Here, we have developed and characterized a cytotoxic LAG-3 chimeric antibody (chimeric A9H12), and evaluated its potential as a selective therapeutic depleting agent in a non-human primate model of delayed-type hypersensitivity (DTH). Chimeric A9H12 showed

a high affinity to its antigen and depleted both cytomegalovirus (CMV)-activated CD4+ and CD8+ human T lymphocytes in vitro. In vivo, a single intravenous injection at either 1 or 0·1 mg/kg was sufficient to deplete LAG-3+-activated T cells in lymph nodes and to prevent the T helper type 1 (Th1)-driven skin inflammation mTOR inhibitor in a tuberculin-induced DTH model in baboons. T lymphocyte and macrophage infiltration into the skin was also reduced. The in vivo effect was long-lasting, as several weeks to months were required after injection to restore a positive reaction after antigen challenge. Our data confirm that LAG-3 is a promising therapeutic target for depleting antibodies that might lead to higher therapeutic indexes compared to traditional immunosuppressive agents in autoimmune diseases and transplantation. Selectively inhibiting or deleting activated T lymphocytes represents a promising therapeutic approach as an alternative to current immunosuppressive treatments in autoimmunity and transplantation. One strategy might be the use of depleting antibodies that target specific antigens on activated T cells. This provides a competitive

advantage of targeting only pathogeneic T cells that are specific for auto- or alloantigens without modifying Napabucasin cost the protective immunity directed against third-party antigens [1]. The proof of concept for selective depletion of pathogeneic T lymphocytes has been demonstrated in an engineered mouse model, whereby their T cells express a viral thymidine kinase suicide gene that metabolizes the non-toxic prodrug ganciclovir into a metabolite that is toxic only to dividing cells. The result was a significant delay in the rejection of skin and heart grafts and the induction of an immune tolerance in a fraction of the recipient mice [2]. However,

the Dynein therapeutic translation of this strategy requires the targeting of an antigen that is highly specific for activated T cells. So far, few molecules that are expressed selectively by activated T cells have been identified. Among these are CD25, CD152, CD154 and CD223 (lymphocyte-activation gene-3; LAG-3[3]). LAG-3 is an important regulator of T cell homeostasis [4] that is related evolutionarily to CD4 and, like CD4, is associated with the T cell receptor. It has retained an affinity 2 logs higher than CD4 for their common ligand, major histocompatibility complex (MHC) class II. LAG-3 is a transmembrane protein that forms dimers at the surface of both CD4+ and CD8+ T lymphocytes [3,5] residing in inflamed secondary lymphoid organs or tissues (i.e. human tumours or rejected allograft), but not in spleen, thymus or blood.

The P. gingivalis -induced production of IL-6 was approximately 2.5-fold higher in patients with GAgP than in healthy controls (P < 0.05), while the corresponding TNF-α production was non-significantly elevated. IL-1β production induced by P. gingivalis, as all cytokine responses induced Pritelivir order by Pr. intermedia, F. nucleatum and TT was similar in the two groups. A reduced IL-12p70 response to Pr. intermedia and F. nucleatum was observed in smokers compared to non-smoking patients (P < 0.02). To assess the role of serum factors in the elevated IL-6 response

to P. gingivalis, MNC from two donors free of disease were stimulated with this bacterium in the presence of the various patient and control sera. An elevated IL-6 and TNF-α response was observed in the presence of patient sera (P < 0.01 and P < 0.04, learn more respectively). The data suggest that an exaggerated production of IL-6 occurs in GAgP, and that pro-inflammatory serum factors play an essential

role in the response. Periodontitis is a widespread destructive inflammatory process affecting the tooth-supporting tissues including gingiva, cementum, alveolar bone and periodontal ligament. An estimated 65% of Scandinavian adults have some form of periodontitis [1]. Untreated, the irreversible destructive process may ultimately result in tooth loss. Inflammation in the peridontium is initiated by bacteria on the surface of the teeth. A pathogen believed to be strongly associated with periodontitis is Porphyromonas gingivalis (P. gingivalis) [2], and this microorganism is also thought to be a key pathogen in the

established relationship between periodontal infection and cardiovascular disease [3]. Periodontitis varies in disease susceptibility and intensity, the Ribonucleotide reductase most severe form being the rapidly progressing generalized aggressive periodontitis (GAgP). The tissue damage observed in GAgP often does not correlate with the amount of bacterial accumulations on the tooth surface [4], which suggests that individual characteristics of the patients’ immune response play a major role in determining the development and severity of periodontitis [5]. The individual differences may be caused by processes involving both the innate and the adaptive immune system [6]. Thus, periodontal inflammation is a double-edged sword: On the one edge, the inflammatory response combats the invading bacteria; on the other edge the production of inflammatory mediators may lead to irreversible destruction of tooth-supporting tissues [7]. Interleukin (IL)-1β, IL-6 and tumour necrosis factor (TNF)-α are considered the most important pro-inflammatory cytokines involved in the destructive processes [8].

16 An alternative approach to expansion of nTregs in vitro may be to use biological therapies such as anti-tumour necrosis factor-α antibodies so as to maximize the function of nTregs in vivo.9,50 The development

of iTregs for clinical applications might provide a superior alternative in IBD. In mouse models, iTregs are known to prevent T-cell driven colitis,37 and it may be easier to generate cells specific for relevant antigens using this approach. In addition to differentiation using compounds such as TGF-β and rapamycin,51 iTregs can be generated when naive T cells are stimulated in vitro by tolerogenic dendritic Selleck Sorafenib cells, which are from the intestine and BAY 80-6946 in vitro induce antigen-specific FoxP3+ Tregs in a TGF-β and retinoic acid dependent manner.52–55 A slight variation on this strategy would be to use vitamin A or its derivative, retinoic acid, to directly enhance tolerance and the generation of iTregs in the intestine in vivo.21,56 Antigens could also be targeted to tolerogenic intestinal dendritic cells in vivo using a single-chain antibody specific for unique cell surface makers as a delivery system.57 This latter strategy is thought to mimic the natural process of oral tolerance where antigens are presented by tolerogenic dendritic cells58 and

so may generate more effective and stable populations of antigen-specific iTregs in comparison with in Edoxaban vitro-derived cells. In addition to FoxP3+ Tregs, Tr1 cells are also candidates for cellular therapy in mucosal diseases. The intestinal environment naturally relies on IL-10 for the maintenance of immune homeostasis; in mouse models, IL-10 secretion by myeloid intestinal cells is required to maintain Treg

suppressive capacity,59 and Tregs themselves must secrete IL-10 to prevent colitis.18,34,35 In a therapeutic setting, subcutaneous delivery of human recombinant IL-10 produced disappointing clinical results, but this was probably the result of protein degradation and a suboptimal route of delivery.7,60 An alternative strategy, delivering IL-10 to the target environment using genetically modified bacteria, is currently being tested in humans.61 Tr1-mediated delivery of IL-10, however, should offer a therapeutic advantage over direct protein delivery because of the possibility of delivering antigen-specific suppression. Following studies in mice showing that ovalbumin (OVA)-specific Tr1 cells prevent colitis following transfer of polyclonal T cells, a Phase I/II clinical trial was initiated to test if OVA-specific Tr1 cell clones could also treat refractory Crohn’s disease.

8%) (Fig. 3A, Supporting Information INCB024360 Fig. 2A) and BAL (44.5%) (data not shown) in 4/5 A7 transgenics. Earlier experiments established that a “public” TCRβ repertoire was consistently detected in wt B6 DbNPVβ8.3+CD8+ populations 14, 15. Loss of this Vβ8.3

bias in the DbNPCD8+ T cells from A7 mice suggests that either the public DbNPVβ8.3+ clonotypes require pairing with one or more DbNP366-specific Vαs for optimal recognition, or that there are structural constrains that limit pairing with the KbOVA257-specific Vα2 chain. However, this did not reflect any overall incapacity of naïve Vβ8.3+CD8+ T cells to pair with at least some Vα2+ TCR, as analysis of naïve CD8+ T cells in B6 mice showed that an average of 8.26±0.35% of Vβ8.3 CD8+ T cells are Vα2+ (Supporting Information Fig. 3). Similar to the restricted and public nature of wt DbNPVβ8.3+ T cells, CH5424802 analysis of the subdominant Vβ4+DbNPCD8+ sets in B6 mice showed a profile of restricted TCRβ repertoire diversity, with several clonotypes being found repeatedly in different individuals 24. Is this also the case for DbNPCD8+ T cells generated from the A7 transgenics? Analysis of 237 CDR3β sequences from seven mice showed that an average of 2.14±1.46 DbNP366-specific Vβ4 clonotypes were detected per mouse (Table 1). Sequencing established a profile of

predominant Jβ1S6 usage, a long CDR3β loop of 12 aa and identified two clonotypes (Table 1A) that were detected previously in the wt B6 SQDRRNSYNSPL and SQDRRSSYNSPL 24. The public DbNPVβ4+CD8+ clonotype SQDRRNSYNSPL found in all B6 mice (Table 1B) was detected in 3/7 A7 transgenics. The Vβ4+DbNPCD8+ cells elicited by influenza virus infection of A7 mice thus display broadly the same TCRβ characteristics as those from the B6 controls 24. Taken together, the Thalidomide DbNPCD8+ T cells generated in the A7 transgenics utilize TCRβ clonotypes that are also found within the subdominant Vβ4+ set from normal mice. Following influenza infection of B6 mice, the DbPACD8+ set displays a strong Vβ7 bias 13. This profile of Vβ7 preference was maintained for the DbPACD8+ T cells in A7 transgenics (Fig. 3B). Similar

to the 53.7% of wt DbPACD8+ T cells that utilize Vβ7 25, the A7 DbPACD8+ set showed preferential usage of Vβ7 (Fig. 3B, Supporting Information Fig. 2B) for those recovered from the spleen (51.2%) and BAL (71.9%, data not shown). A strong Vβ9 bias was also observed in two of the A7 mice (Supporting Information Fig. 2B), suggesting alternate pairing with the OVA257-specific Vα2 for a proportion of the DbPACD8+ T cells. Subsequent analysis of CDR3β sequences for 264 DbPAVβ7+CD8+ T cells from six A7 transgenics established that there is a pattern of limited TCRβ diversity, with an average of 5.3±3.4 clonotypes detected per mouse (Table 2) in contrast to the much broader TCRβ repertoire (20.6±3.8) characteristic of the B6 controls 13, 17. Many of the TCRβ clonotypes identified in the A7 (9/37) had been detected in the B6 mice.

4%), helpful in learning (84.2%), better than traditional MMC (94.7%). Conclusion: A structured MMC is an effective means of engaging physicians, nurses, and key administrative leaders in the discussion of adverse events or patient complications. This systems-based, problem-learning process can promote patient care and safety. RAFIQ KAZI1, Sorafenib purchase SHERAJEE

SHAMSHAD J.1, FUJISAWA YOSHIHIDE2, MOGI MASAKI3, RAHMAN ASADUR1, SUFIUN ABU1, NAKANO DAISUKE1, KOEPSELL HERMANN4, NISHIYAMA AKIRA1 1Department of Pharmacology, Faculty of Medicine, Kagawa University; 2Life Science Research Center, Faculty of Medicine, Kagawa University, Japan; 3Department of Molecular Cardiovascular Biology and Pharmacology, Graduate School of Medicine, Ehime University, Japan; 4Institute of Anatomy and Cell Biology, University of Wuerzburg, Germany Introduction: Overactivity of the sympathetic nervous system has been shown as one of the major contributors to the complex pathophysiology of hypertension, hyperinsulinemia and diabetes. Renal sympathetic denervation (RDX) improves glucose metabolism and insulin sensitivity in addition to reducing blood pressure in patients with resistant hypertension. We investigated the effects of renal sympathetic BAY 80-6946 concentration denervation at early age on the development of hypertension

and glucose metabolism in obese rats. Methods and Results: Uninephrectomized (at 5 week of age) Otsuka Long Evans Tokushima Fatty (OLETF)

and Long Evans Tokushima Otsuka (LETO) were underwent RDX at 6 week of age. RDX-LETO and -OLETF rats had almost undetectable Edoxaban kidney tissues norepinephrine (NE) levels. RDX did not affect blood pressure profiles and heart rate in pre-diabetic stage evaluated by telemetry system. RDX-OLETF rats showed markedly lowered in the blood glucose, plasma insulin levels and their area under the curve in response to oral glucose loading during the oral glucose tolerance test compared to non-denervated OLETF rats. Furthermore, the whole body insulin sensitivity was assessed by the hyperinsulinemic-euglycemic clamp study at 20 week of age, and RDX-OLETF rats showed an increased glucose infusion rate than non-denervated OLETF rats. RDX suppressed plasma and renal tissues NE levels and increased in vivo glucose uptake by adipose tissues, soleus muscle and liver tissues in OLEFT rats. Furthermore, RDX suppressed sodium dependent glucose transporter 2 (SGLT2) translocation and expression in renal proximal tubular brush border membrane as detected by immunofluorescence and western blot followed by markedly increased urinary glucose excretion in OLETF rats.

All skin flaps showed acceptable static 2-point discrimination Selleckchem SB431542 and adequate protective sensation. Patient satisfaction for resurfaced digits averaged 9 on a 10-points visual analogic scale. In conclusion, the free fasciocutaneous flaps used were thin and did not interfere with finger movements. The

patient’s finger formed a smooth contour and acceptable functional results were obtained after reconstruction. This method may be a valuable alternative for reconstruction of entire circumferential avulsion injury of the digits. © 2012 Wiley Periodicals, Inc. Microsurgery, 2013. “
“The object of this study was to compare the outcomes of the vacuum assisted closure (VAC) therapy and conventional wound care with dressing change for treatment of complex wounds in patients with replantation of amputated upper and lower extremities. Data of 43 patients with replantation of amputated extremities from May 2004 to December 2011 were reviewed. There were 18 wounds of 18 patients with replantation, which were treated by dressing change and 26 wounds of Akt inhibitor 25 patients by VAC

therapy. The outcomes were evaluated by the survival rate of replanted extremities, growth of granulation tissue, interval between wound treatment and secondary procedure and eventual secondary wound coverage methods. Vascular thromboses were found in 3 patients with wound treatment by dressing change and 5 by VAC. All replants of two groups of patients survived after salvage procedures. The wound score was 3.6 ± 0.7 in the conventional dressing change group and 5.8 ± 0.7 in the VAC group at the sixth day after treatment, respectively. The intervals between wound treatment and secondary wound coverage procedure were 12.0 ± 1.7 days in the dressing change group and 6.1 ± 0.7 days in

the VAC group. Flaps were applied for wound coverage in 9 out of 18 (50.0%) wounds in the dressing change group and 5 out of 26 (19.2%) in the VAC group (P < 0.05), when the wounds of rest of patients were covered by the skin graft. The results showed that VAC could promote the growth of granulation tissue of wound, decrease the need of flap for wound coverage, and did not change the survival of replantation. © 2013 Wiley Periodicals, Inc. Microsurgery 33:620–624, 2013. "
“The VAV2 aim of this study was to evaluate and compare the effectiveness of classical suture and sutureless repair with fibrin glue, by using or not a resorbable collagen tube, after sciatic nerve transection. Twenty-five mice were used in this study, divided in five groups. They were submitted to sciatic nerve transection and immediate repair of the nerve stumps by either direct suture or fibrin glue adhesion or by the tubulization technique in which the nerves stumps were sutured or glued to a collagen tube (experimental groups). A control group was designed as the best regeneration condition, by using a crush lesion (control group). After eight weeks, the regenerated nerves were processed for light and electron microscopy.

The utility of OCT for distinguishing NMO from MS and other inflammatory conditions with ocular involvement is currently being investigated. Visual evoked potentials show either reduced amplitudes or prolonged latencies, or both; in more severe cases there may be no response at all [262]. Delayed P100 latencies may indicate that the optic nerve is subclinically affected in

patients presenting with LETM, but with no history buy KU-57788 of clinically apparent ON. NMO is still an incurable disease. The goal of treating acute NMO events is to improve relapse symptoms and restore neurological functions; long-term immunosuppression aims to prevent further attacks [4, 263, 264]. Any treatment recommendations are limited by the small size of most studies, which were mostly retrospective case-series. No prospective controlled trials in NMO have been conducted, and most study designs with long placebo treatment would probably be considered unethical. Relapses are treated with high-dose intravenous methylprednisolone; if response is insufficient, patients may benefit from PE [265]. If a patient has previously responded well to PE, PE may be considered as initial treatment

in case of another relapse. In patients in whom both steroids and PE do not improve symptoms, treatment with intravenous immunoglobulins [266] or an escalation to cytoablative Cediranib (AZD2171) therapy such as cyclophosphamide may be considered [264]. For

long-term immunosuppression, click here patients usually receive either B cell-targeted therapies such as intravenous rituximab or oral azathioprine and/or prednisone [87, 110, 113, 267-272]. Other possible options include mycophenolate mofetil [273], methotrexate [274] or mitoxantrone which, however, is limited by major side effects such as cardiotoxicity or leukaemia and thus generally not considered as initial treatment [264, 275-280]. It is beyond the scope of this paper to provide details on dosing schemes and monitoring of the various NMO drugs, and therefore we refer the reader to two recent, excellent overviews on treatment recommendations [264, 281]. However, one aspect deserves mention: less severe lesions have been found in type I interferon (IFN) receptor-deficient mice, suggesting that type I IFNs might be involved in the pathogenesis of NMO. Accordingly, IFN-β, a therapeutic mainstay in MS, has been repeatedly reported to exacerbate disease or to be ineffective in patients with NMO. The use of IFN-β in the treatment of NMO is therefore strongly discouraged. Similarly, lack of efficacy or disease exacerbation has also been reported following treatment with other typical MS drugs such as natalizumab and, in single cases, also fingolimod and alemtuzumab [169-171, 282-290].

Disease penetrance in our NOD colony 3-MA nmr is greater than 90% in 8.3-NOD females and about 50% in males (Fig. 1a,b). Genetic ablation of the Il21 gene abrogated completely T1D incidence in female and male 8.3-NOD mice (Fig. 1a).

Strikingly, a partial reduction in IL-21 availability was sufficient to reduce T1D incidence by 50–60% in Il21+/− females expressing either the 8.3 TCR or a polyclonal TCR repertoire (Fig. 1a,c), although Il21 gene heterozygosity did not diminish T1D incidence in male 8.3-NOD mice (Fig. 1b). IL-21 deficiency completely prevented mononuclear cell infiltration of pancreatic islets in 8.3-NOD mice (Fig. 1d). These results show that the highly diabetogenic 8.3 TCR transgenic CD8+ T cells require IL-21 to induce insulitis and cause diabetes, and that a partial reduction in IL-21 availability is sufficient to attenuate their pathogenic potential. Several reports have shown that IL-21 is required for sustaining the expansion of antigen-specific T cells during chronic viral infections [27-31]. Therefore, we evaluated the ability of IL-21-deficient 8.3 T cells to proliferate in response to cognate IGRP206–214 peptide or to its mimotope NRP. As shown in Fig. 2a–c, IL-21-deficient cells showed significantly reduced proliferation to TCR ligands or to anti-CD3/CD28 cross-linking, but responded similarly to PMA and ionomycin.

These cells also showed comparable levels of proliferation to stimulatory combinations of cytokines, selleck IL-7 or IL-15 along with IL-21, although the magnitude of this response was low compared to antigen-induced proliferation (Fig. 2d). An earlier report suggested a role for IL-21 in T cell homeostasis in the NOD mouse [2]. However, we did not observe

any difference in total T cell numbers or the frequency and numbers of 8.3 T cells in 8.3-NOD.Il21−/− mice (Fig. 3a,b). To evaluate the impact of IL-21 deficiency on homeostatic expansion of CD8+ T cells, we injected CFSE-labelled splenocytes from 8.3-NOD or 8.3-NOD.Il21−/− mice into NOD.Scid or NOD.Scid.Il21−/− recipients. As shown in Fig. 3c, expansion of CD8+ T cells from IL-21-deficient or wild-type Farnesyltransferase donors was comparable in NOD.Scid and NOD.Scid.Il21−/− recipients, suggesting that IL-21 is dispensable for homeostatic expansion of CD8+ T cells. Collectively, the above results indicate that CD8+ T cells that develop in IL-21-deficient mice proliferate to a lesser extent following TCR stimulation, and that this does not arise from a general proliferation defect, as these cells undergo efficient cytokine-driven homeostatic expansion in vivo. Next we addressed the consequence of IL-21 deficiency on antigen-induced effector functions of CD8+ T cells. As shown in Fig. 4a, IL-21-deficient 8.3 T cells displayed normal antigen-specific cytolytic activity as they lysed target cells pulsed with NRP-V7 peptide efficiently (Fig. 4a). IL-21-deficient 8.

SEA possesses a different tropism for the Vβ chain of the TCR, preferentially binding to the Vβ1, 3, 10, 11 and 12 types (75). Intraperitoneal administration of SEA can reactivate MBP-induced EAE after one month of clinical remission (76). Soos et al. have shown that SEA produces new episodes of EAE when given in mice which have previously been immunized with MBP after depletion of Vβ8 cells by SEB pretreatment. As previously mentioned, the explanation relies on the types of lymphocytes that remain in place to be stimulated by SEA. This experiment revealed that it is not only Vβ8 cells that can participate in EAE pathogenesis, as was previously believed (77). To our knowledge, there has been

no study of oral administration of SEA in EAE. In any case, the variable behavior seen after administration of SEB/SEA can be explained by the affinity for certain T cells, different TCR restrictions for effector lymphocytes in different species, and differing GSK3235025 in vitro routes of administration. When administered parenterally, SEA acts as a major stimulant of the systemic lymphocyte compartment. Thus, staphylococcal enterotoxins have the opportunity to reactivate EAE, even in animals which have entered a remission period (78). Insulin is now recognized as the major auto-antigen in type 1 diabetes (79). As a consequence, a number of clinical trials have tested the possibility of producing oral tolerance to insulin,

in the hope of preventing or delaying the onset of the disease in non-diabetic relatives at high risk of diabetes. The Diabetes Prevention Trial–Type 1 showed that 7.5 Gemcitabine supplier mg of oral insulin daily did not confer a benefit when compared to placebo. In a subgroup

of this trial which included only those relatives who had tested positive on two occasions for anti-insulin autoantibodies, orally administered insulin proved to be useful in preventing the onset of diabetes, compared with placebo (80). Currently, Dapagliflozin the Pre-POINT (Primary Oral/intranasal Insulin Trial) is addressing the group of children who are at high risk of developing type 1 diabetes and who have not yet developed anti-insular autoantibodies. This trial is ongoing (81). There has been no trial in humans or animals that has tested the efficacy of SEA as an adjuvant for augmenting oral tolerance to insulin or any other peptides that function as autoantigens in type 1 diabetes. In animal models the results of SEA usage appear to be in conflict. Kawamura et al. have shown that staphylococcal enterotoxins (SEA, SEC1, SEC2, or SEC3), when injected iv into non-obese diabetic female mice at 4 and 10 weeks of age, significantly reduce the incidence of diabetes at 32 weeks compared with a saline treated group (82). The explanation, according to the authors, originates in the fact that SAs are able to stimulate a CD4+ fraction of T lymphocytes which is capable of immunoregulatory activity. Ellerman et al.

To further determine IL-22 production by naive and memory CD8+ T cells, we purified subsets of naive (CD45RA+) and memory (CD45RO+) CD8+ T cells from PBMCs and stimulated the two populations with anti-CD3 plus anti-CD28 in the presence or absence of IL-21 or IL-15. Interleukin-21 induced a large amount of IL-22 production by activated naive CD8+ T cells (Fig. 3d left graph). Anti-CD3 plus anti-CD28 induced a low level

of IL-22 and addition of IL-21 slightly increased IL-22 production by memory CD8+ T cells (Fig. 3d right graph). Naive CD8+ T cells produced selleck inhibitor IL-22 in greater amounts than memory CD8+ T cells with IL-21 stimulation. In addition, IL-15 had no effect on IL-22 production in naive CD8+ T cells but could induce IL-22 production by memory CD8+ T cells. Purified naive CD8+ T cells were labelled with CFSE and stimulated with anti-CD3 and anti-CD28 in the presence or absence of IL-21 for the indicated times. Cells were then collected for flow cytometric analysis for cell division. On day 3, both CD8+ T cells from CBMCs and CD8+ CD45RA+ T cells from PBMCs treated with IL-21 had more divisions than those cells without IL-21 treatment. On day 6, the proliferation of IL-21-treated CD8+ T cells was markedly higher than non-stimulated and anti-CD3 plus anti-CD28-stimulated https://www.selleckchem.com/products/PLX-4032.html cells (Fig. 4a). In addition, on day 3, the cell number of CD8+ T cells from CBMCs was threefold to fourfold higher in culture with IL-21 than

in culture with anti-CD3 and anti-CD28 alone (Fig. 4b). Purified CD8+ T cells from CBMCs were cultured with anti-CD3 and anti-CD28 in the presence or absence of IL-21, and the expression of IL-21R was assessed by flow cytometry. The results showed that IL-21R was expressed at a low level on resting naive CD8+ T cells. Interleukin-21 up-regulated the expression of Rapamycin purchase IL-21R following stimulation with anti-CD3 plus anti-CD28 (Fig. 5a). Moreover, stimulation of CD8+ T cells with anti-CD3 plus anti-CD28 resulted in higher levels of mean

fluorescence intensity (MFI) of IL-21R expression than untreated cells (P < 0·05). Addition of IL-21 further increased the MFI of IL-21R (Fig. 5a). We further examine the expression of granzyme B in IL-21-treated naive CD8+ T cells. The results showed that a low frequency of CD8+ T cells expressed granzyme B following anti-CD3 and anti-CD28 stimulation. Addition of IL-21 markedly enhanced granzyme B expression and IL-22+ CD8+ T cells produced granzyme B simultaneously (Fig. 5b). These findings indicate that both IL-22+ CD8+ and IL-22− CD8+ T cells contribute to the cytolytic function. Signalling through the IL-21R/γc may involve different JAK/STAT molecules in different responding cells. We therefore examined the phosphorylation of STATs in human naive CD8+ T cells following IL-21 stimulation. Stimulation of CD8+ T cells with IL-21 resulted in phosphorylation of STAT1 in more than 60% of cells and more than 30% of CD8+ T cells expressed phosphor-STAT3 and phosphor-STAT5.