CD25 Foxp3 Research Articles

In vitro effects of binuclear (η (6)-p-cymene)ruthenium(II) complex containing bridging bis(nicotinate)-polyethylene glycol ester ligand on differentiation pathways of murine Th lymphocytes activated by T cell mitogen.

T cell differentiation into distinct T helper (Th) subpopulations is crucial in governing acquired immune responses as well as some inflammatory and autoimmune disorders. This study investigated potential of the novel neutral binuclear ruthenium(II) complexes 1-8 with general formula [{RuCl2(η(6)-p-cym)}2μ-(N(∩)N)] (N(∩)N = bis(nicotinate)- and bis(iso-nicotinate)-polyethylene glycol esters; (3-py)COO(CH2CH2O) n CO(3-py) and (4-py)COO(CH2CH2O) n CO(4-py); n = 1-4), as well as [RuCl2(η(6)-p-cym)(nic)] (R1, nic = nicotinate) and [RuCl2(η(6)-p-cym)(inic)] (R2, inic = isonicotinate) as an immunomodulatory agents capable to direct Th cell differentiation. From all investigated complexes, [{RuCl2(η(6)-p-cym)}2μ-{(3-py)COO(CH2CH2O)4CO(3-py)}] (4) was selected for further study because it did not affect splenocyte viability (in concentration up to 50 μM), but significantly reduced secretion of representative Th1 cytokine, IFN-γ induced by T cell mitogen. Besides IFN-γ, 4 inhibited dose dependently expression and production of representative Th17 cytokine, IL-17, in these cells. Otherwise, the production of anti-inflammatory cytokines IL-4 and IL-10 was upregulated. Also, 4 significantly increased CD4(+)CD25(+)FoxP3(+) Treg cell frequency in the activated splenocytes. Moreover, ConA-induced expression of Th1 transcription factors, T-bet and STAT1, as well as of Th17-related protein STAT3 was attenuated upon exposure to 4, while the expression of Th2-related transcription factor GATA3 remained stable. In conclusion, ruthenium(II) complex 4 modulates immune system cell functions in vitro by inhibiting T cell differentiation towards pathogenic Th1/Th17 phenotype and inducing a regulatory phenotype characterized by IL-10 and IL-4 production, which may provide novel therapeutic opportunities for immune-inflammatory and/or autoimmune disorders.

Low expression of CD39 on regulatory T cells as a biomarker for resistance to methotrexate therapy in rheumatoid arthritis

Rheumatoid arthritis (RA) is an inflammatory autoimmune disease characterized by joint destruction and severe morbidity. Methotrexate (MTX) is the standard first-line therapy of RA. However, about 40% of RA patients are unresponsive to MTX treatment. Regulatory T cells (Tregs, CD4(+)CD25(+)FoxP3(+)) are thought to play an important role in attenuating RA. To investigate the role of Tregs in MTX resistance, we recruited 122 RA patients (53 responsive, R-MTX; 69 unresponsive, UR-MTX) and 33 healthy controls. Three months after MTX treatment, R-MTX but not UR-MTX showed higher frequency of peripheral blood CD39(+)CD4(+)CD25(+)FoxP3(+) Tregs than the healthy controls. Tregs produce adenosine (ADO) through ATP degradation by sequential actions of two cell surface ectonucleotidases: CD39 and CD73. Tregs from UR-MTX expressed a lower density of CD39, produced less ADO, and had reduced suppressive activity than Tregs from R-MTX. In a prospective study, before MTX treatment, UR-MTX expressed a lower density of CD39 on Tregs than those of R-MTX or control (P < 0.01). In a murine model of arthritis, CD39 blockade reversed the antiarthritic effects of MTX treatment. Our results demonstrate that MTX unresponsiveness in RA is associated with low expression of CD39 on Tregs and the decreased suppressive activity of these cells through reduced ADO production. Our findings thus provide hitherto unrecognized mechanism of immune regulation in RA and on mode of action of MTX. Furthermore, our data suggest that low expression of CD39 on Tregs could be a noninvasive biomarker for identifying MTX-resistant RA patients.

A Bone Anabolic Effect of RANKL in a Murine Model of Osteoporosis Mediated Through FoxP3+ CD8 T Cells.

TNF-α and IL-17 secreted by proinflammatory T cells (T(EFF)) promote bone erosion by activating osteoclasts. We previously demonstrated that in addition to bone resorption, osteoclasts act as antigen-presenting cells to induce FoxP3 in CD8 T cells (Tc(REG)). The osteoclast-induced regulatory CD8 T cells limit bone resorption in ovariectomized mice (a murine model of postmenopausal osteoporosis). Here we show that although low-dose receptor activator of NF-κB ligand (RANKL) maximally induces Tc(REG) via Notch signaling pathway to limit bone resorption, high-dose RANKL promotes bone resorption. In vitro, both TNF-α and IL-17, cytokines that are abundant in ovariectomized animals, suppress Tc(REG) induction by osteoclasts by repressing Notch ligand expression in osteoclasts, but this effect can be counteracted by addition of RANKL. Ovariectomized mice treated with low-dose RANKL induced Tc(REG) that suppressed bone resorption, decreased T(EFF) levels, and increased bone formation. High-dose RANKL had the expected osteolytic effect. Low-dose RANKL administration in ovariectomized mice lacking CD8 T cells was also osteolytic, confirming that Tc(REG) mediate this bone anabolic effect. Our results show that although RANKL directly stimulates osteoclasts to resorb bone, it also controls the osteoclasts' ability to induce regulatory T cells, engaging an important negative feedback loop. In addition to the conceivable clinical relevance to treatment of osteoporosis, these observations have potential relevance to induction of tolerance and autoimmune diseases.

Immunomodulatory effects of Trichinella spiralis-derived excretory-secretory antigens.

Helminth-derived products, either released into the circulation during the course of the infection or isolated after in vitro cultivation of the parasite and applied by the injection, are able to suppress the host immune response to autoantigens and allergens, but mechanisms could differ. Prophylactic application of Trichinella spiralis excretory-secretory muscle larvae (ES L1) products ameliorates experimental autoimmune encephalomyelitis (EAE) with the same success as infection did. However, a shift to the Th2-type response in the periphery and in the central nervous system, accompanied by activation of regulatory mechanisms, had a striking, new feature of increased proportion of unconventional CD4(+)CD25(-)Foxp3(+) regulatory cells both in the periphery and in the central nervous system of animals treated with ES L1 before the induction of EAE.

CD103 marks a subset of human CD34+-derived langerin+ dendritic cells that induce T-regulatory cells via indoleamine 2,3-dioxygenase-1

Indoleamine 2,3-dioxygenase 1 (IDO1) is an immunosuppressive molecule expressed in some subsets of normal and neoplastic cells. Mature human dendritic cells (DCs) have been shown to express IDO1, but little is known about its expression and function during DC differentiation from bone marrow hematopoietic stem/progenitor cells (HSPCs). Here, we show that during in vitro differentiation along the myeloid DC lineage, CD34(+) HSPCs acquire IDO1 expression, which acts in a tolerogenic manner by inducing a population of fully functional CD4(+)CD25(+) FOXP3(+) T-regulatory cells. Phenotypically, CD1a(+)CD14(-) HPSC-derived DCs expressed IDO1, langerin, CD11b, and CD1c. Cell-sorting experiments demonstrated that IDO1 expression is found in a subset of CD1a(+)CD14(-)langerin(+) cells, expressing CD103, which is capable of inducing T-regulatory cells in an IDO1-dependent manner. In conclusion, DC differentiation from CD34(+) HSPCs results in the expression of a functionally active IDO1 protein in CD1a(+)langerin(+), CD103-expressing DCs. These data point toward IDO1 expression as part of a tolerogenic signature during DC development.

Manipulation of regulatory T cells and antigen-specific cytotoxic T lymphocyte-based tumour immunotherapy.

The most potent killing machinery in our immune system is the cytotoxic T lymphocyte (CTL). Since the possibility for self-destruction by these cells is high, many regulatory activities exist to prevent autoimmune destruction by these cells. A tumour (cancer) grows from the cells of the body and is tolerated by the body's immune system. Yet, it has been possible to generate tumour-associated antigen (TAA) -specific CTL that are also self-antigen specific in vivo, to achieve a degree of therapeutic efficacy. Tumour-associated antigen-specific T-cell tolerance through pathways of self-tolerance generation represents a significant challenge to successful immunotherapy. CD4(+) CD25(+) FoxP3(+) T cells, referred to as T regulatory (Treg) cells, are selected in the thymus as controllers of the anti-self repertoire. These cells are referred to as natural T regulatory (nTreg) cells. According to the new consensus (Nature Immunology 2013; 14:307-308) these cells are to be termed as (tTreg). There is another class of CD4(+) Treg cells also involved in regulatory function in the periphery, also phenotypically CD4(+) CD25(±) , classified as induced Treg (iTreg) cells. These cells are to be termed as peripherally induced Treg (pTreg) cells. In vitro-induced Treg cells with suppressor function should be termed as iTreg. These different Treg cells differ in their requirements for activation and in their mode of action. The current challenges are to determine the degree of specificity of these Treg cells in recognizing the same TAA as the CTL population and to circumvent their regulatory constraints so as to achieve robust CTL responses against cancer.

Correlation of CD₄⁺ CD₂₅⁺ Foxp₃⁺ Treg with the recovery of joint function after total knee replacement in rats with osteoarthritis.

In this study, we observed changes in CD4(+) CD25(+) Foxp3(+) Treg expression in rats with osteoarthritis (OA) to explore the role that CD4(+) CD25(+) Foxp3(+) Treg plays in the decline in the condition of OA rats. Thirty rats were randomly divided into 2 groups equally and OA was induced in rats in the model group by injection of papain and l-cysteine into the right knee joint. Cartilage lesions were scored by the modified Mankin scale; pulmonary function was assessed by spirometry; interleukin (IL)-17 and IL-4 levels were evaluated by the enzyme-linked immunosorbent assay; and the levels of CD4(+) CD25(+) Foxp3(+) Treg in peripheral blood were measured by flow cytometry. The left knee joints of the model rats appeared palpable swelling and osteophytes, while the body weight, heart and lung function of these rats decreased. The serum IL-4 level was lower, whereas the serum IL-17 level was higher in the model group (P < 0.05). The peripheral blood CD4(+) CD25(+) Foxp3(+) Treg of CD4(+)T cells was significantly lower. Correlation of the changes in the levels of IL-4, IL-17, and Treg suggests that the underlying mechanism may be a reduction of the regulatory effect of Treg. The specific mechanism still requires further study.

Promises and paradoxes of regulatory T cells in inflammatory bowel disease.

Since their discovery two decades ago, CD4(+)CD25(+)Foxp3(+) regulatory T cells (Tregs) have become the subject of intense investigation by immunologists. Unlike other T cells, which promote an immune response, Tregs actively inhibit inflammation when activated by their cognate antigen, thus raising hope that these cells could be engineered into a highly targeted, antigen-specific, immunosuppressant therapy. Although Tregs represent less than 10% of circulating CD4(+)T cells, they have been shown to play an essential role in preventing or limiting inflammation in a variety of animal models and human diseases. In particular, spontaneous intestinal inflammation has been shown to occur in the absence of Tregs, suggesting that there may be a Treg defect central to the pathogenesis of human inflammatory bowel disease (IBD). However, over the past decade, multiple groups have reported no qualitative or quantitative deficits in Tregs from the intestines and blood of IBD patients to explain why these cells fail to regulate inflammation in Crohn's disease and ulcerative colitis. In this review, we will discuss the history of Tregs, what is known about them in IBD, and what progress and obstacles have been seen with efforts to employ them for therapeutic benefit.

Influence of plasma exchange on transplantation related immune function in patients with liver failure

This study aimed to evaluate the influence of plasma exchange (PE) treatment of patients with liver failure on the patient's immune function, including peripheral blood T lymphocytes and cytokines. Patients accepting PE for liver failure from October 2011 to February 2012 were included prospectively in the research group. Peripheral blood samples were collected at set time points. The percentages of T lymphocyte subtypes were detected by flow cytometry using different fluorescence labels including CD3-FITC, CD4-PerCP, CD8-PE, CD25-FITC, and Foxp3-PE. Changes in serum IL-17 concentration were followed by ELISA. In all fifteen patients who accepted PE, the percentages of CD3(+) and CD8(+) T cells increased immediately after the procedure and then reduced gradually. These significant changes were confirmed by statistical analysis (P < 0.05). The percentage of CD4(+) T cells also increased after PE to a certain extent, but failed to show statistical significance. The positive ratio of CD4(+)CD25(+)Foxp3(+) T cells (Treg) increased after the treatment with statistical difference (P < 0.05). The concentration of IL-17 in patient serum increased significantly following PE treatment (P < 0.05). These results demonstrated that T lymphocyte subgroups of patients with liver failure could be influenced after PE treatment, and that cellular immunity could be recovered. PE treatment, therefore, can be viewed as providing reliable protection for the reconstruction of the patient immune system.

Association of CD4(+)CD25(+)FoxP3(+) regulatory T cells with natural course of childhood chronic immune thrombocytopenic purpura.

PurposeThe purpose of this study was to determine the frequency of CD4+CD25+FoxP3+ regulatory T cells (Treg) in the peripheral blood of patients with childhood chronic immune thrombocytopenic purpura (ITP) exhibiting thrombocytopenia and spontaneous remission. The findings of this study indicate the possibility of predicting spontaneous recovery and pathogenesis of childhood chronic ITP.MethodsEleven children with chronic ITP (seven thrombocytopenic and four spontaneous remission cases; mean age, 8.8 years; range, 1.7-14.9 years) were enrolled in this study. Five healthy children and eight healthy adults were included as controls. The frequency of Treg was evaluated by flow cytometry in the peripheral blood.ResultsIn this study, four patients (36%) achieved spontaneous remission within 2.8 years (mean year; range, 1.0-4.4 years). The frequency of Treg was significantly lower in patients with persisting thrombocytopenia (0.13%±0.09%, P<0.05), than that in the patients with spontaneous remission (0.30%±0.02%), healthy adults controls (0.55%±0.44%), and healthy children controls (0.46%±0.26%). A significantly positive correlation was found between the frequency of Treg and the platelet count in children.ConclusionThese data suggest that a lower frequency of Treg contributes to the breakdown of self-tolerance, and may form the basis for future development of specific immunomodulatory therapies. Furthermore, Treg frequency has prognostic implication toward the natural course and long-term outcomes of childhood chronic ITP.

Changes of naturally occurring CD4(+)CD25(+) FOXP3(+) regulatory T cells in patients with acute coronary syndrome and the beneficial effects of atorvastatin treatment.

To determine the number and function of naturally occurring CD4(+)CD25(+)FOXP3(+) regulatory T cells (nTregs) in patients with acute coronary syndrome (ACS) and to determine the effects of a low dose of atorvastatin treatment (20 mg/day) on nTregs.Patients with ACS were randomly divided into a group receiving conventional therapy (n = 60) or conventional therapy supplemented with atorvastatin (20 mg/day) (n = 60). A group of healthy volunteers that did not suffer from ACS was used as controls (n = 60). T lymphocytes were isolated from ACS patients, both before and 4 weeks after treatment, or control patients and the percentage of nTregs was determined by flow cytometry. Serum levels of cytokines were determined by enzyme-linked immunosorbent assays. A mixed lymphocyte reaction was used to determine the ability of nTregs to inhibit proliferation of effector T cells. Quantitative PCR and Western blot were used to analyze (forkhead box P3) FOXP3 mRNA transcript levels and the expression of FOXP3 protein.In ACS patients, the percentage and inhibitory properties of nTregs were reduced, IFN-γ and hsCRP levels were increased, and IL-10 and TGF-β1 levels were lowered. Atorvastatin treatment increased the percentage and inhibitory ability of nTregs, decreased serum IFN-γ and hsCRP levels, and decreased IL-10 and TGF-β1 levels, as compared with the non-atorvastatin group.Our findings suggest that nTregs play an atheroprotective role in atherosclerosis. The inhibitory effects of atorvastatin on inflammation in ACS may be due to its beneficial effects on nTregs and restoration of immune homeostasis.

Increase in Activated Treg in TIL in Lung Cancer and In Vitro Depletion of Treg by ADCC Using an Antihuman CCR4 mAb (KM2760)

Tregs infiltrate tumors and inhibit immune responses against them. We investigated subpopulations of Foxp3 CD4 T cells previously defined by Miyara et al. (Immunity 30, 899-911, 2009) in peripheral blood mononuclear cells (PBMCs) and tumor infiltrating lymphocytes (TILs) in lung cancer. We also showed that Tregs in healthy donors that express CCR4 could be efficiently eliminated in vitro by cotreatment with antihuman (h) CCR4 mAb (KM2760) and NK cells. In lung cancer, the number of activated/effector Tregs and non-Tregs, but not resting/naive Tregs, was increased in TILs compared with the number of those cells in PBMCs. The non-Treg population contained Th2 and Th17. CCR4 expression on activated/effector Tregs and non-Tregs in TILs was down-regulated compared with that on those cells in PBMCs. Chemokinetic migration of CD25 CD4 T cells containing the Treg population sorted from the PBMCs of healthy donors to CCL22/MDC was abrogated by pretreatment with anti-hCCR4 mAb (KM2760). The inhibitory activity of CD25 CD127 CD4 Tregs on the proliferative response of CD4 and CD8 T cells stimulated with anti-CD3/CD28 coated beads was abrogated by adding an anti-hCCR4 mAb (KM2760) and CD56 NK cells to the culture. The findings suggested the CCR4 on activated/effector Tregs and non-Tregs was functionally involved in the chemokinetic migration and accumulation of those cells to the tumor site. In vitro findings of efficient elimination of Tregs may give the basis for implementation of a clinical trial to investigate Treg depletion by administration of an anti-hCCR4 mAb to solid cancer patients.

Hepatitis C virus core protein triggers expansion and activation of CD4+CD25+ regulatory T cells in chronic hepatitis C patients

CD4(+)CD25(+)FoxP3(+) regulatory T cells (Tregs) are increased in patients with chronic hepatitis C, which may contribute to the sustained suppression of hepatitis C virus (HCV)-specific T-cell responses and viral persistence in HCV-infected individuals. We postulated that HCV core protein (HCVc) directly contributes to the expansion of Tregs in HCV-infected patients, and we provide evidence to support this hypothesis in the report. Peripheral blood mononuclear cells (PBMCs) and sera were collected from 87 treatment-naïve chronic HCV-infected patients, CD4(+)CD25(+) Tregs were measured by flow cytometry, and HCV RNA and HCVc levels were detected using qPCR and enzyme-linked immunosorbent assay (ELISA), respectively. CD4(+), CD8(+), CD4(+)CD25(+) and CD4(+)CD25(-) T cells were purified from healthy donors and cultured with recombinant HCVc and Toll-like receptor (TLR) ligands. Flow cytometry was used to analyze cell proliferation, and ELISA was performed to measure cytokine production. In the 87 chronic HCV-infected patients, HCVc showed a significant correlation with HCV RNA and CD4(+)CD25(+) Tregs. Mechanistic studies showed that HCVc, together with anti-CD3 antibody, augmented CD4(+)CD25(+) Treg proliferation, but inhibited CD4(+)CD25(-) T-cell proliferation and IFN-γ production, in a dose-dependent and Treg-dependent manner. Moreover, unlike the TLR3 ligand (poly I:C) and the TLR4 ligand (lipopolysaccharide, LPS), the TLR2 ligand (lipoteichoic acid, LTA) and HCVc both inhibited TCR-induced CD4(+) T-cell proliferation and IFN-γ secretion in a Treg-dependent manner. These data indicate that HCVc, like other TLR2 ligands, triggers CD4(+)CD25(+) Treg activation and expansion to inhibit host immune responses, which may play a critical role in viral persistence in HCV-infected patients.

Treadmill exercise induces murine cardiac allograft survival and generates regulatory T cell.

Exercise therapy has been associated with improvement in functional capacity and quality of life. The role of exercise therapy in heart transplant recipients is of great interest for the transplant society, although concerning the effect of exercise therapy, there is little knowledge at present. We analyzed the effects of exercise on alloimmune responses in murine cardiac allograft transplantation. CBA mice (H2k) underwent transplantation of C57Bl/6 (H2b) hearts and exercised on a treadmill. Untreated CBA recipients rejected C57Bl/6 cardiac grafts acutely (median survival time [MST], 7 days). CBA recipients treated with treadmill for 1 week after transplantation, and for 1 week both before and after transplantation prolonged allograft survivals (MSTs, 35 and 18 days, respectively). However, treadmill exercise recipients for 1 week before transplantation were not effective to allograft survival (MST, 8 days). Adoptive transfer of whole splenocytes and CD4+ cells from treadmill exercise recipients significantly prolonged allograft survival in naive secondary recipients (MSTs, 30 and 52 days, respectively), suggesting that regulatory cells was generated after treadmill exercise. Moreover, flow cytometry studies showed that CD4+CD25+Foxp3+ cell population increased in treadmill exercise recipients. Therefore, postoperative but not pre-operative exercise could induce prolongation of survival of fully allogeneic cardiac allografts and generate CD4+CD25+Foxp3+ regulatory T cells.

Rapamycin Can Reduce Apoptosis of CD4+CD25+ Regulatory T Cells from Mouse Model of Severe Aplastic Anemia

CD4+CD25+ regulatory T cells (Tregs) are believed to control the development and progression of autoimmunity by suppressing autoreactive T cells and decreased numbers of these cells are associated with impaired immune homeostasis and development of aplastic anemia (AA). We have developed a novel mouse model of severe aplastic anemia (SAA) induced by interferon-gamma in combination with Busulphan, in which excessive apoptotic CD4+CD25+ Treg cells was found in our previous experiments. It might be one of the reasons for reduction of CD4+CD25+ Treg cells in patients with AA. Rapamycin (RAPA) is a macrolide antibiotic produced by streptomyces hygroscopicus and is considered as a potential immunosuppressant in the treatment of AA, while the effect of RAPA on apoptosis of CD4+CD25+ Tregs from patients with AA is not clear. In this study, the SAA model female mice were intraperitoneal injection with RAPA at daily dose of 0.5 mg/kg for 5 days (the RAPA-treated group, n=60), the SAA group (n=60) and the un-treated group (n=60) were as control. All mice selected to set up the SAA model developed the typical clinical and pathological patterns of SAA from day 10 post dosing. Most of them presented bleedings in association with anemia and infections. In order to explore the effect of RAPA on apoptosis of CD4+CD25+ Tregs from the novel mouse SAA model, the mononuclear cells of the peripheral blood and spleen were subjected to assess the intracellular Foxp3 expression in CD4+CD25+ Tregs by flow cytometry (FCM). In the SAA group, the FCM analysis showed down-expression of Foxp3 in CD4+CD25+ Tregs compared with the un-treated group (P<0.05). These results were in accordance with our previous observations. However, after treatment with RAPA, the expression of Foxp3 in CD4+CD25+ Tregs was increased (P<0.05). In addition, after being pured by immunomagnetic beads, the splenic CD4+CD25+ Tregs was subjected to assess apoptosis by FCM and the Akt and Stat3 phosphorylation using western blot. Compared with the un-treated group, increased CD4+CD25+ Tregs apoptosis with increased Akt phosphorylation accompanied by increased Stat3 phosphorylation was found in SAA group (P<0.05, respectively). On the contrary, RAPA-treated group exhibited CD4+CD25+ Tregs with a reduction in apoptosis rate, Akt phosphorylation and Stat3 phosphorylation compared with the SAA group (P<0.05, respectively). These results indicated that RAPA may increase the expression of Foxp3 by down-regulation the levels of Akt and Stat3 phosphorylation in splenic CD4+CD25+ Tregs from the mice model of SAA, through which might RAPA reduce apoptosis of the CD4+CD25+ Tregs from this model. DisclosuresNo relevant conflicts of interest to declare.

Differential requirement for IL-2 and IL-15 during bifurcated development of thymic regulatory T cells.

The developmental pathways of regulatory T cells (T(reg)) generation in the thymus are not fully understood. In this study, we reconstituted thymic development of Zap70-deficient thymocytes with a tetracycline-inducible Zap70 transgene to allow temporal dissection of T(reg) development. We find that T(reg) develop with distinctive kinetics, first appearing by day 4 among CD4 single-positive (SP) thymocytes. Accepted models of CD25(+)Foxp3(+) T(reg) selection suggest development via CD25(+)Foxp3(-) CD4 SP precursors. In contrast, our kinetic analysis revealed the presence of abundant CD25(-)Foxp3(+) cells that are highly efficient at maturing to CD25(+)Foxp3(+) cells in response to IL-2. CD25(-)Foxp3(+) cells more closely resembled mature T(reg) both with respect to kinetics of development and avidity for self-peptide MHC. These population also exhibited distinct requirements for cytokines during their development. CD25(-)Foxp3(+) cells were IL-15 dependent, whereas generation of CD25(+)Foxp3(+) specifically required IL-2. Finally, we found that IL-2 and IL-15 arose from distinct sources in vivo. IL-15 was of stromal origin, whereas IL-2 was of exclusively from hemopoetic cells that depended on intact CD4 lineage development but not either Ag-experienced or NKT cells.

Endometrial regenerative cells as a novel cell therapy attenuate experimental colitis in mice.

BackgroundEndometrial regenerative cells (ERCs) are mesenchymal-like stem cells that can be non-invasively obtained from menstrual blood and are easily grown /generated at a large scale without tumorigenesis. We previously reported that ERCs exhibit unique immunoregulatory properties in vitro, however their immunosuppressive potential in protecting the colon from colitis has not been investigated. The present study was undertaken to determine the efficacy of ERCs in mediating immunomodulatory functions against colitis.MethodsColitis was induced by 4% dextran-sulfate-sodium (DSS, in drinking water) in BALB/c mice for 7 days. ERCs were cultured from healthy female menstrual blood, and injected (1 million/mouse/day, i.v.) into mice on days 2, 5, and 8 following colitis induction. Colonic and splenic tissues were collected on day 14 post-DSS-induction. Clinical signs, disease activity index (DAI), pathological and immunohistological changes, cytokine profiles and cell populations were evaluated.ResultsDSS-induced mice in untreated group developed severe colitis, characterized by body-weight loss, bloody stool, diarrhea, mucosal ulceration and colon shortening, as well as pathological changes of intra-colon cell infiltrations of neutrophils and Mac-1 positive cells. Notably, ERCs attenuated colitis with significantly reduced DAI, decreased levels of intra-colon IL-2 and TNF-α, but increased expressions of IL-4 and IL-10. Compared with those of untreated colitis mice, splenic dendritic cells isolated from ERC-treated mice exhibited significantly decreased MHC-II expression. ERC-treated mice also demonstrated much less CD3+CD25+ active T cell and CD3+CD8+ T cell population and significantly higher level of CD4+CD25+Foxp3+ Treg cells.ConclusionsThis study demonstrated novel anti-inflammatory and immunosuppressive effects of ERCs in attenuating colitis in mice, and suggested that the unique features of ERCs make them a promising therapeutic tool for the treatment of ulcerative colitis.

Chaperoned amyloid proteins for immune manipulation: α-Synuclein/Hsp70 shifts immunity toward a modulatory phenotype.

α-Synuclein (αSyn) is a 140-residue amyloid-forming protein whose aggregation is linked to Parkinson's disease (PD). It has also been found to play a critical role in the immune imbalance that accompanies disease progression, a characteristic that has prompted the search for an effective αSyn-based immunotherapy. In this study, we have simultaneously exploited two important features of certain heat-shock proteins (HSPs): their classical “chaperone” activities and their recently discovered and diverse “immunoactive” properties. In particular, we have explored the immune response elicited by immunization of C57BL/6 mice with an αSyn/Hsp70 protein combination in the absence of added adjuvant. Our results show differential effects for mice immunized with the αSyn/Hsp70 complex, including a restrained αSyn-specific (IgM and IgG) humoral response as well as minimized alterations in the Treg (CD4+CD25+Foxp3+) and Teff (CD4+Foxp3−) cell populations, as opposed to significant changes in mice immunized with αSyn and Hsp70 alone. Furthermore, in vitro-stimulated splenocytes from immunized mice showed the lowest relative response against αSyn challenge for the “αSyn/Hsp70” experimental group as measured by IFN-γ and IL-17 secretion, and higher IL-10 levels when stimulated with LPS. Finally, serum levels of Th1-cytokine IFN-γ and immunomodulatory IL-10 indicated a unique shift toward an immunomodulatory/immunoprotective phenotype in mice immunized with the αSyn/Hsp70 complex. Overall, we propose the use of functional “HSP-chaperoned amyloid/aggregating proteins” generated with appropriate HSP-substrate protein combinations, such as the αSyn/Hsp70 complex, as a novel strategy for immune-based intervention against synucleinopathies and other amyloid or “misfolding” neurodegenerative disorders.

Administration of Low Doses of IL-2 Combined to Rapamycin Promotes Allogeneic Skin Graft Survival in Mice

Human CD4(+) CD25(+) FoxP3(+) regulatory T cells (Tregs) prevent allogeneic graft rejection by inhibiting T cell activation, as has been shown in mouse models. Recently, low-dose IL-2 administration was shown to specifically activate Tregs but not pathogenic conventional T cells, leading to resolution of type 1 diabetes in nonobese diabetic mice. We therefore tested the ability of low-dose IL-2 to prevent allogeneic skin graft rejection. We found that while IL-2 alone was inefficient in preventing rejection, combined with rapamycin, IL-2 treatment promoted skin graft survival both in minor disparate and semi-allogeneic skin graft combinations. Tregs are activated by this combined treatment while conventional CD4(+) cell expansion and activation are markedly inhibited. Co-administration of anti-CD25 antibodies dramatically reduces the effect of the IL-2/rapamycin treatment, strongly supporting a central role for Treg activation. Thus, we provide the first preclinical data showing that low-dose IL-2 combined with rapamycin can significantly delay transplant rejection in mice. These findings may form the rational for clinical evaluation of this novel approach for the prevention of transplant rejection.

The immune pathogenesis of immune reconstitution inflammatory syndrome associated with highly active antiretroviral therapy in AIDS.

The present study investigated the immunological pathogenesis of immune reconstitution inflammatory syndrome (IRIS) in acquired immunodeficiency syndrome (AIDS) patients undergoing highly active antiretroviral therapy (HAART). A total of 238 patients with AIDS who received initial HAART were included in this prospective cohort study. Blood samples were collected immediately, at baseline, at week 12, and at week 24 after initial HAART and at the onset of IRIS. Lymphocyte subsets, Th1 and Th2 cytokines, and interleukin (IL)-7 levels were measured by flow cytometry or ELISA. Among the 238 patients with AIDS who received HAART, 47 patients developed IRIS. The percentages of CD4(+) and CD8(+) naive, memory, and activated cells exhibited no significant differences between AIDS patients with and without IRIS 24 weeks after initial HAART. The percentage of CD4(+)CD25(+)Foxp3(+) regulatory T cells was lower in IRIS patients than in non-IRIS patients before HAART, 12 weeks after HAART, 24 weeks after HAART, and at the onset of IRIS. IL-2 and interferon (IFN)-γ levels were significantly higher at week 4 and at the onset of IRIS in IRIS patients than in non-IRIS patients. In contrast, IL-4 and IL-10 levels were significantly lower at week 4 and at the onset of IRIS in IRIS patients than in non-IRIS patients. Plasma IL-7 decreased gradually with the progression of HAART. The level of IL-7 was higher in IRIS patients than in non-IRIS patients at all follow-up time points. An imbalance of Th1/Th2 cytokines, a consistently low CD(+)CD25(+)Fox3(+) percentage, and a high IL-7 level may be crucial in the pathogenesis of IRIS in AIDS patients who had received HAART.