Assessment of chemokine receptor expression by human Th1 and Th2 cells in vitro and in vivo.

The preferential association of some chemokine receptors with human Th1 or Th2 cells has recently been reported. In this study, the expression of CCR3, CCR5, CXCR3, and CXCR4 were analyzed by flow cytometry in three distinct in vitro models of Th1/Th2 polarization, activated naive and memory T cells, and T-cell clones, in which the intracellular synthesis of interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) and the surface expression of CD30 and LAG-3 were also assessed. Moreover, by using immunohistochemistry the in vivo expression of CCR3, CCR5, CXCR3, and CXCR4 was examined in the gut of patients suffering from Crohn's disease, a Th1-dominated disorder, and in the skin of patients suffering from systemic sclerosis, a Th2-dominated disorder. CCR5 and LAG-3 exhibited the same pathway of Th1 association, whereas CXCR3 did not discriminate between Th1- and Th2-dominated responses. On the other hand, CCR3 was found only occasionally in a small proportion of allergen-specific memory T cells with Th2/ThO profile of cytokine production in vitro. However, it was neither seen in Th2-polarized activated naive T cells nor in established Th2 clones and could be detected in vivo only on non-T cells. Finally, whereas CXCR4 expression was not limited to Th2 cells in vivo, it was markedly up-regulated by IL-4 and down-regulated by IFN-gamma in vitro. Thus, the results of this study confirm the existence of flexible programs of chemokine receptor expression during the development of Th1 and Th2 cells. However, caution is advised in interpreting these receptors as surrogate markers of a given type of effector response.     

Switch in Chemokine Receptor Phenotype on Memory T Cells without a Change in the Cytokine Phenotype

Th1 and Th2 cells as defined by their cytokine profile are associated with the expression of the chemokine receptors CCR5 and CCR3, respectively. In committed human memory Th1 cells the cytokine profile is irreversibly expressed. However, it is not known if the chemokine receptor phenotypes of Th1 and Th2 cells are permanently associated to the cytokine profile or if it can be changed. To analyze the possibility of inducing a switch in chemokine receptor phenotype on memory Th cells we used differentiated memory Th cells isolated from synovial tissue (ST) samples of patients with rheumatoid arthritis (RA). Freshly isolated T cells, T-cell lines and T-cell clones from these tissues were manipulated with Th1 (interleukin (IL)-12 + anti IL-4) or Th2 (IL-4 + anti IL-12) inducing conditions. The surface expression of CCR5 and CCR3 was analyzed by flowcytometry and interferon (IFN)-gamma and IL-4 production by ELISA. A Th1-inducing cytokine environment increased the expression of CCR5 in Th1 cells and induced the expression of CCR5 in Th2 cells as compared to culture condition with only IL-2. Induction of CCR5 expression on Th2 clones was associated with secretion of some IFN-gamma. Moreover, the Th2-associated chemokine receptor CCR3 could be expressed on both Th1-dominant cell lines, and clones of Th1 and Th0 type after culture conditions with IL-4. This expression of CCR3 was associated with a reduced IFN-gamma production, but no IL-4 production could be induced. The IL-4-treated Th1 clones had a reduced migratory capacity against chemokines produced by ST cells compared to nonmanipulated T-cell clones. In contrast, the same IL-12-treated Th1 clones showed an increased migratory potential. Induction of the Th2-associated marker CCR3 on memory Th1 cells demonstrates that a change in chemokine receptor phenotype related to the Th2 type can be induced on terminally differentiated Th1 cells, without a change in the cytokine profile.     

Systemic chemokine and chemokine receptor responses are divergent in allergic versus non-allergic humans

T(h)1- and T(h)2-polarized human T cell clones display distinct patterns of chemokine receptor expression and selective chemokine responsiveness in vitro. We hypothesized that natural exposure to environmental grass pollen would induce differential systemic chemokine and chemokine receptor expression patterns in individuals with allergic rhinitis compared to healthy controls with type 2- and type 1-dominated responses to allergen respectively. To this end, we compared chemokine receptor expression on peripheral blood T cells directly ex vivo and plasma chemokine levels between these two groups of study participants prior to and during the grass pollen season. T(h)1-associated CXC chemokine receptor (CXCR) 3 was strongly expressed on >50% CD4(+)/CD45RO(+) cells of all subjects. When examined longitudinally, CXCR3 expression increased over the grass pollen season (P < 0.0001), solely in non-allergic subjects. In contrast, for both allergic and non-allergic subjects, CC chemokine receptor (CCR) 5 (T(h)1-associated) and CCR3 (T(h)2-associated) were weakly expressed on <10% of CD4(+)/CD45RO(+) cells both prior to and during the grass pollen season. Type 1 chemokines CXC chemokine ligand (CXCL) 9 and CXCL10 (monokine induced by IFN-gamma and IFN-gamma-inducible protein of 10 kDa: CXCR3 ligands), and type 2 chemokines CC chemokine ligand (CCL) 11 (eotaxin: CCR3 ligand), CCL17 (thymus and activation-regulated chemokine: CCR4 ligand) and CCL22 (monocyte-derived chemokine: CCR4 ligand) were readily detectable in the plasma of most participants. Systemic CXCL9 levels decreased from pre- to grass pollen season in allergics (P < 0.05), whereas CCL17 decreased in non-allergics (P < 0.05) over the same period. Taken together, these longitudinal data suggest a systemic shift to more intensely type 1-dominated responses in non-allergic individuals and, conversely, to more type 2-dominated responses in allergic individuals upon natural re-exposure to grass pollen.     

Characterization of Th1- and Th2-associated Chemokine Receptor Expression in Spleens of Patients with Immune Thrombocytopenia

Purpose

In view of the numerous clinical observations and laboratory studies that suggest a critical role for the spleen in immune thrombocytopenia (ITP) pathophysiology, we aimed to characterize Th1-associated chemokine receptors CXCR3 and CCR5 and Th2-associated chemokine receptor CCR3 in spleens of ITP patients and assess the significance of their differential expression in the clinical setting.

Methods

The histopathology of spleens was observed using hematoxylin-eosin staining (HE), and the positive rate of CXCR3, CCR5 and CCR3 expression in spleens of 24 ITP patients and 12 patients with traumatic splenic rupture as normal controls was detected by immunohistochemistry using the SP method. CXCR3, CCR5 and CCR3 protein expression was analyzed by Western blot and mRNA levels were investigated by real-time polymerase chain reaction (RT-PCR).

Results

Reactive hyperplasia could be seen in follicles of the white pulp, the germinal central zone was enlarged and the marginal zone was thickened, the central arteries were thickened and fibrotic, and the density of the capillary vessel was increased in ITP patients. ITP group displayed a higher rate of expression of Th1-associated chemokine receptors CXCR3 and CCR5 (83.3 % vs. 75 %, 100 % vs. 83.3 %) but lower rate of expression of Th2-associated chemokine receptor CCR3 (50 % vs. 66.7 %) compared with the controls (P?<?0.05, respectively). Western blot analysis revealed that CXCR3 and CCR5 protein expression was significantly increased in ITP patients while CCR3 was significantly reduced (P?<?0.05, respectively). Meanwhile, ITP patients displayed increased mRNA levels of CXCR3 and CCR5 but decreased gene expression of CCR3 (P?<?0.05, respectively).

Conclusion

The data suggested that the abnormal expression of Th1/Th2 chemokine receptors may participate in splenic immune disorder in patients with ITP. Using corresponding inhibitors may inhibit Th1-dominant expression and mitigate the progress of the disease.     

Impaired CD4 and CD8 T cell phenotype and reduced chemokine secretion in recent-onset type 1 diabetic children

Although the role of the T cell-mediated autoimmune reaction in type 1 diabetes (T1D) is conclusive, studies including data from human circulating CD4(+) and CD8(+) lymphocytes subsets during the disease onset and posterior development are scarce. Further, chemokines and chemokine receptors are key players in the migration of pathogenic T cells into the islets of non-obese diabetic mice developing T1D, but few studies have investigated these markers in human T1D patients. We studied the expression of T helper 1 (Th1)- and Th2-associated chemokine receptors, and the two isoforms of CD45 leucocyte antigen on CD4(+) and CD8(+) lymphocytes from T1D and healthy children, as well as the secretion of chemokines in cell supernatants in peripheral blood mononuclear cells. Our results showed increased expression of CCR7 and CD45RA and reduced CD45RO on CD8(+) cells among recent-onset T1D patients. The percentages of CD4(+) cells expressing CXC chemokine receptor 3 (CXCR3), CXCR6 and CCR5, and the secretion of interferon-gamma-induced protein-10, monocyte chemoattractant protein-1, macrophage inflammatory protein (MIP)-1alpha and MIP-1beta was lower among diabetics. Low expression of Th1-associated receptors and secretion of chemokines, together with an increased amount of CD8(+) cells expressing CD45RA and CCR7 in T1D patients therefore might represent suboptimal Th function in T1D, leading to impaired T cytotoxic responses or alternatively reflect a selective recruitment of Th1 cells into the pancreas.     

Downregulation of CXCR6 and CXCR3 in lymphocytes from birch-allergic patients

Preferential expression of chemokine receptors on Th1 or Th2 T-helper cells has mostly been studied in cell lines generated in vitro or in animal models; however, results are less well characterized in humans. We determined T-cell responses through chemokine receptor expression on lymphocytes, and cytokine secretion in plasma from birch-allergic and healthy subjects. The expression of CCR2, CCR3, CCR4, CCR5, CCR7, CXCR3, CXCR4, CXCR6, IL-12 and IL-18R receptors was studied on CD4⁺ and CD8⁺ cells from birch-allergic ( n  = 14) and healthy ( n  = 14) subjects by flow cytometry. The concentration of IL-4, IL-5, IL-10, IL-12, IFN-γ and TNF-α cytokines was measured in plasma from the same individuals using a cytometric bead array human cytokines kit. The similar expression of CCR4 in T cells from atopic and healthy individuals argues against the use of the receptor as an in vivo marker of Th2 immune responses. Reduced percentages of CD4⁺ cells expressing IL-18R, CXCR6 and CXCR3 were found in the same group of samples. TNF-α, IFN-γ, IL-10, IL-5, IL-4 and IL-12 cytokines were elevated in samples from allergic individuals. Reduced expression of Th1-associated chemokine receptors together with higher levels of Th1, Th2 and anti-inflammatory cytokines in samples from allergic patients indicate that immune responses in peripheral blood in atopic diseases are complex and cannot be simplified to the Th1/Th2 paradigm. Not only the clinical picture of atopic diseases but also the clinical state at different time points of the disease might influence the results of studies including immunological markers associated with Th1- or Th2-type immune responses.     

Dynamic T-lymphocyte chemokine receptor expression induced by interferon-beta therapy in multiple sclerosis

Treatment with interferon (IFN)-beta reduces clinical disease activity in multiple sclerosis (MS). Using flow cytometry, an enzyme-linked immunosorbent assay and a real-time polymerase chain reaction, we studied in vivo IFN-beta-induced effects on CD4(+) T-lymphocyte chemokine receptor expression as these influence central nervous system (CNS) transmigration and inflammation. At 'steady state' (>/=1 day after the most recent IFN-beta injection), IFN-beta treatment increased CD4(+) T-cell surface expression of CC chemokine receptor (CCR)4, CCR5 and CCR7 after 3 months of treatment, whereas that of CXC chemokine receptor (CXCR)3 was unaltered. Conversely, at 9-12 h after the most recent IFN-beta injection, CCR4, CCR5 and CCR7 expressions were unaltered, while CXCR3 expression was reduced. CD4(+) T-cell surface expression of CCR4 was significantly lower in untreated MS patients compared with healthy volunteers. Of the plasma chemokines, only CXCL10 was increased by IFN-beta treatment; CCL3, CCL4, CCL5 and CXCL9 were unaltered. CCR5 mRNA expression in blood mononuclear cells correlated with the expression of T-helper type 1 (Th1)-associated genes whereas CCR4 and CCR7 mRNA expression correlated with Th2 and immunoregulatory genes. In conclusion, IFN-beta treatment caused 'steady-state' increases of several chemokine receptors relevant for CD4(+) T-lymphocyte trafficking and function, possibly facilitating lymphocyte migration into the CNS. An important therapeutic effect of IFN-beta treatment may be the normalization of a decreased Th2-related CD4(+) T-cell CCR4 expression in MS patients. Surface chemokine receptor expression and CXCL10 varied according to the timing of blood sampling in relation to the most recent IFN-beta injection. Thus, it is imperative to distinguish acute effects of IFN-beta from steady-state effects.     

Chemoattractant receptors expressed on type 2 T cells and their role in disease

The existence of two functionally distinguished populations among T cells has been established in both mice and humans. Type 1 T helper (Th1) cells are involved in the defense against intracellular bacteria and many viruses, while type 2 Th cells (Th2) are the major actors in the response against parasites and play a central role in allergic inflammation. More recently, several data have suggested that some chemokine receptors are tightly regulated on T cells, and in accordance with this selective expression, Th1 and Th2 cells can be differentially recruited by specific chemokines to the inflammatory sites. Among Th2-associated chemokine receptors, CCR3, CCR4 and CCR8 have been described to play a central role in allergic inflammation. However, CCR3 is mainly expressed on basophils, eosinophils and mast cells, but it is poorly expressed by Th2 cells, and CCR4 is also expressed by Th subsets different from Th2 cells. So far, the chemoattractant receptors which among T cells appear to be selectively expressed by Th2 cells or their subsets are CCR8 and CRTH2. The ligand for CRTH2 is not a chemokine, but is prostaglandin (PG)D2, which is able to attract basophils, eosinophils, Th2 cells and type 2 cytotoxic (Tc2) CD8+ T lymphocytes. The selective expression of CRTH2 on Th2 and Tc2 cells may be useful to develop new therapeutic strategies against allergic diseases and against other immune disorders. Additional studies, however, are required to understand its effective importance in the induction and maintenance of Th2- or Tc2-mediated response and inflammation.     

Involvement of CCR5 in the passage of Th1-type cells across the blood-retina barrier in experimental autoimmune uveitis

Although the recruitment of T helper cell type 1 (Th1)/Th2 cells into peripheral tissues is essential for inflammation and the host response to infection, the traffic signals that enable the distinct positioning of Th1/Th2 cells are unclear. We have determined the role of CC chemokine receptor 5 (CCR5) in this using experimental autoimmune uveitis (EAU) as a model system. In EAU, Th1-like cells are preferentially recruited into the retina across the blood-retina barrier, partly as a result of expression of the adhesion molecules P-selectin glycoprotein ligand 1 and lymphocyte function-associated antigen-1 on these cells. CD3+ T cells, infiltrating the retina, also expressed the chemokine receptor CCR5, and CCR5 ligands, macrophage-inflammatory protein-1alpha (MIP-1alpha), MIP-1beta, and regulated on activation, normal T expressed and secreted (RANTES), were strongly expressed in the retina at peak EAU. Th1-like cells, polarized in vitro, expressed high levels of CCR5. The trafficking of these CCR5+ cells was examined by tracking them after adoptive transfer in real time in vivo at an early disease stage using scanning laser ophthalmoscopy. Treatment of the cells with antibody against CCR5 prior to transfer resulted in a reduction in their infiltration into the retina. However, rolling velocity, rolling efficiency, and adherence of the cells to retinal endothelium were not reduced. CCR5 is clearly important for Th1 cell recruitment, and this study demonstrates for the first time in vivo that CCR5 may act at the level of transendothelial migration rather than at the earlier stage of rolling on the endothelium.     

The chemokines CCL5, CCL2 and CXCL12 play significant roles in the migration of Th1 cells into rheumatoid synovial tissue

As the T-cell population in the synovial tissue (ST) in rheumatoid arthritis (RA) is dominated by T helper (Th) 1 cells, this study was designed to examine whether there is a preferential migration of polarized T cells to ST, and to identify the chemokines responsible for the migration. This was done by developing 10 T-cell clones specific for an arbitrary antigen (mouse immunoglobulin G (IgG)) from the peripheral blood (PB) of a healthy donor sensitized to mouse IgG. The Th polarizations of the clones were determined by measuring secreted interferon-gamma and interleukin-4, following anti-CD3 stimulation. Migration to pools of RA ST cell-derived supernatants was analysed. Expression of the chemokine receptors CCR1, CCR2, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CXCR3 and CXCR4 were analysed by flow cytometry. Th1 clones showed significantly higher migration to RA ST cell-derived supernatant compared with Th2 clones. Blocking of either of the chemokines, CCL5 or CCL2, strongly inhibited migration of the Th1 cells between 56 and 77%, while blocking of CXCL12 inhibited migration between 44 and 61%. Blocking of CXCL10 had only a minor inhibitory effect. Our results demonstrate a selective migration of Th1 cells to RA ST supernatant and that blocking either CCL5, CCL2 or CXCL12 significantly inhibits T-cell migration. This indicates that CCL5, CCL2 and CXCL12 play significant roles in attracting Th1 cells towards the RA ST, and may prove potent targets for obstructing T-cell migration to the synovium.     

Experimental autoimmune encephalomyelitis: CC chemokine receptor expression by trafficking cells

Chemokines are chemotactic cytokines, which stimulate migration of inflammatory cells towards tissue sites of inflammation. The largest chemokine group, termed CC chemokines (CCLs), act primarily on T cells and monocytes, through CC chemokine receptors (CCRs) belonging to the superfamily of G-protein coupled seven transmembrane domain receptors. CCR expression is a critical determinant of cellular responses to CCLs. In this report, we describe the expression pattern of mRNA encoding selected CCRs in the spinal cord and spleen of perfused and non-perfused mice at different stages of chronic-relapsing EAE (ChREAE). We detected increased expression of receptors (CCR1, CCR5) associated with T helper-1 (Th1) but not those (CCR3, CCR4) associated with Th2 T cells in spinal cord during initial attack and relapse of ChREAE. Expression of these CCRs correlated temporally and spatially with reported previously expression of corresponding CCLs. The principal cells expressing CCR5 were inflammatory cells invading the spinal cord. Our results supported the implication of Th1-associated CCRs in the CNS-specific inflammatory reaction of ChREAE.     

Evidence for PI-3K-dependent migration of Th17-polarized cells in response to CCR2 and CCR6 agonists

IL-17-producing Th cells (Th17) are a distinct subset of effector cells that bridge the innate and adaptive immune system and are implicated in autoimmune disease processes. CD4(+) splenocytes from DO11.10 mice were activated with OVA peptide(323-339) and maintained under Th17 polarization conditions, resulting in significantly higher proportions of IL-17(+) T cells compared with nonpolarized (Th0) cells. Th17-polarizing conditions significantly increased the proportion of cells expressing the chemokine receptors CCR2, CCR6, and CCR9 when compared with Th0 cells. In contrast, there was a significant decrease in the proportion of cells expressing CXCR3 under Th17-polarizing conditions compared with nonpolarizing conditions. The respective chemokine agonists for CCR2 (CCL2 and CCL12), CCR6 (CCL20), and CCR9 (CCL25) elicited migration and PI-3K-dependent signaling events in Th17-polarized cells, thus indicating that all three receptors were functionally and biochemically responsive. Furthermore, postmigration phenotypic analysis demonstrated that the agonists for CCR2 and CCR6, but not CCR9, stimulated a modest enrichment of IL-17(+) cells compared with the premigration population. Pan-isoform inhibitors of PI-3K/Akt signaling prevented CCR2- and CCR6-mediated, polarized Th17 cell migration in a concentration-dependent manner. The unique chemokine receptor expression pattern of Th17 cells and their corresponding PI-3K-dependent migratory responses are important for understanding the pathogenesis of autoimmune diseases and may provide opportunities for the application of CCR2 and CCR6 antagonists and PI-3K isoform-selective inhibitors in defined inflammatory settings.     

Airway infiltration of CD4+ CCR6+ Th17 type cells associated with chronic cigarette smoke induced airspace enlargement

Recently, patients with tobacco smoke induced emphysema have been shown to exhibit classical signs of T cell mediated autoimmunity characterized by autoantibody production and Th1 type responses. As the recently described Th17 type subset has been found to play a role in the pathogenesis of a number of autoimmune diseases previously considered to be Th1 driven, we sought to examine whether a Th17 type response was associated with airspace enlargement in a murine model of emphysema. Six to eight months exposure of mice to inhalation of mainstream cigarette smoke led to progressive airspace enlargement as defined by morphometric analysis. Flow cytometric analysis of the bronchoalveolar lavage (BAL) from these mice demonstrated a significant increase in the overall number of both CD4+ and CD8+ T cells present. These cells were subsequently examined for skewing towards a Th1, Th2 or Th17 phenotype by intracellular cytokine analysis. Distinct populations of BAL CD4+ T cells were found to express IFN-gamma or IL-17 demonstrating the presence of both a Th1 and Th17 type response. No expression of the Th2 associated cytokine IL-4 was detected. Further analysis of this Th17 subset demonstrated that the majority of cells with this effector phenotype express the chemokine receptor CCR6. Together these data identify a novel T cell subset associated with pulmonary inflammation as a result of cigarette smoke exposure. Given the reported roles of CCR6 and IL-17 in promoting pulmonary inflammation, this subset may play an important role in the pathogenesis of cigarette smoke induced autoimmunity.     

Modulation of chemokine receptor expression and chemotactic responsiveness during differentiation of human naive T cells into Th1 or Th2 cells

Chemokines and their receptors direct movements and encounters of lymphocytes and professional APC into specific microenvironments of lymphoid tissues. Chemokine receptors such as CCR7, CXCR5 and CCR4 that are differentially expressed and modulated in distinct subsets of T cells contribute to establish functionally and spatially segregated microenvironments within secondary lymphoid tissues where T cell activation and differentiation occur. Here, we have explored the modulation of CCR7, CCR4, CCR8 and CXCR5 expression and chemotactic responsiveness to their ligands during commitment of human naive T cells along the Th1 or Th2 differentiation pathway in vitro. Our results document that activation of human naive T cells and differentiation in Th1 or Th2 cells result in progressive down-modulation of CCR7 expression and CCL19 responsiveness. By contrast, expression of CCR4 and responsiveness to CCL22 is rapidly induced at the early stages of both Th1/Th2 cell development. However, while CCR4 expression is further up-regulated upon differentiation into Th2 cells, it is lost on fully differentiated Th1 cells. CCR8 is detected at later time points than CCR4 and exclusively on differentiated Th2 cells as revealed by analysis of mRNA expression and responsiveness to CCL1. Expression of CXCR5 is transiently induced at the early stages of Th cell differentiation, but with distinct kinetics in developing Th1 and Th2 cells. Analysis of human tonsillar CD4(+) T cells reveals a consistent pattern of chemotactic responsiveness and chemokine receptor expression in distinct transitional stages of human T cell activation and differentiation in vivo.     

Tryptase-Chymase Double-Positive Human Mast Cells Express the Eotaxin Receptor CCR3 and Are Attracted by CCR3-Binding Chemokines

Eosinophils, basophils, and Th2 cells express the chemokine receptor CCR3, which binds eotaxin, RANTES, and some other chemokines. Using immunohistochemistry and flow cytometry, we demonstrate that CCR3 is also expressed by a variable proportion of human mast cells in gut, skin, and lung tissue. By contrast, with the same anti-CCR3 antibody (B711), CCR3 was poorly if at all detectable on human Th2 cells in vitro and in vivo. Eotaxin neither induced histamine release from purified human mast cells nor increased anti-IgE-stimulated histamine secretion. However, both eotaxin and RANTES elicited mast cell migration in vitro with a similar efficacy. High percentages of CCR3-expressing mast cells were present in the skin and in the intestinal submucosa; much lower percentages were found in the intestinal mucosa and in lung interstitium. Double immunostaining with anti-CCR3 and anti-chymase antibody showed that the vast majority of CCR3-expressing mast cells in the various tissues examined were tryptase-chymase double-positive. Therefore, tryptase-chymase double-positive mast cells express CCR3 and are attracted by CCR3-binding chemokines, eotaxin, and RANTES. Our findings indicate that these chemokines may play an important role in the differentiation and/or migration of this mast cell subset in connective tissues, as well as in sites of allergic inflammation.     

Clonal Th2 cells associated with chronic hypereosinophilia: TARC-induced CCR4 down-regulation in vivo

We analyzed the expression of chemokine receptors on clonal Th2-type CD4(+)CD3(- )lymphocytes isolated from blood of two patients with chronic hypereosinophilia. First, we observed that these Th2 cells express membrane CCR5 and CXCR4 but neither CCR3 nor CCR4 when analyzed immediately after purification. However, CCR4 appeared following culture in human serum-free medium, suggesting that it was down-regulated in vivo. Indeed, patient's serum, but not control human serum, strongly down-regulated CCR4 expression on cultured Th2 cells. As high levels of TARC, a CCR4 ligand, were detected in the serum of four hypereosinophilic patients with CD3(-)CD4(+) clonal Th2 cells, we evaluated the effect of TARC neutralization in this system. Addition of a neutralizing anti-TARC mAb inhibited CCR4 down-regulation by patient's serum, indicating that circulating TARC contributed to CCR4 down-regulation on Th2 cells in vivo. Clonal Th2 cells did not secrete high levels of TARC themselves but induced a sustained production of TARC by monocyte-derived dendritic cells, a phenomenon that was inhibited by addition of blocking mAb against IL-4 receptor. We conclude that high circulating levels of TARC in serum of patients with chronic hypereosinophilia, most likely derived from antigen-presenting cells stimulated by Th2-type cytokines, induce down-regulation of CCR4 on Th2 cells in vivo.     

Th1 and Th2 in human IVF pregnancy with allogenic fetus

PROBLEM: Positive obstetric outcome of allogenic in vitro fertilization (IVF) pregnancies makes interesting the subject of additional regulatory mechanisms after oocyte donation. METHOD OF STUDY: Eighty eight women: 23 donation of oocytes (DO), 33 IVF, 32 natural conception (NC) were studied in first trimester of pregnancy. Intracellular production of cytokines and chemokine receptors expression were studied by flow cytometry. RESULTS: Intracellular production of interferon-gamma (IFN-gamma), interleukin (IL)-4, tumor necrosis factor-alpha by CD4 T lymphocytes in DO women was higher than in IVF and NC women. Ratio IFN-gamma/IL-4 in DO was lower than in IVF. We found higher expression of chemokine receptor CCR4 but not CXCR3 on CD4 T cells in DO compared with IVF and NC. Ratio CXCR3/CCR4 in DO was lower than in NC. CONCLUSION: Hyperactivation of T helper 1 (Th1) and T helper 2 (Th2) by allogenic fetus is specific for DO pregnancy. Preferable activation of Th2 and relative suppression of Th1 chemokine expression reflect additional regulatory counteractive mechanism(s).     

Distinctive features of classic and nonclassic (Th17 derived) human Th1 cells

T helper17 (Th17) lymphocytes represent a third arm of the CD4⁺ T‐cell effector responses, in addition to Th1 and Th2 cells. Th17 cells have been found to exhibit high plasticity because they rapidly shift into the Th1 phenotype in inflammatory sites. In humans, Th1 cells derived from Th17 cells express CD161, whereas classic Th1 cells do not; these Th17‐derived Th1 cells have been termed nonclassic Th1 cells. In this study, we examined similarities and differences between classic and nonclassic human Th1 cells by assessing a panel of T‐cell clones, as well as CD161⁺ or CD161^? CD4⁺ T cells derived ex vivo from the circulation of healthy subjects or the synovial fluid of patients with juvenile idiopathic arthritis. The results show that nonclassic Th1 cells can be identified based on CD161 expression, as well as the consistent expression of retinoic acid orphan receptor C, IL‐17 receptor E, CCR6, and IL‐4‐induced gene 1, which are all virtually absent in classic Th1 cells. The possibility to distinguish these two‐cell subsets by using such a panel of markers may allow the opportunity to better establish the respective pathogenic roles of classic and nonclassic (Th17 derived) Th1 cells in different chronic inflammatory disorders.