Search

Background : Vitamin D has regulatory effects on different cells of the immune system and low levels are associated with several immune-mediated diseases. Aim : To investigate the association between neonatal 25-hydroxy vitamin D (25-OHD) level and the expression of lymphocyte activation markers (HLA-DR, CD69, CD25, CD45RA) on T-lymphocyte subpopulations and its impact in neonatal infection. Methods : 25-OHD level was measured in the cord blood of 56 neonates and their mothers using an enzyme immune-assay method. Based on the 25-OHD level, infants were categorised into four groups: severe deficiency ( n =  7), moderate deficiency ( n =  21), mild deficiency ( n =  15) and normal 25-OHD level ( n =  13). Mothers were classified into deficient ( n =  18), insufficient ( n =  21) and normal levels ( n =  17). T-lymphocyte subpopulations and lymphocyte activation markers were investigated using flow cytometry. Results : There was a positive correlation between maternal and cord blood 25-OHD levels ( r  = 0.503, p  = 0.001). The group with severe 25-OHD deficiency had the significantly lowest level of total lymphocytes, CD3+ T lymphocytes, CD4+ T-helper and CD8+ T-cytotoxic lymphocytes and CD4+CD45RA+ naïve T-cells compared with the other groups. The frequencies of CD8+CD25+, CD4+CD25+ and CD4+HLA-DR+ activated T-lymphocytes were significantly lower in the severe, moderate and mild deficiency groups than in the normal group. Seven of 43 (16.27%) infants with 25-OHD deficiency were admitted with sepsis to the neonatal intensive care unit and there were no cases of sepsis in the normal 25-OHD group. Conclusion : Vitamin D deficiency is associated with a reduction of lymphocyte subsets and altered T-lymphocyte activation which are considered to be risk factors for neonatal infection.

CD83 is a widely recognized surface marker for mature dendritic cells, which are essential for priming naïve CD4+ T cells into effector cells. However, CD83 is also expressed on activated CD4+ T cells, which remains an enigma in T‑cell mediated immunity. Therefore, the identification of the biological features and regulation of the expression of CD83 on activated CD4+ T cells is important in understanding the function of CD83 in the adaptive immune response. The present study revealed a time‑dependent manner of the expression of CD83 on anti‑CD3/CD28‑stimulated human CD4+ T cells, which is characterized by the maximum expression at day 2 and a significant decrease at day 3. The reduced expression is not a result of a reduced rate of cell proliferation. The activation of interleukin‑2 and secretion of interferon‑γ accumulated progressively from day 1 to 3. Of note, sustained expression of CD83 was observed when CD4+ T cells were induced by transforming growth factor-β to differentiate into CD4+CD25+ forkhead box P3+ regulatory T (iTreg) cells. Confocal immunofluorescence microscopy analysis demonstrated that CD83 was highly co‑localized with CD25 on activated CD4+ T cells. In conclusion, the findings of the present study suggested that the continuous expression of CD83 on activated human CD4+ T cells is correlated with their differentiation into iTreg cells.

Introduction: Kaqun-water contains a high amount of stable oxygen, which absorbed through the skin and intestinal tract, increases tissue oxygenation., Aim: The aim of the authors was to evaluate the effect of 21 days of Kaqun-water treatment on the immune parameters of healthy volunteers., Method: Subpopulations of lymphocytes were determined by immune phenotyping, and CD25 and CD71 activation antigens were used to assess lymphocyte activation. Production of reactive oxygen intermediates was measured to determine the killing capacity of neutrophil granulocytes. Data was analysed with repeated measures ANOVA., Results: The reactive oxygen intermediate production of neutrophils increased significantly in stimulated samples during three weeks of Kaqun-water treatment. The percent of activated, CD25 positive T and helper T cells, and the ratio of NK cells increased., Conclusions: The increase in oxygen concentration caused by Kaqun-water treatment affects several immune functions: the killing potential of neurophil granulocytes is enhanced, the activation of lymphocytes shows an increased activity of immune function, and the elevated ratio of NK cells may help combat virally infected and tumorous cells.

Extrapulmonary tuberculosis may be due to underlying immune compromise. Immunosuppressive regulatory T cells (Treg cells), and CD4(+) T lymphocytes in general, are important in the host immune response to Mycobacterium tuberculosis. We evaluated T lymphocytes from patients after recovery from extrapulmonary tuberculosis, which may reflect conditions before M. tuberculosis infection. A case-control study was conducted among HIV-uninfected adults with previously treated extrapulmonary tuberculosis and 3 sets of controls: (i) subjects with previously treated pulmonary tuberculosis, (ii) close tuberculosis contacts with M. tuberculosis infection, and (iii) close tuberculosis contacts with no infection. Monocyte-depleted peripheral blood mononuclear cells (PBMC-M) were stained for CD4(+) CD25(hi) CD127(low) FoxP3(+) cell (Treg cell) and T lymphocyte activation. Both characteristics were compared as continuous variables between groups with the Kruskal-Wallis test. There were 7 extrapulmonary tuberculosis cases, 18 pulmonary tuberculosis controls, 17 controls with M. tuberculosis infection, and 18 controls without M. tuberculosis infection. The median Treg cell proportion was highest among persons with previous extrapulmonary tuberculosis (1.23%) compared to subjects with pulmonary tuberculosis (0.56%), latent M. tuberculosis infection (0.14%), or no M. tuberculosis infection (0.20%) (P = 0.001). The median proportion of CD4(+) T lymphocytes that expressed the activation markers HLA-DR and CD38 was highest for CD4(+) T lymphocytes from persons with previous extrapulmonary tuberculosis (0.79%) compared to subjects with pulmonary tuberculosis (0.44%), latent M. tuberculosis infection (0.14%), or no M. tuberculosis infection (0.32%) (P = 0.005). Compared with controls, persons with previously treated extrapulmonary tuberculosis had the highest Treg cell frequency, but also the highest levels of CD4(+) T lymphocyte activation. Immune dysregulation may be a feature of individuals at risk for extrapulmonary tuberculosis.

Background: Regulatory T cells have been implicated in the pathogenesis of COPD by the increased expression of CD25 on helper T cells along with enhanced intracellular expression of FoxP3 and low/absent CD127 expression on the cell surface., Method: Regulatory T cells were investigated in BALF from nine COPD subjects and compared to fourteen smokers with normal lung function and nine never-smokers., Results: In smokers with normal lung function, the expression of CD25+CD4+ was increased, whereas the proportions of FoxP3+ and CD127+ were unchanged compared to never-smokers. Among CD4+ cells expressing high levels of CD25, the proportion of FoxP3+ cells was decreased and the percentage of CD127+ was increased in smokers with normal lung function. CD4+CD25+ cells with low/absent CD127 expression were increased in smokers with normal lung function, but not in COPD, when compared to never smokers., Conclusion: The reduction of FoxP3 expression in BALF from smokers with normal lung function indicates that the increase in CD25 expression is not associated with the expansion of regulatory T cells. Instead, the high CD127 and low FoxP3 expressions implicate a predominantly non-regulatory CD25+ helper T-cell population in smokers and stable COPD. Therefore, we suggest a smoking-induced expansion of predominantly activated airway helper T cells that seem to persist after COPD development.

Rheumatoid arthritis (RA) is a chronic inflammatory disease, with a clinical manifestation both systemic and in joints. It has been suggested that age at disease onset and/or patients' age have influence on disease activity and clinical outcome. The reasons for the different course of RA in older people are not known; however, the activation status of peripheral blood lymphocytes could be responsible. Our aim was to relate expression of activation markers in peripheral blood CD4(+) T cells of RA patients with patients' age and/or onset age and disease activity measured by DAS28. Seventy RA patients were included into the immunological study. Two separation criteria were performed: based on age of RA onset and on the biological age of patients. We examined different activation markers, CD69, CD25, CD95 and human leucocyte antigen D-related (HLA-DR), on the CD4(+) T cell surface. Division of RA patients in 10-year intervals at 40, 50 and 60 years revealed that RA patients with later disease onset were characterized by higher DAS28. This phenomenon was not limited to the division at 60 years of age but, surprisingly, the major differences were found for the 40-year onset division. Analysis of all four components of DAS28 revealed that disease activity in older disease onset was dependent on all components. Older-onset RA patients had a higher percentage of CD4(+) CD25(+) and CD4(+) CD95(+) T cells. Summarizing the major differences in DAS28 and activation status of CD4(+) T cells observed for onset of disease at 40 years seems to be the most informative about the immunological status of RA patients., (© 2010 The Authors. Clinical and Experimental Immunology © 2010 British Society for Immunology.)

Vγ9Vδ2 T lymphocytes have been shown to respond to a variety of non-peptide antigens including alkylamines and phosphoantigens. Recently, aminobisphosphonates have also been shown to stimulate this subset of γδ+ T cells. In this study we analyzed the proliferative responses of freshly isolated γδ T lymphocytes obtained from human cord blood when challenged with pyrophosphomonoesters or aminobisphosphonates. Nitrogen-containing aminobisphopsphonates, in contrast to phoshoantigens, readily stimulated expansion of Vδ2Vγ9 cells in human cord blood. Expanded cells displayed an activated mature phenotype, and were capable of producing TNFalpha and IFNgamma but not perforin following secondary stimulation, consistent with the development of a regulatory, as opposed to cytotoxic, phenotype. This approach may provide a useful strategy for a new approach to the treatment of neonatal pathologies.

Objectives: Regulatory T cells (T(REG)) represent a T cell subset able to modulate immune response by suppressing autoreactive T-lymphocytes. The evidence of a reduced number and an impaired function of this cell population in autoimmune/inflammatory chronic diseases led to the hypothesis of its involvement in the pathogenesis of these disorders. Glucocorticoid-induced TNFR-related protein (GITR) is a well known marker of murine T(REG) cells, but little is known in humans. The aim of this study was to investigate the characteristics of T(REG) cells in systemic lupus erythematosus (SLE) and the potential role of GITR as marker of human T(REG)., Methods: Nineteen SLE patients and 15 sex- and age-matched normal controls (NC) were enrolled. CD4(+) T cells were magnetic sorted from peripheral blood by negative selection. Cell phenotype was analyzed through flow-cytometry using primary and secondary antibodies and real time polymerase-chain reaction (PCR) using TaqMan probes., Results: The CD25(high)GITR(high) subset was significantly decreased in SLE patients with respect to NC (0.37±0.21% vs 0.72±0.19%; p<0.05). On the opposite, the CD25⁻GITR(high) cell population was expanded in the peripheral blood of SLE patients (3.5±2.25 vs 0.70±0.32%, p<0.01). Interestingly, FoxP3 at mRNA level was expressed in both CD25⁻GITR(high) and CD25(high)GITR(high) cells, suggesting that both cell subsets have regulatory activity., Conclusions: CD4(+)CD25⁻GITR(high) cells are increased in SLE as compared to NC. The expression of high level of GITR, but not CD25, on FoxP3+ cells appears to point to a regulatory phenotype of this peculiar T cell subset.

Objectives: Researchers recently discovered a group of semimature dendritic cells that induce autoimmune tolerance by activating host antigen-specific CD4+CD25+ T-regulatory cells. We hypothesized that donor semimature dendritic cells injected into recipients would induce effector T-cell hyporesponsiveness by activating CD4+CD25+ T-regulatory cells., Materials and Methods: Donor myeloid semimature dendritic cells were cultivated for 6 days and were then stimulated with tumor necrosis factor a for 24 hours. BALB/c mice were pretreated with semimature dendritic cells to generate antigen-specific CD4+CD25+ T-regulatory cells in vivo. The role of CD4+CD25+ T-regulatory cells in transplant immunity was studied via mixed lymphocyte culture in vitro., Results: Surface markers and cytokines secreted by semimature dendritic cells differed from those secreted by immature myeloid dendritic cells or mature dendritic cells. Semimature dendritic cells and immature myeloid dendritic cells did not activate allogenic lymphocyte responses in coculture studies. CD4+CD25+ T-regulatory cells of recipients challenged by donor semimature dendritic cells, which expressed a high level of interleukin-10, induced hyporesponsiveness in host effector T cells that were stimulated by donor splenocytes. In contrast, CD4+CD25+ T-regulatory cells did not induce hyporesponsiveness in effector T cells when the host T cells were stimulated by third-party antigen from DBA2 mice splenocytes., Conclusions: Our findings confirm that semimature dendritic cells are an independent subgroup of dendritic cells in both immune function and morphologic profile. It may be the cytokine secretion profile of semimature dendritic cells (rather than that of surface markers) that has a key role in inducing CD4+CD25+ T-regulatory cells to express a high level of interleukin-10. Immunization with donor semimature dendritic cells may be an effective method of inducing transplant tolerance, but further evidencebased studies of that topic are necessary.

Background: Abnormally high serum uric acid levels have been associated with several disease conditions including gout and kidney stone disease. More recently, it was shown that uric acid crystals stimulate dendritic cell maturation, activate the NALP3 inflammasome, and enhance antigen-specific immune responses. We hypothesize that uric acid can also stimulate T cells directly and in the absence of antigen presentation., Methods: Purified primary human T cells were incubated with and without uric acid at concentrations of 50, 100, 150, and 200 microg/mL. The expression of T-cell activation markers CD25 and CD70 was assessed by flow cytometry. In other experiments, Jurkat T cells were used and the expression of the costimulatory molecule CD70 was determined at the mRNA level., Results: Uric acid directly activates primary human T cells in the absence of antigen presentation. Furthermore, primary human T cells and Jurkat T cells treated with uric acid overexpress the costimulatory molecule CD70, which plays an important role in T cell-B cell interaction and antibody production., Conclusions: The finding that uric acid directly promotes T-cell activation in an antigen-independent system is novel and might play a mechanistic role in the inflammatory response observed in gouty arthritis and other immune-mediated diseases.

T lymphocytes play central roles in adaptive immunity. Lycium barbarum L. (L. barbarum), also known as wolfberry, is a Chinese herbal medicine with various biological activities, such as enhancing immunity, protecting liver damage, and reducing the side effects of chemotherapy and radiotherapy. Here, we report that polysaccharide-protein complex from L. barbarum (LBP) is able to activate T cells. LBP was isolated from L. barbarum and separated to five homogenous fractions, designated LBPF1, LBPF2, LBPF3, LBPF4, and LBPF5. We found that LBP, LBPF4, and LBPF5 significantly stimulated mouse splenocyte proliferation. The proliferation proved to be of T cells, but not B cells. Cell cycle profile analysis indicated that LBP, LBPF4, and LBPF5 markedly reduced sub-G1 cells. LBP, LBPF4, and LBPF5 could activate transcription factors NFAT and AP-1, prompt CD25 expression, and induce IL-2 and IFN-gamma gene transcription and protein secretion. LBP (i.p. or p.o.) significantly induced T cell proliferation. Our results suggest that activation of T lymphocytes by LBP may contribute to one of its immuno-enhancement functions.

Purpose of Review: Although a plethora of data indicate the importance of regulatory T cells (Tregs) in experimental and clinical transplantation, are we any closer to seeing these cells as therapeutic tools in the clinic? This review discusses the functional and practical aspects of using CD4+CD25hiFoxp3+ Tregs as cellular therapeutic products in clinical transplantation, focusing on the requirements in terms of phenotype, antigen specificity and preparation of Tregs., Recent Findings: Following the emergence of new phenotypic markers of Tregs as well as improved isolation methods, a few milestone clinical trials employing the adoptive transfer of Tregs are now underway. Although mounting data suggest that alloantigen-specific Tregs may provide higher therapeutic benefits in solid organ transplantation compared with polyclonal Tregs, it seems that the specificty of Treg selected for use will need to be tailored to each clinical transplantation setting. In addition, recent findings imply that immunosuppressive regimes will also need to be reevaluated in order to complement this therapeutic strategy., Summary: Although many key questions about Tregs remain, we are undoubtedly entering an exciting era of Treg research in clinical transplantation. As renewed efforts focus on translational medical research, it seems as though, whether ready or not, Tregs are finally crossing from bench to bedside.

Background: It has been suggested that allergic diseases are caused by defective suppression of allergen-specific Th2 cells by CD4(+)CD25(+) regulatory T cells. However, such studies have been hampered by the difficulty in distinguishing regulatory T cells from CD25-expressing activated T cells. Recently, it was shown that conventional T cells expressed high levels of CD127, whereas regulatory T cells were CD127(lo), allowing discrimination between these distinct T cell subpopulations., Objective: The aim of this study was to study whether the putative regulatory subset defined as CD4(+)CD25(+)CD127(lo) was involved in grass pollen-reactive T cell responses., Methods: Peripheral blood mononuclear cells (PBMCs) were obtained from allergic donors and non-atopic controls out of season. Grass pollen-induced cytokine production and proliferation were compared in cultures of undepleted cells and cells depleted of CD4(+)CD25(+), CD4(+)CD25(+)CD127(hi) or CD4(+)CD25(+)CD127(lo) T cells., Results: Undepleted cell cultures from allergic patients showed significantly increased proliferation and Th2 cytokine production compared with non-atopic controls. Depletion of all CD25(+) T cells did not increase cytokine production or proliferation, and more importantly, no increase in Th2 cytokine production or proliferation was observed in cell cultures depleted of CD4(+)CD25(+)CD127(lo) cells (putative regulatory T cells) compared with undepleted PBMCs in both the allergic and the non-atopic group., Conclusion: Our study showed that T cells from grass pollen-allergic patients and non-atopic controls responded very differently to grass pollen extract, but this difference could not be explained by differences in regulatory T cell function. Further studies are needed to understand the importance of regulatory T cells in allergy.

The activity of substance P (SP) in the rat thymus seems to be tightly controlled by its bioavailability. In this study, we provide evidence for the expression of the SP-degrading enzyme, neutral endopeptidase (NEP)/CD10, by rat thymocyte subsets, and we illustrate its involvement in the in vivo SP/neurokinin-1 receptor (NK(1)R)-mediated regulation of thymocyte survival and proliferation. NEP/CD10 was expressed at both mRNA and protein levels on a substantial portion (45.5%) of CD5(+) thymocytes, namely on the CD4(+)CD8(+) (double positive; DP) and CD4(+) subsets. Continuous administration of thiorphan, a specific NEP/CD10 inhibitor, by means of miniosmotic pumps, enhanced rat thymocyte preprotachykinin-A (PPT-A) and NK(1)R mRNA expression as well as SP and NK(1)R protein levels in an NK(1)R-dependent manner. Thiorphan increased CD10(+)CD4(+) and CD10(+)DP thymocyte numbers, and an NK(1)R antagonist, (S)1-{2-[3(3-4-dichlorophenyl)-1-(3-iso-propoxyphenylacetyl)-piperidine-3-yl]ethyl}-4-pheny-1-azoniabicyclo[2.2.2]octane, chloride (SR140333), abrogated these stimulatory effects. In addition, the NEP/CD10 inhibitor stimulated interleukin (IL)-2 production, IL-2 receptor alpha chain expression, and concanavalin A-induced proliferation of CD5(+) thymocytes, and it inhibited spontaneous and NK(1)R-dependent thymocyte apoptosis. The thiorphan-protective antiapoptotic and proliferative effects involved the activation of Akt serine-threonine kinase, subsequent up-regulation of survivin mRNA, down-regulation of procaspase-3 mRNA levels, and suppression of caspase-3 activity, which were inhibited by SR140333 and mimicked by exogenous SP administration. Overall, our findings suggest that by controlling SP availability, NEP/CD10 negatively regulates thymocyte homeostasis and development.

Background and Aim: Hepatitis A virus (HAV) causes a transient illness leaving permanent protection against reinfection. Few data are available on the regulatory mechanisms involved in the CD4+ T helper activation. We aimed to investigate the frequency and function of CD3+/CD4+/CD25+ T cells with regulatory function (Tregs) during acute HAV infection., Methods: We enrolled 35 consecutive patients and 15 healthy donors, enumerated Tregs by flow cytometry assay and evaluated, after immunomagnetical sorting with magnetic beads, their ability to inhibit the proliferation of CD4+/CD25- T lymphocytes at different ratios (1:1, 1:10, 1:20)., Results: All patients had the usual course of infection. Our immunological analysis showed Tregs frequency in these patients (6.5% [range, 5-8.8%]; 36 [range, 10-87] cells) did not have any statistical difference compared with healthy donors (6% [range, 5-8%]; 48 (range, 23-71) cells), while their ability to suppress CD4+/CD25- was drastically reduced at different ratios (Mann-Whitney U-test; ratio 1:1, 93% vs 72%, z = -3.34, P < 0.0001; ratio 1:10, 86% vs 51%, z = -4.04, P < 0.001; ratio 1:20, 56% vs 30%, z = -3.43, P < 0.0001). After the seroconversion, CD4+/CD25+ frequency and function in HAV-infected patients did not differ from healthy individuals., Conclusion: CD4+/CD25+ T cells seem to be impaired in their function during the HAV acute infection. This evidence might help to determine an optimal T helper cell immune network that is a predisposing factor for a self-limiting disease.

Engagement of glucocorticoid-induced TNFR-related protein (GITR) enables the costimulation of both CD25(-)CD4(+) effector (Teff) and CD25(+)CD4(+) regulatory (Treg) cells; however, the effects of GITR-costimulation on Treg function remain controversial. In this study, we examined the effects of GITR ligand (GITRL) binding on the respective functions of CD4(+) T cells. GITRL-P815 transfectants efficiently augmented anti-CD3-induced proliferation and cytokine production by Teff cells. Proliferation and IL-10 production in Treg were also enhanced by GITRL transfectants when exogenous IL-2 and stronger CD3 stimulation was provided. Concomitant GITRL-costimulation of Teff and Treg converted the anergic state of Treg into a proliferating state, maintaining and augmenting their function. Thus, GITRL-costimulation augments both effector and regulatory functions of CD4(+) T cells. Our results suggest that highly activated and increased ratios of Treg reverse the immune-enhancing effects of GITRL-costimulation in Teff, which may be problematic for therapeutic applications using strong GITR agonists.

CD4+CD25+ human regulatory T cells (Treg cells), which express the transcription factor FoxP3, suppress T cell activation. In this study, we sought to define cellular and molecular mechanisms of human Treg cell differentiation. A subset of human naive CD4+ T cells that are CD25+ express high levels of FoxP3. We show that upon activation through the TCR, these FoxP3-expressing naive T cells (termed TNreg cells) greatly expand in vitro. Expanded TNreg cells acquire a full Treg phenotype with potent suppressive activity and display low IL-2 production upon TCR stimulation. TNreg cells in which FoxP3 expression was reduced through RNA interference lost their suppressive activity, but retained their low IL-2 secretion in response to TCR stimulation. Furthermore, in support of the notion that TNreg cells represent a separate lineage of naive cells, we found that they were more susceptible to HIV infection as compared with naive CD4+ T cells. Based on these findings, we propose that TNreg cells are precursors for human Treg cells and that these cells require a high level of FoxP3 expression to maintain their suppressive function. Accordingly, modulation of TNreg cell numbers during infections such as HIV may disrupt human Treg cell development, and contribute to chronic immune activation.

Foxp3+ regulatory T cells (Tregs) play a pivotal role in the maintenance of peripheral T cell tolerance and are thought to interact with dendritic cells (DC) in secondary lymphoid organs. We analyzed here the in vivo requirements for selective expansion of Ag-specific Treg vs CD4+CD25- effector T cells and engagement of Ag-specific Treg-DC interactions in secondary lymphoid organs. Using i.v. Ag delivery in the absence of inflammation, we found that CD4+CD25+Foxp3+ Tregs undergo vigorous expansion and accumulate whereas naive CD4+CD25-Foxp3- T cells undergo abortive activation. Quantifying directly the interactions between Tregs and CD11c+ DC, we found that Tregs establish cognate contacts with endogenous CD11c+ DC in spleen and lymph nodes at an early time point preceding their expansion. Importantly, we observed that as few as 10(3) Tregs selectively expanded by i.v. Ag injection are able to suppress B and T cell immune responses in mouse recipients challenged with the Ag. Our results demonstrate that Tregs are selectively mobilized by Ag recognition in the absence of inflammatory signals, and can induce thereafter potent tolerance to defined Ag targets.

In vivo immunomodulatory effect of essential oil of niaouli (EON) was investigated using a mouse model, in which mice were immunized with keyhole limpet hemocyanin (KLH) and intraperitoneally given EON (less than 500 microl kg(-1) body weight). In vivo efficacy of EON for immune potentiation was convinced by significantly higher expression of an activation marker, CD25, on freshly isolated draining lymph node (LN) T cells, but not B cells. However, immunofluoresence analysis failed to show any proportional change in T/B and CD4(+)/CD8(+) T cell ratios. Data of KLH-specific immunoglobulin serum levels showed that EON does not affect humoral immune response. Instead, proliferative response and IFNgamma production of LN T cells ex vivo stimulated with KLH were significantly higher in EON-treated group, but not IL-2 and IL-4 production. These results clearly show that EON preferentially upregulates T-cell mediated cellular immunity. We further clarified the accessory cells' contribution to the EON-mediated potentiation of cellular immunity and found considerably higher production of and TNF-alpha and IL-12 by splenic macrophages from EON-treated mice when stimulated with lipopolysaccharide (LPS) and IFNgamma. Collectively, in vivo EON treatment potentiates T cell-mediated cellular immunity and macrophage activity, but not humoral immunity. The current study provides a rationale for clinical application of EON to control infectious diseases, in particular, those caused by intracellular pathogens.

The concept of immune regulation/suppression has been well-established and, besides thymus-derived CD4+CD25+ regulatory T (TR) cells, it became clear that a variety of additional peripherally induced TR cells play vital roles in protection from many harmful immune responses including intestinal inflammation. In the present study, we have analyzed in vivo-induced Ag-specific CD4+ TR cells with respect to their molecular and functional phenotype. By comparative genomics we could show that these Ag-specific TR cells induced by chronic Ag stimulation in vivo clearly differ in their genetic program from naturally occurring thymus-derived CD4+CD25+ TR cells. This distinct population of induced TR cells express neither CD25 nor the TR-associated transcription factor Foxp3. Strikingly, CD25 is not even up-regulated upon stimulation. Despite the lack in Foxp3 expression, these in vivo-induced CD25- TR cells are able to interfere with an Ag-specific CD8+ T cell-mediated intestinal inflammation without significant increase in CD25 and Foxp3 expression. Thus, our results demonstrate that in vivo-induced Ag-specific TR cells represent a distinct population of Foxp3-CD25- TR cells with regulatory capacity both in vitro and in vivo.

Naturally occurring Foxp3(+)CD4(+)CD25(+) T cells (nTregs) isolated from lungs of naive mice regulate allergic airway hyperresponsiveness (AHR) and inflammation. Here, we demonstrate the critical requirement for engagement of MHC class I on CD4(+)CD25(+) T cells by CD8 for the functional activation of these nTregs. Suppression of allergen-induced AHR and inflammation by nTregs was abolished in mice treated with anti-CD8. Correspondingly, decreased levels of IL-10 and TGF-beta and increased levels of Th2 cytokines in bronchoalveolar lavage were detected in these treated mice. Similarly, nTregs isolated from beta2m(-/-) mice or from mice treated with anti-MHC I antibody in vitro before intratracheal transfer failed to modulate AHR or inflammation. Coculture of nTregs with CD8(+) T cells increased IL-10 and TGF-beta. Addition of anti-MHC I or anti-CD8 reduced IL-10 and TGF-beta. These results demonstrate that functional activation of nTregs requires the interaction between MHC I on CD4(+)CD25(+) T cells and CD8.

The Cdk9/Cyclin T1 complex is very important in controlling specific differentiative pathways of several cell types, including muscle cells and neurons. We recently demonstrated the involvement of this complex in B cell activation/differentiation. To check whether the Cdk9/Cyclin T1 complex is also involved in the T cell activation/differentiation process, we isolated different T cell populations by magnetic separation, based on their surface antigens. We observed that the expression level of Cdk9/Cyclin T1 increases in effector T cells (CD27(+)), as well as in activated T cells (CD25(+)) and memory T cells (CD45RA(-)), thus suggesting a specific upregulation of the Cdk9/Cyclin T1 complex following antigen encounter. We have previously demonstrated that in B cells, Cdk9 interacts in vivo with the E2A gene products E12/E47 (members of the basic helix-loop-helix family) which are involved in differentiation. In this article, we show that this interaction also occurs in T cells. This suggests an active role for the Cdk9/Cyclin T1 complex during lymphoid differentiation, through physical binding with E12 and E47. These preliminary results suggest that the Cdk9/Cyclin T1 complex may be important in the activation and differentiation program of lymphoid cells and that its upregulation, which is due to still unknown mechanisms, may contribute to malignant transformation.

Naturally occurring CD4(+)CD25(+) regulatory T cells (nT(R)) comprise a separate lineage of T cells that are essential for maintaining immunological tolerance to self. Here we demonstrate that the level of phosphorylation of the TCR zeta-chain is approximately 1.5- to 4-fold higher in nT(R) as compared with CD4(+)CD25(-) T cells. The increased level of TCR zeta-chain phosphorylation is presumably secondary to their higher affinity for self, resulting in a stronger TCR signal as it was completely blocked by treatment with anti-MHC class II. The enhanced level of TCR zeta-chain phosphorylation was correlated with the capacity of nT(R) to develop non-specific suppressor effector function following culture with IL-2 or IL-4 in the absence of TCR stimulus. Thus, a sub-population of nT(R) is activated by recognition of self-peptide-MHC class II ligands in vivo, resulting in their capacity to be induced to mediate suppressor function in vitro in the absence of TCR stimulation.

Although standardized protocols are widely used for the generation of monocyte-derived immunostimulatory dendritic cells (DC(ims)), the inducibility of Th1 cells by DC(ims) may considerably differ. As a measure for the quality of DC(ims) generated from an individual donor at a certain time point, CD83 is used in combination with HLA-DR and CD86 to assess DC maturation. When phenotypically analyzing DC(ims), we identified a subpopulation ( approximately 60%) of CD83+, CD86+, and HLA-DR+ DC(ims) that co-expressed CD25. DC within a given DC(ims) preparation identified by lower expression of CD83 and by selective expression of CD14, however, did not co-express CD25. In order to establish CD25 as an additional maturation marker of DC(ims), we studied the DC phenotype of these cells as well as the DC-dependent T-cell proliferation and T-cell cytokine production profile after co-incubation with sorted CD25(high) and CD25(low) subpopulations of CD83+, HLA-DR+, CD86+ DC(ims). CD25(high) DC(ims) showed significant up-regulation of the DC activation molecule CD43 and induced increased levels of IL-2 secretion in allogeneic T-cells (170.7+/-86.7pg/mL) as compared to T-cells coincubated with CD25(low) DC(ims) (86.6+/-37.6pg/mL) [p=0.0224]. This was reflected by a significantly lower T-cell stimulatory capacity of CD25(low) DC(ims) (84.0% of CD25(high) DC(ims), 1:10 ratio; p=0.014) whereas the T-cell stimulatory capacity of CD25(low) DC(ims) was much higher when compared to IL-10 induced regulatory DC (55.3% of CD25(high) DC(ims); 1:10 ratio). With regard to cancer vaccination protocols, we propose to use CD25 and CD43 as additional markers for DC quality control, assessment of maturational status, and positive selection.

Toll-like receptor (TLR) ligands are notable for their ability to induce APC maturation, which in turn facilitates optimal T cell mediated immune responses. Toll-like receptor ligands, such as CpG DNA, can also modulate immune responses by blocking the suppressive effects of CD4+CD25+ regulatory T cells (Tregs). Recently, we have demonstrated that CpG DNA, in addition to its actions on APCs and Tregs, can provide direct costimulatory signals to CD4+CD25- T cells. Here we show that this costimulatory effect is sufficient to abrogate suppression by Tregs. These data indicate a previously undefined role for TLR ligands in directly modulating CD4+ T cell responses.

Regulatory T cells are crucial for maintaining T cell tolerance to-self-antigens and autoimmune process prevention. They are known to be able to cut down in vitro autologous lymphocyte proliferation in mice. It is suggested that their increased levels are responsible for immunosuppression in human tumor growth. Our investigation was concerned with identifying the extent of mitogen-stimulated T- and B-cell concentration changes as well as phenotypic variation in peripheral blood lymphocyte level in lung, stomach, breast and colorectal cancer. It was found that malignancy involves reduced mitogen (PHA, ConA, PWM)-stimulated lymphocyte proliferation and enhanced peripheral blood CD25+-cell concentration which should be accounted for by regulatory T cell increase. However, both the extent of proliferation changes and CD25+-lymphoid cell concentration considerably depended on tumor localization, which might have an indicator of a varying degree of immunosuppression involved in different neoplasms.

Mild zinc deficiency is a common condition in healthy elderly individuals leading to impaired cell-mediated immune response. Here we report the effect of improved zinc status on TH1/TH2 balance and on the activation status of T helper cells in 19 healthy elderly subjects aged 69.8 +/- 5.1 years. Our investigations revealed a mild zinc deficiency which was adjusted by oral zinc supplementation for seven weeks. Improved serum zinc levels significantly reduced levels of activated T helper cells whereas changes in TH1/TH2 ratio (determined by CCR4 and CCR5 expression) were not observed. These findings suggest that elderly individuals may benefit from moderate zinc supplementation due to improved immune response leading to reduced incidences of autoimmune diseases and infections.

T cells are known to develop a critical role in the pathogenesis of atopic dermatitis (AD) and bronchial asthma. T cells involved in AD express the skin homing receptor CLA, but no lung homing receptor has been identified in bronchial asthma. We compared different cell markers and the cytokine production in T cells from children with AD or bronchial asthma. We studied the involvement of CLA+ and CLA- T-cell subpopulations in these diseases. We studied 20 children with acute AD lesions, 15 with mild persistent asthma, and 15 non-atopic controls. All patients were sensitized to house dust mite (DP) and evaluated during the acute phase. Total and specific IgE were measured by immunoassay and the expression of different cell markers and the cytokine production was analyzed by flow cytometry in peripheral blood mononuclear cells. Total IgE was significantly higher in AD children and IgE to DP in the asthmatic children. There was a significant increase in CD25+ CD4+ cells in asthmatic children and in HLA-DR+ CD4+ and HLA-DR+ CD8+ cells in AD. In the CD4+ subsets, there was an increase in IL-13, IL-5 and TNF-alpha in AD compared to controls, a decrease in IFN-gamma in asthmatic children compared to controls, and an increase in IL-13, IL5, IL2, TNF-alpha, and IFN-gamma in the AD compared to asthmatic children. Changes in cytokine production were mainly detected in CLA+ cells in AD and in CLA- cells in asthma. Differences exist in total and specific IgE, activation markers, and cytokine patterns between AD children and children with asthma, with the former expressing a Th2 pattern whereas in asthmatic children we only detected a decrease in IFN-gamma. Moreover, the subpopulations (CLA+ vs. CLA-) expressing these changes were different, indicating that the underlying mechanisms in the two diseases are not exactly the same.