Characterization of CD4+ CD25+ T cells in solid organ transplant recipients

Although important progresses have been achieved in the therapeutic management of transplant recipients, acute and chronic rejections remain the leading causes of premature graft loss after solid organ transplantation. This, together with the undesirable side effects of immunosuppressive drugs, has significant implications for the long-term outcome of transplant recipients. Thus, a better understanding of the immunological events occurring after transplantation is essential. The immune system plays an ambivalent role in the outcome of a graft. On one hand, some T lymphocytes with effector functions (called alloreactive) can mediate a cascade of events eventually resulting in the rejection, either acute or chronic, of the grafted organ ; on the other hand, a small subset of T lymphocytes, called regulatory T cells, has been shown to be implicated in the control of these harmful rejection responses, among other things. Thus, we focused our interest on the study of the balance between circulating effectors (alloreactive) and regulatory T lymphocytes, which seems to play an important role in the outcome of allografts, in the context of kidney transplantation. The results were correlated with various variables such as the clinical status of the patients, the immunosuppressive drugs used as induction or maintenance agents, and past or current episodes of rejection. We observed that the percentage of the alloreactive T lymphocyte population was correlated with the clinical status of the kidney transplant recipients. Indeed, the highest percentage was found in patients suffering from chronic humoral rejection, whilst patients on no or only minimal immunosuppressive treatment or on sirolimus-based immunosuppression displayed a percentage comparable to healthy non-transplanted individuals. During the first year after renal transplantation, the balance between effectors and regulatory T lymphocytes was tipped towards the detrimental effector immune response, with the two induction agents studied (thymoglobulin and basiliximab). Overall, these results indicate that monitoring these immunological parameters may be very useful for the clinical follow-up of transplant recipients ; these tests may contribute to identify patients who are more likely to develop rejection or, on the contrary, who tolerate well their graft, in order to adapt the immunosuppressive treatment on an individual basis.

Homeostasis and in vivo effector function of antigen-specific CD4+CD25+ regulatory T cells in experimental transplantation.

CD4+CD25+ regulatory T cells (Tregs) play a critical role in the prevention of autoimmune diseases as well as in the induction and maintenance of dominant tolerance in transplantation models. While their suppressive function has been extensively studied in vitro, their homeostasis and mechanisms of immunoregulation still remain to be clarifi ed in vivo. Using a murine adoptive transfer and skin allograft model, we analysed the expansion, effector function and traffi cking of effector T cells in the presence or absence of donor-specifi c Tregs. Although hyporesponsive to allogeneic and polyclonal stimulation in vitro, transferred Tregs survived and expanded, in response to an allograft in vivo. When co-transferred with naive CD4+CD25- effector T cells, they specifi cally prevented donor but not 3rd party allograft rejection by inhibiting the production of effector cytokines rather than the proliferation of effector T cells in response to alloantigens. The co-transfer of donor-specifi c Tregs did not affect the homing of effector T cells towards the graft draining lymph nodes, but it markedly reduced the infi ltration of the allograft by these pathogenic cells. Furthermore, in recipients where donor-specifi c transplantation tolerance was induced, Tregs preferentially accumulated in the allograft draining lymph nodes and within the grafted skin itself. Taken together, our results suggest that the suppression of graft rejection is an active process that involves the persistent presence of Tregs at the site of antigenic challenge.

Long-term persistence of HIV-1 vaccine-induced CD4+CD45RA-CD62L-CCR7- memory T-helper cells.

OBJECTIVE: To determine in chimpanzees if candidate HIV-1 subunit protein vaccines were capable of eliciting long-lasting T-cell memory responses in the absence of viral infection, and to determine the specific characteristics of these responses. DESIGN: A longitudinal study of cell-mediated immune responses induced in three chimpanzees following immunization with subunit envelope glycoproteins of either HIV-1 or herpes simplex virus (HSV)-2. Following these pre-clinical observations, four human volunteers who had been immunized 7 years previously with the same HIV-1 vaccine candidate donated blood for assessment of immune responses. METHODS: Responses were monitored by protein and peptide based ELISpot assays, lymphocyte proliferation, and intracellular cytokine staining. Humoral responses were assessed by enzyme-linked immunosorbent assay and virus neutralization assays. RESULTS: Although antigen (Ag)-specific CD4 T-cell responses persisted for at least 5 years in chimpanzees, CD8 T-cell responses were discordant and declined within 2 years. Detailed cellular analyses revealed that strong Th1 in addition to Th2 type responses were induced by AS2/gp120 and persisted, whereas CD8 T-cell memory declined in peripheral blood. The specificity of both Th and cytotoxic T-lymphocyte responses revealed that the majority of responses were directed to conserved epitopes. The remarkable persistence of Ag-specific CD4 T-cell memory was characterized as a population of the CD45RA-CD62L-CCR7- "effector phenotype" producing the cytokines IFNgamma, IL-2 and IL-4 upon epitope-specific recognition. Importantly, results in chimpanzees were confirmed in peripheral blood of one of four human volunteers studied more than 7 years after immunization. CONCLUSION: These studies demonstrate that epitope-specific Th1 and Th2 cytokine-dependent Th responses can be induced and maintained for longer than 5 years by immunization with subunit proteins of HIV-1.

Influence of CD4+/CD25+ regulatory T cells on atherogenesis in patients with end-stage kidney disease.

Atherosclerosis, which is influenced by both traditional and nontraditional cardiovascular risk factors and has been characterized as an inflammatory process, is considered to be the main cause of the elevated cardiovascular risk associated with chronic kidney disease. The inflammatory component of atherosclerosis can be separated into an innate immune response involving monocytes and macrophages that respond to the excessive uptake of lipoproteins and an adaptive immune response that involves antigen-specific T cells. Concurrent with the influx of immune cells to the site of atherosclerotic lesion, the role of the adaptive immune response gradually increases. One of those cells are represented by the CD4+/CD25+ Tregs, which play indispensable roles in the maintenance of natural self-tolerance and negative control of pathological, as well as physiological, immune responses. Altered self-antigens such as oxidized LDLs may induce the development of CD4+/CD25+ Tregs with atheroprotective properties. However, atherosclerosis may be promoted by an imbalance between regulatory and pathogenic immunity that may be represented by the low expression of the forkhead box transcription factor (Foxp3) in CD4+/CD25+ Tregs. Such a defect may break immunologic tolerance and alter both specific and bystander immune suppression, leading to exacerbation of plaque development. Patients with end-stage kidney disease (ESKD) display a cellular immune dysfunction and accelerated atherosclerosis. Uremic solutes that accumulate during ESKD may be involved in these processes. In patients with ESKD and especially in those that are chronically hemodialyzed, oxidative stress induced by oxidized LDLs may increase CD4+/CD25+ Treg sensitivity to Fas-mediated apoptosis as a consequence of specific dysregulation of IL-2 expression. This review will focus on the current state of knowledge regarding the influence of CD4+/CD25+ Tregs on atherogenesis in patients with ESKD, and the potential effect of statins on the circulating number and the functional properties of these cells.

CD4+CD25+Foxp3+ regulatory T cells: from basic research to potential therapeutic use.

Regulatory T cells control immune responses to self- and foreign-antigens and play a major role in maintaining the balance between immunity and tolerance. This article reviews recent key developments in the field of CD4+CD25+Foxp3+ regulatory T (TREG) cells. It presents their characteristics and describes their range of activity and mechanisms of action. Some models of diseases triggered by the imbalance between TREG cells and effector pathogenic T cells are described and their potential therapeutic applications in humans are outlined.

Role of CD4+CD25+ regulatory T cells in the antigen specific induction of tolerance "via" the nasal route

ABSTRACT Asthma is a complex inflammatory syndrome caused by environmental factors in predisposed individuals (atopics). Its severity correlates with the presence of activated T lymphocytes and eosinophils in the bronchoalveolar lavage fluid (BALF). Induction of tolerance via the nasal route results in reduced recruitment of eosinophils into BALF upon challenge, inhibition of TH2 pro-inflammatory cytokine secretion and T cell hyporesponsiveness. Recently, CD4+CD25+ natural regulatory T cells (Treg) were proposed as key players in controlling the development of asthma and allergic disease. The objective of the present study is to investigate the role of CD4+CD25+ regulatory T cells in the mechanisms leading to tolerance in an established model of asthma. In this goal we depleted CD4+CD25+ T cells at different times during asthma and tolerance induction protocol in mice and looked at efficiency of tolerization (intranasal application of high dose of allergen) in the absence of natural Tregs. First, ovalbumin-sensitized mice were depleted of CD25+ T cells by intraperitoneal injection of anti-CD25 mAb (PC61) either for along-term (repeated injections of anti-CD25 from day 31 until the end of the protocol) or a short-term period (single injection of anti-CD25 before or after tolerance induction). We demonstrated that the long-term depletion of CD4+CD25+ T cells severely hampered tolerance induction (marked enhancement in eosinophil recruitment into BALF and a vigorous antigen specific T cell response to OVA upon allergen challenge) whereas transient depletions were not sufficient to do so. We then characterized T cell subsets by flow cytometry and observed that a large part of CD4+CD25+ T cells express Foxp3, an established marker of regulatory T cells. We also tested in-vitro suppressor activity of CD4+CD25+ T cells from tolerized mice by cell proliferation assay in coculture and observed a strong suppressive activity. Our data suggest that CD4+CD25+ T cells with regulatory properties play a crucial role in the induction of tolerance via the nasal route. The relationship between CD25+ natural Treg and inducible IL-10+ TRl-type Treg will have to be defined. RESUME L'asthme est un syndrome inflammatoire complexe provoqué par des facteurs environnementaux chez des individus génétiquement prédisposés (atopiques). Sa sévérité corrèle avec la présence des lymphocytes T activés et d'éosinophiles dans le lavage bronchoalvéolaire (BAL). L'induction de la tolérance par la voie nasale résulte en une diminution du recrutement des eosinophils dans le BAL, une inhibition de la sécrétion de cytokines pro-inflammatoires de type TH2 et de l'hypo-réponse des cellules T à l'allergène. Récemment, les cellules régulatrices «naturelles » de type CD4+CD25+ T (Tregs) ont été proposées comme acteurs essentiels dans le développement de l'asthme et de l'allergie. L'objectif de cette étude est d'étudier le rôle des cellules régulatrices CD4+CD25+ T dans les mécanismes menant à la tolérance dans un modèle établi d'asthme. Dans ce but nous avons déplété les cellules de CD4+CD25+ T à différents temps au cours du protocole d'induction d'asthme et de tolérance et nous avons regardé l'efficacité de l'induction de tolérance (application intranasale d'une dose importante d'allergène) en l'absence de Tregs. Dans un premier temps des souris sensibilisées à l'ovalbumine (OVA) ont été déplétées en cellules CD25+ T par l'injection intrapéritonéale d'anti-CD25 mAb (PC61) pour une longue période (injections répétées d'anti-CD25 du jour 31 jusqu'à la fin du protocole) ou pour une courte période (injection unique d'anti-CD25 avant ou après l'induction de tolérance). Nous avons démontré que la déplétion à long t erme des cellules de CD4+CD25+ T a empêché l'induction de tolérance (recrutement accru d'éosinophiles dans le BAL et une réponse vigoureuse des cellules T spécifiques de l'antigène après exposition à l'allergène) tandis des déplétions à court-terme n'ont pas cet effet. Nous avons ensuite caractérisé des sous-populations de cellules T par cytométrie de flux. Nous avons observé que la majorité des cellules CD4+CD25+ T expriment Foxp3, un marqueur établi des cellules régulatrices. Nous avons également examiné in vitro l'activité régulatrice des cellules T CD4+CD25+ issues de souris tolérisées. La prolifération de cellules T en coculture a démontré une forte activité suppressive des cellules CD4+CD25+. Nos données suggèrent que des cellules T CD4+CD25+ ayant des propriétés régulatrices jouent un rôle crucial dans l'induction de la tolérance par la voie nasale. Le rapport entre les cellules régulatrices naturelles CD4+CD25+ et les cellules régulatrices inductible de type TR1 I1-10+ devra être défini. RESUME DESTINE A UN LARGE PUBLIC L'asthme est une maladie inflammatoire des bronches, caractérisée par des crises de dyspnée (gêne respiratoire) témoignant d'une activation brutale des muscles bronchoconstricteurs, auxquelles s'associent un oedème et une hypersécrétion des muqueuses des voies aériennes ainsi qu'une importante production d'anticorps de l'allergie (IgE). Chez la plupart des enfants atteints et chez près de la moitié des adultes concernés par l'asthme, c'est une allergie à des substances présentes dans l'air environnant (acariens, pollens ou poils d'animaux) qui est à l'origine de la maladie. . Le traitement actuel de l'asthme repose d'une part sur le soulagement des symptômes grâce à des produits à base de stéroïdes ou des bronchodilatateurs. D'autre part, l'immunothérapie spécifique (aussi appelée désensibilisation) permet d'améliorer l'asthme et de «reprogrammer» le système immunitaire. C'est à ce jour, le seul moyen connu de faire régresser une allergie. Cependant l'immunothérapie prend beaucoup de temps (3 à 5 ans) et ne marche pas à tous les coups ni pour tous les antigènes. Il est donc important de mieux comprendre les mécanismes impliqués lors d'un tel traitement afin d'en améliorer l'efficacité. Af n de pouvoir investiguer en détail ces mécanismes des modèles d'immunothérapie ont été mis au point chez la souris. Notre étude se base sur un modèle d'asthme allergique chez la souris. Des souris sont rendues allergiques à l'ovalbumine (OVA) et présentent alors les caractéristiques majeures de l'asthme humain (recrutement de cellules inflammatoires dans les poumons, augmentation de la production d'IgE et de la résistance des bronches aux flux respiratoires). Ces souris asthmatiques une fois traitées par l'application nasale d'OVA (forme d'immunothérapie muqueuse) ne développent plus de réaction allergique lors d'une ré-exposition à l'allergène. Notre hypothèse est que cette «guérison» (tolérance) est liée à l'action de cellules (lymphocytes T CD4) dites «régulatrices» et caractérisées par le marqueur CD25. Pour le démontrer, nous avons éliminé ces cellules «régulatrices» CD25 de nos souris asthmatiques grâce à un anticorps monoclonal spécifique. Nous n'avons dès lors plus été en mesure d'induire une tolérance à l'allergène. Ceci suggère donc un rôle clé des cellules «régulatrices» T CD4+CD25+ dans la réussite de l'immunothérapie nasale dans notre modèle. Nos résultats n'excluent pas la participation d'autres cellules telles que les lymphocytes producteurs d'IL-10 (lymphocytes régulateurs induits). Le rôle respectif de ces sous-populations régulatrices devra être examiné dans les études à venir. Une meilleure maîtrise des mécanismes de régulation pourrait s'avérer cruciale pour améliorer les thérapies de l'asthme.

Patterns of intracellular cytokines in CD4 and CD8 T cells from patients with mycobacterial infections

Using a short-term bulk culture protocol designed for an intracellular-staining method based on a flow cytometry approach to the frequencies of cytokine-producing cells from tuberculosis and leprosy patients, we found distinct patterns of T cell subset expression. The method also reveals the profile of peak cytokine production and can provide simultaneous information about the phenotype of cytokine-producing cells, providing a reliable assay for monitoring the immunity of these patients. The immune response of Mycobacterium leprae and purified protein derivative (PPD) in vitro to a panel of mycobacteria-infected patients from an endemic area was assessed in primary mononuclear cell cultures. The kinetics and source of the cytokine pattern were measured at the single-cell level. IFN-gamma-, TNF-alpha-, IL-4- and IL-10-secreting T cells were intracytoplasmic evaluated in an attempt to identify M. leprae- and PPD-specific cells directly from the peripheral blood. The analysis by this approach indicated that TNF-alpha was the first (8 h) to be produced, followed by IFN-gamma (16 h), IL-10 (20 h) and IL-4 (24 h), and double-staining experiments confirmed that CD4+ were a greater source of TNF-alpha than of CD8+ T cells (P < 0.05). Both T cell subsets secreted similar amounts of IFN-gamma. We conclude that the protocol permits rapid evaluation of cytokine production by different T cell populations. The method can also be used to define immune status in non-infected and contact individuals.

In vivo fluctuation of Tax, Foxp3, CTLA-4, and GITR mRNA expression in CD4+CD25+ T cells of patients with human T-lymphotropic virus type 1-associated myelopathy

HTLV-1 Tax expression exerts an inhibitory effect on the Foxp3 transcription factor in CD4+CD25+ T-regulatory cells (Treg). For a better understanding of the role of Tax mRNA in the gene expression of cellular markers we measured Tax, Foxp3, CTLA-4, GITR, TGF-&#946;, and IL-10 mRNA in Treg cells of 50 patients with human T-lymphotropic virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP; 27 women and 23 men; mean age: 56.7 years). The control group consisted of 23 non-infected subjects (12 women and 11 men) with a mean age of 51.3 years. Real-time PCR was used to measure mRNA of Tax proteins and several cellular markers of Treg function. Determinations revealed a high level of Tax mRNA in HAM/TSP (124.35 copies/100 CD4+CD25+ T cells). Foxp3, GITR, and CTLA-4 mRNA levels were lower in HAM/TSP patients (mean ± SD, 22.07 ± 0.78, 9.63 ± 0.36, and 4.54 ± 0.39, respectively) than in non-infected controls (47.15 ± 12.94, 22.14 ± 1.91, and 21.07 ± 2.31). Both groups had similar levels of TGF-&#946; and IL-10. An inverse relationship was found between Tax levels and Foxp3, CTLA-4, and GITR levels. Conversely, there was a direct correlation between levels of Foxp3, GITR, and CTLA-4. Disease severity and evolution time did not correlate with Tax or Foxp3 levels. The present results suggest that Tax and Foxp3 mRNA vary with the same degree of disease severity in HAM/TSP patients. Tax fluctuations may affect CTLA-4 and GITR expression via the Foxp3 pathway, causing virus-induced dysfunction of CD4+CD25+ T cells in HAM/TSP patients.

Function of the immunoregulatory CD4-CD8- T cells in the context of autoimmune diabetes

La tolérance immunitaire dépend de la distinction entre le soi et le non soi par le système immunitaire. Un bris dans la tolérance immunitaire mène à l'auto-immunité, qui peut provoquer la destruction des organes, des glandes, des articulations ou du système nerveux central. Le diabète auto-immun, également connu sous le nom diabète juvénile et diabète de type 1, résulte d'une attaque auto-immune sur les cellules β pancréatiques sécrétrices d’insuline, localisées au niveau des îlots de Langerhans du pancréas. Bien que le diabète auto-immun soit traitable par une combinaison d’injections quotidiennes d’insuline d’origine exogène, de régime et d'exercices, beaucoup de complications chroniques peuvent se manifester chez les patients, y compris, mais non limitées à, la cécité, les maladies cardiovasculaires, l’insuffisance rénale et l'amputation. En raison des nombreuses complications liées au diabète auto-immun à long terme, la recherche continue afin de mieux comprendre tous les facteurs impliqués dans la progression de la maladie dans le but de développer de nouvelles thérapies qui empêcheront, renverseront et/ou traiteront cette maladie. Un rôle primordial dans la génération et l'entretien de la tolérance immunitaire a été attribué au nombre et à la fonction des sous-populations de cellules régulatrices. Une de ces populations est constituée de cellules T CD4-CD8- (double négatives, DN), qui ont été étudiées chez la souris et l'humain pour leur contribution à la tolérance périphérique, à la prévention des maladies et pour leur potentiel associé à la thérapie cellulaire. En effet, les cellules de T DN sont d'intérêt thérapeutique parce qu'elles montrent un potentiel immunorégulateur antigène-spécifique dans divers cadres expérimentaux, y compris la prévention du diabète auto-immun. D’ailleurs, en utilisant un système transgénique, nous avons démontré que les souris prédisposées au diabète auto-immun présentent peu de cellules T DN, et que ce phénotype contribue à la susceptibilité au diabète auto-immun. En outre, un transfert des cellules T DN est suffisant pour empêcher la progression vers le diabète chez les souris prédisposées au diabète auto-immun. Ces résultats suggèrent que les cellules T DN puissent présenter un intérêt thérapeutique pour les patients diabétiques. Cependant, nous devons d'abord valider ces résultats en utilisant un modèle non-transgénique, qui est plus physiologiquement comparable à l'humain. L'objectif principal de cette thèse est de définir la fonction immunorégulatrice des cellules T DN, ainsi que le potentiel thérapeutique de celles-ci dans la prévention du diabète auto-immun chez un modèle non-transgénique. Dans cette thèse, on démontre que les souris résistantes au diabète auto-immun présentent une proportion et nombre absolu plus élevés de cellules T DN non-transgéniques, lorsque comparées aux souris susceptibles. Cela confirme une association entre le faible nombre de cellules T DN et la susceptibilité à la maladie. On observe que les cellules T DN éliminent les cellules B activées in vitro par une voie dépendante de la voie perforine et granzyme, où la fonction des cellules T DN est équivalente entre les souris résistantes et prédisposées au diabète auto-immun. Ces résultats confirment que l'association au diabète auto-immun est due à une insuffisance en terme du nombre de cellules T DN, plutôt qu’à une déficience fonctionnelle. On démontre que les cellules T DN non-transgéniques éliminent des cellules B chargées avec des antigènes d'îlots, mais pas des cellules B chargées avec un antigène non reconnu, in vitro. Par ailleurs, on établit que le transfert des cellules T DN activées peut empêcher le développement du diabète auto-immun dans un modèle de souris non-transgénique. De plus, nous observons que les cellules T DN migrent aux îlots pancréatiques, et subissent une activation et une prolifération préférentielles au niveau des ganglions pancréatiques. D'ailleurs, le transfert des cellules T DN entraîne une diminution d'auto-anticorps spécifiques de l'insuline et de cellules B de centres germinatifs directement dans les îlots, ce qui corrèle avec les résultats décrits ci-dessus. Les résultats présentés dans cette thèse permettent de démontrer la fonction des cellules T DN in vitro et in vivo, ainsi que leur potentiel lié à la thérapie cellulaire pour le diabète auto-immun.

PAS-1, an Ascaris suum Protein, Modulates Allergic Airway Inflammation via CD8(+) gamma delta TCR(+) and CD4(+) CD25(+) FoxP3(+) T Cells

We have previously demonstrated that PAS-1, a 200 kDa protein from Ascaris suum, has a potent immunomodulatory effect on humoral and cell-mediated responses induced by APAS-3 (an allergenic protein from A. suum) or unrelated antigens. In this study, we investigated the mechanisms by which PAS-1 is able to induce this effect on an allergic airway inflammation induced by OVA in mice. C57BL/6 mice were adoptively transferred on day 0 with seven different PAS-1-primed cell populations: PAS-1-primed CD19(+) or B220(+) or CD3(+) or CD4(+) or CD8(+) or CD4(+) CD25) or CD4(+) CD25(+) lymphocytes. These mice were immunized twice with OVA and alum by intraperitoneal route (days 0 and 7) and challenged twice by intranasal route (days 14 and 21). Two days after the last challenge, the airway inflammation was evaluated by antibody levels, cellular migration, eosinophil peroxidase levels, cytokine and eotaxin production, and pulmonary mechanical parameters. Among the adoptively transferred primed lymphocytes, only CD4(+) CD25(+), CD8(+) or the combination of both T cells impaired the production of total IgE and OVA-specific IgE and IgG1 antibodies, eosinophilic airway inflammation, Th2-type cytokines (IL-4, IL-5 and IL-13), eotaxin release and airway hyperreactivity. Moreover, airway recruited cells from CD4(+) CD25(+) and CD8(+) T-cell recipient secreted more IL-10/TGF-beta and IFN-gamma, respectively. Moreover, we found that PAS-1 expands significantly the number of CD4(+) CD25(+) FoxP3(+) and CD8(+) gamma delta TCR(+) cells. In conclusion, these findings demonstrate that the immunomodulatory effect of PAS-1 is mediated by these T-cell subsets.

CD4(+) CD25(+) Foxp3(+) Regulatory T Cells, Dendritic Cells, and Circulating Cytokines in Uncomplicated Malaria: Do Different Parasite Species Elicit Similar Host Responses?

Clearing blood-stage malaria parasites without inducing major host pathology requires a finely tuned balance between pro- and anti-inflammatory responses. The interplay between regulatory T (Treg) cells and dendritic cells (DCs) is one of the key determinants of this balance. Although experimental models have revealed various patterns of Treg cell expansion, DC maturation, and cytokine production according to the infecting malaria parasite species, no studies have compared all of these parameters in human infections with Plasmodium falciparum and P. vivax in the same setting of endemicity. Here we show that during uncomplicated acute malaria, both species induced a significant expansion of CD4(+) CD25(+) Foxp3(+) Treg cells expressing the key immunomodulatory molecule CTLA-4 and a significant increase in the proportion of DCs that were plasmacytoid (CD123(+)), with a decrease in the myeloid/plasmacytoid DC ratio. These changes were proportional to parasite loads but correlated neither with the intensity of clinical symptoms nor with circulating cytokine levels. One-third of P. vivax-infected patients, but no P. falciparum-infected subjects, showed impaired maturation of circulating DCs, with low surface expression of CD86. Although vivax malaria patients overall had a less inflammatory cytokine response, with a higher interleukin-10 (IL-10)/tumor necrosis factor alpha (TNF-alpha) ratio, this finding did not translate to milder clinical manifestations than those of falciparum malaria patients. We discuss the potential implications of these findings for species-specific pathogenesis and longlasting protective immunity to malaria.

Reduced frequency of CD4(+)CD25(HIGH)FOXP3(+) cells and diminished FOXP3 expression in patients with Common Variable Immunodeficiency: A link to autoimmunity?

Common Variable Immunodeficiency (CVID) is a primary immunodeficiency disease characterized by defective immunoglobulin production and often associated with autoimmunity. We used flow cytometry to analyze CD4(+)CD25(HIGH)FOXP3(+) T regulatory (Treg) cells and ask whether perturbations in their frequency in peripheral blood could underlie the high incidence of autoimmune disorders in CVID patients. In this study, we report for the first time that CVID patients with autoimmune disease have a significantly reduced frequency of CD4(+)CD25(HIGH)FOXP3(+) cells in their peripheral blood accompanied by a decreased intensity of FOXP3 expression. Notably, although CVID patients in whom autoimmunity was not diagnosed had a reduced frequency of CD4(+)CD25(HIGH)FOXP3(+) cells, FOXP3 expression levels did not differ from those in healthy controls. In conclusion, these data suggest compromised homeostasis of CD4(+)CD25(HIGH)FOXP3(+) cells in a subset of CVID patients with autoimmunity, and may implicate Treg cells in pathological mechanisms of CVID. (C) 2009 Elsevier Inc. All rights reserved.

Expansion of CD4(+) CD25(+) Foxp3(+) T cells by bone marrow-derived dendritic cells

Dendritic cells (DCs) are the most important antigen-presenting cells of the immune system and have a crucial role in T-lymphocyte activation and adaptive immunity initiation. However, DCs have also been implicated in maintaining immunological tolerance. In this study, we evaluated changes in the CD4(+) CD25(+) Foxp3(+) T-cell population after co-culture of lymph node cells from BALB/c mice with syngeneic bone marrow-derived DCs. Our results showed an increase in CD4(+) CD25(+) Foxp3(+) T cells after co-culture which occurred regardless of the activation state of DCs and the presence of allogeneic apoptotic cells; however, it was greater when DCs were immature and were pulsed with the alloantigen. Interestingly, syngeneic apoptotic thymocytes were not as efficient as allogeneic apoptotic cells in expanding the CD4(+) CD25(+) Foxp3(+) T-cell population. In all experimental settings, DCs produced high amounts of transforming growth factor (TGF)-beta. The presence of allogeneic apoptotic cells induced interleukin (IL)-2 production in immature and mature DC cultures. This cytokine was also detected in the supernatants under all experimental conditions and enhanced when immature DCs were pulsed with the alloantigen. CD4(+) CD25(+) Foxp3(+) T-cell expansion during co-culture of lymph node cells with DCs strongly suggested that the presence of alloantigen enhanced the number of regulatory T cells (Tregs) in vitro. Our data also suggest a role for both TGF-beta and IL-2 in the augmentation of the CD4(+) CD25(+) Foxp3(+) population.

Leukotriene B(4) is essential for selective eosinophil recruitment following allergen challenge of CD4(+) cells in a model of chronic eosinophilic inflammation

Subcutaneous heat-coagulated egg white implants (EWI) induce chronic, intense local eosinophilia in mice, followed by asthma-like responses to airway ovalbumin challenge. Our goal was to define the mechanisms of selective eosinophil accumulation in the EWI model. EWI carriers were challenged i.p. with ovalbumin and the contributions of cellular immunity and inflammatory mediators to the resulting leukocyte accumulation were defined through cell transfer and pharmacological inhibition protocols. Eosinophil recruitment required Major Histocompatibility Complex Class It expression, and was abolished by the leukotriene B4 (LTB4) receptor antagonist CP 105.696, the 5-lipoxygenase inhibitor BWA4C and the 5-lipoxygenase activating protein inhibitor MK886. Eosinophil recruitment in EWI carriers followed transfer of: a) CD4(+) (but not CD4(-)) cells, harvested from EWI donors and restimulated ex vivo; b) their cell-free supernatants, containing LTB4. Restimulation in the presence of MK886 was ineffective. CC chemokine receptor ligand (CCL)5 and CCL2 were induced by ovalbumin challenge in vivo. mRNA for CCL17 and CCL11 was induced in ovalbumin-restimulated CD4(+) cells ex vivo. MK886 blocked induction of CCL17 Pretreatment of EWI carriers with MK886 eliminated the effectiveness of exogenously administered CCL11, CCL2 and CCL5. In conclusion, chemokine-producing, ovalburnin-restimulated CD4(+) cells initiate eosinophil recruitment which is strictly dependent on LTB4 production. (C) 2008 Elsevier B.V. All rights reserved.