathway. Conclusions/Significance: These data suggest that PP14 arrests T cells in a unique activated state that is not accompanied with the acquisition of effector function, together with promoting the generation of regulatory T cells. Taken together, our results may elucidate the role of PP14 in supporting immune tolerance in pregnancy by reducing T cell effector function

Key role for ubiquitin protein modification in TGFβ signal transduction

The transforming growth factor β (TGFβ) superfamily of signal transduction molecules plays crucial roles in the regulation of cell behavior. TGFβ regulates gene transcription through Smad proteins and signals via non-Smad pathways. The TGFβ pathway is strictly regulated, and perturbations lead to tumorigenesis. Several pathway components are known to be targeted for proteasomal degradation via ubiquitination by E3 ligases. Smurfs are well known negative regulators of TGFβ, which function as E3 ligases recruited by adaptors such as I-Smads. TGFβ signaling can also be enhanced by E3 ligases, such as Arkadia, that target repressors for degradation. It is becoming clear that E3 ligases often target multiple pathways, thereby acting as mediators of signaling cross-talk. Regulation via ubiquitination involves a complex network of E3 ligases, adaptor proteins, and deubiquitinating enzymes (DUBs), the last-mentioned acting by removing ubiquitin from its targets. Interestingly, also non-degradative ubiquitin modifications are known to play important roles in TGFβ signaling. Ubiquitin modifications thus play a key role in TGFβ signal transduction, and in this review we provide an overview of known players, focusing on recent advances

T cells at the site of autoimmune inflammation show increased potential for trogocytosis

CD4+ T cells acquire membrane fragments from antigen-presenting-cells via a process termed trogocytosis. Identifying which CD4+ T cells undergo trogocytosis in co-culture with Ag-loaded APC can enrich for antigen-reactive T cells without knowledge of their fine specificity or cytokine-production profiles. We sought to assess the suitability of this method to identify disease relevant effector and regulatory T cells during autoimmune inflammation. Trogocytosis efficiently identified MBP-reactive T cells in vitro and ex-vivo following immunization. However, Foxp3+ regulatory T cells constitutively displayed a higher rate of trogocytosis than their Foxp3- counterparts which limits the potential of trogocytosis to identify antigen-reactive Treg cells. During inflammation a locally elevated rate of trogocytosis (seen in both effector and regulatory T cells isolated from the inflamed CNS) precludes the use of trogocytosis as a measure of antigenic reactivity among cells taken from inflammatory sites. Our results indicate trogocytosis detection can enrich for Ag-reactive conventional T cells in the periphery but is limited in its ability to identify Ag-reactive Treg or T effector cells at sites of inflammation. Increased trogocytosis potential at inflammatory sites also draws into the question the biological significance of this phenomenon during inflammation, in Treg mediated suppression and for the maintenance of tolerance in health and disease