8 The use of herbal medicine has increased in developed countries.9 Alternative remedies are perceived to be innocuous and may provide placebo effects from the rituals associated with their ingestion.10 Use of herbal medicines increased in the USA by 25% between 1990 and 1997.11 Approximately 10% of US adults were using herbal remedies in 1999. Approximately $US 4.2 out of the $US 17.8 billion spent on ‘dietary supplements’ in 2001 were for herbs and other botanical remedies.12 Approximately $US 5 billion worth of over-the-counter Small Molecule Compound Library herbal medicines were sold in the European countries in 2003.13 Herbal medicine accounted for approximately

26% of all alternative and complimentary medicine use in Australia.14 The global annual turnover in herbal medicines is estimated at $US60 billion, representing approximately 20% of the overall drug market.15 The nephrotoxic potential of herbal remedies is being increasingly recognized.3,16,17 Causality is suspected on the basis of a temporal association between the intake of an agent and the injury. This is easier to establish in the case of acute toxicity where the interval between intake and presentation is short and the history of use of the offending agent is easy to recall, but harder in chronic

diseases that progress slowly. Remote exposure may be forgotten or even denied for fear of social stigmatization. Herbal toxicity can develop in any of the following situations:3,18,19 selleck chemicals (i) consumption of a herb with unknown toxicity; (ii) incorrect identification leading to substitution of an innocuous herb with a toxic one; (iii) deliberate or inadvertent contamination with nephrotoxic non-herbal drugs (e.g. non-steroidal anti-inflammatory agents), pesticides or chemicals (e.g. heavy metal contamination from soil or water); (iv) potentiation of the toxic effect

of a conventional drug due to interaction with a compound Methisazone present in the herb; and (v) consumption of meat from an animal that has grazed on toxic plants (e.g. hemlock). The kidney is the route of excretion of most of the substances present in the herbs. The high blood flow rate and large endothelial surface area of the kidneys ensures delivery of large amounts of toxin to the renal parenchyma. High concentrations may be reached in the medulla because of active tubular transport, especially during a state of fluid deprivation. Renal involvement associated with the use of traditional medicinal products can take several forms,16–18 including acute kidney injury, tubular function defects, dyselectrolytaemias, systemic hypertension, chronic kidney disease (CKD), renal papillary necrosis, urolithiasis and urothelial cancer.

The study was approved by local Ethics Committees and informed consent was obtained from the donors. In the following sections references

are given to papers which have used the same samples for other purposes. A recombinant fragment of MASP-1 comprising the CCP1-CCP2-SP domains (rCCP1-CCP2-SP) was produced in Escherichia coli, and refolded and purified as described previously [13]. A synthetic peptide representing the 15 C-terminal amino acid residues of human MASP-1 (CHHNKDWIQRVTGVR) was coupled to keyhole limpet haemocyanin. Three Wistar rats were immunized four times subcutaneously with 10 µg of this conjugate, emulsified first in complete Freund’s adjuvant and then in incomplete Freund’s adjuvant for boosts. Sera from the animals were tested for reactivity towards Protease Inhibitor Library order rCCP1-CCP2-SP coated onto microtitre wells. All rats responded and the rat with the highest titre was selected. IgG was purified from the anti-serum by affinity chromatography on Protein G-coupled beads. The serum was diluted 1/1 in phosphate-buffered saline (PBS; 137 mM NaCl, 2·7 mM KCl, 1·5 mM KH2PO4, 8·1 mM Na2HPO4, pH 7·4) selleck compound containing 10 mM EDTA (PBS/EDTA), and passed through the beads. After washing, the

bound IgG was eluted with 0·1 M glycine, pH 2·4. The immunoglobulin concentration was determined by spectroscopy at 280 nm. The purified IgG was biotinylated by standard procedure [26] using 167 µg biotin-N-hydroxysuccinimide ester (Sigma-Aldrich, Vildagliptin St Louis, MO, USA) per mg antibody. The anti-MASP-1 anti-serum was tested by Western blotting. MBL/MASP complexes were purified from serum by affinity chromatography on mannan coupled to Sepharose beads, as described previously [8]. The complexes, as well as a preparation of rCCP1-CCP2-SP, were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) in non-reducing conditions followed by blotting onto a membrane. For the preparation of Western blotting strips, MBL/MASP

complexes corresponding to 30 µg MBL were loaded onto a single-well XT-Criterion pre-cast 4–12% gradient Bis-Tris polyacrylamide gel (Bio-Rad, Copenhagen, Denmark) and cut into 2·5-mm-wide strips after blotting, resulting in approximately 1 µg MBL (+ associated proteins) per strip. The proteins were blotted onto a nitrocellulose membrane (Hybond-ECN; GE Healthcare, Hilleroed, Denmark) in transfer buffer (25 mM Tris, 0·192 M glycine, 20% v/v ethanol, 0·1% w/v SDS, pH 8·3) for 500 volt-hours. The membrane was blocked in 0·1% Tween 20 in Tris-buffered saline (TBS) (10 mM Tris–HCl, 140 mM NaCl, 1·5 mM NaN3, pH 7·4) before being cut into strips. The strips were incubated with primary antibodies (normal rat IgG or rat anti-MASP-1 antibody) diluted in primary buffer (TBS with 0·05% Tween 20 (TBS/Tw), 1 mM EDTA, with 1 mg human serum albumin (HAS) and 100 µg normal human IgG (hIgG) added per ml) in eight-well trays (Octaline, Pateof, Denmark) for 2·5 h on a rocking table.

Genetic analysis of various TB proteins has confirmed that MPB64 is identical to MPT64, a protein produced by M. tuberculosis. Non-tuberculous mycobacteria do not produce MPB64; it is specifically secreted by M. tuberculosis complex (17–21). MPB64 was first

isolated by Harboe and Nagai in 1986, whereas Li and colleagues identified it as a secreted protein specific to tuberculous mycobacteria in 1993 (7, 3). Hasegawa and colleagues confirmed the high sensitivity and specificity of the Capilia TB assay, which employs an anti-MPB64 monoclonal antibody to detect MPB64 protein and concluded that this assay was useful for the diagnosis of TB (8). In the present study, we MK-8669 supplier assayed urine and serum samples obtained from patients with TB in the active and healing phases by the dot-blot method to assess the profile of reactivity with MPB64 antigen. Rashid and colleagues reported that patients admitted to hospital with TB had a mean ESR 97.04 mm/hr, 57.6% being ≥100 mm/hr (22, 23). In the present study, we investigated the correlation between our dot-blot assay and ESR. In one representative patient, the ESR was around 100 mm/hr one month after commencing treatment and gradually decreased from two months. Our dot blot assays showed that both serum and urine samples paralleled the changes in ESR over time (Fig. AZD3965 nmr 4a, d, e). All patients with

active TB were positive by dot-blot assay of both serum and urine samples and all patients with a strongly positive result had active TB. Thus, a weak reaction on the dot-blot assay suggests TB and a strong reaction indicates active TB. As shown in Figure 6, analysis that included

data obtained from both TB patients and uninfected individuals revealed a strong correlation between the results obtained by dot-blot assay of urine and serum samples (n = 34, r = 0.672). Analysis of TB patients alone revealed an even stronger correlation between results obtained with urine and serum samples (n = 23, r = 0.841) (data not shown). These findings confirm that the results obtained by assay of urine samples are consistent with those for serum samples. In the present study, we evaluated NADPH-cytochrome-c2 reductase the specificity of a dot-blot test for M. tuberculosis infection by comparing data from infected and uninfected individuals and from patients with active and inactive disease. Moreover, the results obtained from urine samples are closely correlated with those obtained from serum samples. Testing of serum is currently the main method for diagnosis of TB. However, there is a need for an assay kit that allows rapid diagnosis of active TB in the field. In particular, a kit for urine testing would be desirable. Collection of urine requires less skill than does collection of blood, has a smaller risk of contamination and requires no special equipment such as centrifuges. Therefore, urine tests are suitable for mass screening.

89 Several studies have suggested that DC can be infected with HCV, but the role of HCV in DC development and function is still elusive.59,90,91 Virologically, HCV first attaches itself to the host cell surface by means of weak interactions with glycosylaminoglycans or the

low-density lipoprotein receptor. Once bound and concentrated on the cell surface, virions are able to interact with entry receptors such as CD81 and SR-BI with high affinity. The virus–receptor complex then translocates to the tight junctions where claudin and occludin act as cofactors and induce receptor-mediated endocytosis.92 Barth et al.35 used HCV-like particles (HCV-LPs) to study the interaction of HCV with human DC. The iDC exhibited an envelope-specific and saturable binding of HCV-LPs, indicating receptor-mediated DC–HCV-LP interaction. They GDC-0068 in vivo revealed that HCV-LPs were rapidly taken up by DC in a temperature-dependent manner, and C-type lectins such as mannose receptor or DC-SIGN (DC-specific intercellular adhesion molecule 3-grabbing non-integrin) were not sufficient for mediating HCV-LP binding. Lambotin et al.93 suggested that HCV cell entry factors, which are crucial for viral uptake in hepatocytes, do not support the cell culture-produced HCV (HCVcc) uptake in DC subsets.

HCVcc acquisition by DC subsets does not depend on the C-type lectin DC-SIGN, but is partially selleck kinase inhibitor mediated by HCVcc E2 protein interaction at the cell surface. To date, the mechanisms whereby HCV affects DC function remain largely elusive.55 It is possible that HCV proteins play a role in suppressing protective immunity through interactions with host immune cells, such as DC. Indeed, the HCV core protein has been reported to impair the function of DC. The HCV core protein was able to selectively inhibit TLR4-induced IL-12 production after interacting with the gC1q receptor on the surface of MDDC by activating the phosphatidyl

inositol 3-kinase pathway, leading to reduced T helper type 1 (Th1) cell development.94,95 Dolganiuc et al.96 demonstrate that HCV core and NS3 proteins, but not envelope 2 proteins (E2), activate monocytes and inhibit DC differentiation in the absence of the intact virus, and induced production of the anti-inflammatory Fenbendazole cytokine IL-10 associated with elevated IL-10 and decreased IL-2 levels during T-cell proliferation. They also found that treatment-naive patients with chronic HCV infection had a reduced frequency of circulating PDC as the result of increased apoptosis and showed diminished IFN-α production after stimulation with TLR9 ligands.97 The HCV core protein reduced TLR9-triggered IFN-α and increased TNF-α and IL-10 production in peripheral blood mononuclear cells (PBMCs) but not in isolated PDC, suggesting that HCV core induces PDC defects. The addition of rTNF-α and IL-10 induced apoptosis and inhibited IFN-α production in PDC.

An important finding of our study is the presence of monoclonal gammopathy and proliferative glomerulonephritis. Recently, Nasr et al. described a novel

form of proliferative Nutlin-3a nmr glomerulonephritis associated with monoclonal IgG deposits (PGNMID) characterized by diffuse proliferative, membranoproliferative, or membranous features on light microscopy and glomerular monoclonal IgG deposits restricted to a single IgG subclass and a single light-chain isotype on IF microscopy.[3] On EM, granular, non-organized deposits were detected, typically in a sub-endothelial and mesangial distribution. Thirty per cent of patients have a detectable level of circulating monoclonal protein with the same heavy- and light-chain isotypes as those of the glomerular deposits. Over 40 additional patients Selleck GSK2126458 with PGNMID in the native kidney have been reported by

other groups.[4-9] The present case may be similar to those discussed in these studies, except for the presence of mesangial and segmental endocapillary proliferation secondary to monoclonal IgA2 λ light-chain deposition. Although the existence of underlying lymphoplasmacytic disorders remains to be determined by bone marrow biopsy, we believe that the capillary wall deposition of other monoclonal Igs, including monoclonal IgA, can result in a proliferative glomerulonephritis pattern of injury. Recurrent glomerular diseases usually develop early post transplantation, whereas de novo glomerular diseases usually develop several years after kidney transplantation. Furthermore, the possible development of recurrent or de novo PGNMID after kidney transplantation has been reported.[10-12] Whether the present case represents recurrent or de novo glomerulonephritis in terms of IgA2-λ monoclonality remains to be determined, and we lack the native kidney biopsy material to prove the similarity of the morphological features and the presence of

monoclonal deposits. However, because the patient had obvious IgA2 mesangial and glomerular capillary deposits 1 year post transplantation, it is likely that the clinical history was consistent with recurrent disease. The initial three allograft biopsies performed without immunostaining for anti-light chain antibodies showed recurrent IgAN. Because of the lack of proven effective therapeutic approaches for recurrent IgAN,[1, 13] we treated see more the patient with rituximab, which has been shown to be effective in treating patients with nephrotic syndrome.[14, 15] However, the treatment failed to improve renal function. A recent small trial conducted by Sugiura et al. in adults with IgAN found no benefit of rituximab for the reduction of proteinuria at 6 months, although the dose of steroids was reduced.[16] The optimum dose of rituximab is also unknown, although prescribing the minimal dose needed to achieve B-cell depletion may be as clinically effective and cost-effective as conventionally prescribed doses.

MSC-mediated immunomodulation requires both cell–cell contact and release of soluble factors, although there is great controversy concerning the molecules involved both in the direct immunosuppressive effect of MSCs and in Treg induction [20].

Many possible candidates are currently under investigation, including transforming growth factor (TGF)-β and interleukin (IL)-6 [21]. It is well known that TGF-β is involved in MSC immunosuppression via a significant increase of its production NVP-AUY922 [22-24]; as far as IL-6 is concerned, it has been proposed that its increased production is associated directly with ageing [25], and probably playing a role in triggering the immunosuppressive effect of MSCs [26]. Furthermore, a recent report suggests that, although the number of natural Tregs is increased significantly during SSc, an impairment

in their ability to suppress PF-02341066 cost CD4+ effector T cells has been shown and their defective function correlates strongly with lower expression of surface CD69 [27]. Taken together, these few data do not address completely the immunoregulatory status during SSc, and might suggest a possible defect in effector cell immunosuppression. In this paper we have gained insight into the multi-step immunosuppressive function of MSCs in SSc, permitting these cells, although senescent, to save their specific ability by exploring some pathways involved in this function, with a special interest in IL-6 and TGF-β production, which are considered pivotal cytokines in the pathology of SSc, and finally addressing the potential role of SSC–MSC in generating inducible Tregs. After ethics committee approval and written informed consent (Helsinki

Declaration), human MSCs were obtained by aspiration from the iliac crest from 10 SSc patients (four with diffuse and six with a limited form of the disease) and 10 healthy bone marrow (BM) donors [nine women and one man; mean age 35 years (age range 23–45 years)] undergoing BM harvest. The demographic features of our SSc patients are shown in Table 1. Due to the possible effects of immunosuppressive and cytotoxic agents on MSCs, SSc patients treated with high TCL doses of both corticosteroids and cyclophosphamide were not included into this study. Samples were placed into tubes containing ethylenediamine tetraacetic acid (EDTA) and the BM cells were obtained by density gradient sedimentation on 12% hydroxyethyl amide. The upper phase was harvested, centrifuged at 700 g for 10 min and plated at a concentration of 5 × 103 cells/cm2 in Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (FBS; Gibco), 2 mmol/l L-glutamine (EuroClone, Milan, Italy) and 100 U penicillin, 1000 U streptomycin (Biochrom AG, Berlin, Germany).

1). Splenic lymphocytes from mice

immunized with AMH formulated Epacadostat molecular weight with adjuvants DDA and BCG PSN secreted high levels of IFN-γ upon stimulation with Ag85B, HspX, Mpt64190–198 and PPD (Fig. 2). Splenic lymphocytes from mice immunized with AMH produced higher level of IFN-γ than those immunized with Ag85B, AMM, BCG and PBS with the stimulation of HspX, Mpt64190–198 and PPD. When stimulated with antigen Ag85B, the level of IFN-γ induced by AMH vaccine was lower than that by AMM (P < 0.05) and Ag85B (P > 0.05) vaccines, but was still higher than that receiving BCG (P < 0.05). With the aid of adjuvant DDA + BCG PSN, AMH induced higher levels of antigen-specific IgG1 and IgG2a than Ag85B and AMM (Table 1). Ag85B-specific IgG2a and HspX-specific IgG1 and IgG2a from AMH group were the highest among all groups. PPD-specific IgG1 and IgG2a from the mice immunized with AMH were higher than Ag85B and BCG group. The ratio of Ag85B-specific IgG2a/IgG1 from AMH group was lower than that of BCG group but higher than that of AMM and Ag85B groups. The ratio of HspX-specific IgG2a/IgG1 from AMH group was the highest among all groups. High IgG2a/IgG1 ratio reflects Th1 activity which produces IFN-γ to promote intracellular killing activity by activating

macrophages and cytotoxic T cells [17]. Cell-mediated immune responses in mice primed with BCG and boosted by AMH, AMM, or AMM + AMH were analysed with the stimulation of Ag85B and PPD. The results showed that there were higher levels of IFN-γ C-X-C chemokine receptor type 7 (CXCR-7) production MK0683 clinical trial in mice boosted with AMH, AMM and AMM + AMH vaccines than the group of BCG (Fig. 3). It

indicated that Ag85B-, PPD-specific cell-mediated immunity were highly induced by AMH, AMM and AMM + AMH boosting. Unlike the fusion proteins, single-protein Ag85B boosting did not significantly induce high cell-mediated immunity compared with BCG alone. There was no significant difference among AMM, AMH and AMM + AMH groups. The boost with subunit vaccines induced a higher humoral immune response against Ag85B (data not shown). PBS control did not produce antibodies. The titres of IgG1 and IgG2a against Ag85B from mice immunized with BCG and boosted with subunit vaccines were higher than that primed with BCG alone (P < 0.05), whereas there were no significant differences among boosting groups. Protective efficacy was evaluated by CFU count in mice boosted with different protein vaccines followed by challenging with virulent M. tuberculosis H37Rv. The CFUs from the lungs of mice boosted with the subunit vaccines AMM + AMH and AMM were significantly lower than PBS injection, although AMH subunit vaccine boosting did not lead to a significant decrease in CFUs. The bacilli were effectively inhibited in the lungs of mice boosted by AMM + AMH in DDA-BCG PSN, which even induced significantly lower CFU than BCG group (P < 0.05) (Fig. 4).

Then sequential treatments of these prepared JAWS II iDCs and examination of them were performed as described in the Results section. The effects on DCs of chemokine pre-treatment followed by LPS stimulus (to initiate

maturation) were assessed by measuring levels of endocytic ability. To quantify endocytic ability, DCs collected on Day 1 (24 hr after no treatment or the described chemokine treatment) and on Day 2 (24 hr after subsequent LPS treatment) were resuspended in medium (without phenol red) at 1 × 106 cells/ml. Then, each sample received 3·33 μg/ml fluorescent Alexa Fluor 488-ovalbumin (OVA) (a model antigen) (Invitrogen) for 30 min at 37°. After incubation, selleck inhibitor any excess fluorochrome bound to the cell surface see more was

quenched for 3–4 min on ice using a 0·5% Trypan Blue/2% FBS/1× PBS solution. After two repetitive quenching steps, cells were thoroughly washed using ice-cold FACS buffer (2% FBS/1× PBS) and then immediately examined using a FACS Canto (BD Biosciences, San Jose, CA). Negative control DCs were separately prepared by incubation of DCs with the model antigen on ice. The mean fluorescence intensity (MFI) of the ice control cells was subtracted from that of cells incubated at 37° with OVA per treatment or control. Data were analysed using the FlowJo Software (Tree Star Inc., Ashland, OR). The model antigen (OVA) degradation (processing) by DCs was also examined using flow cytometry. Here, DCs were treated with BODIPY-conjugated DQ-OVA (Molecular Probes/Invitrogen), IMP dehydrogenase a self-quenched conjugate of OVA that exhibits

bright green fluorescence only upon proteolytic cleavage releasing the dye molecule from the OVA. To quantify antigen degradation kinetics, this assay was carried out at 30 min, 1 hr and 2 hr after OVA incubation. DQ-OVA was applied at the concentration identical to the OVA of the antigen uptake assay above. Briefly, after DCs were collected on Day 1 and Day 2, DCs from control or sample wells were divided into three groups and resuspended in medium (without phenol red) at 1 × 106 cells/ml per group. Then, each group was incubated with 3·33 μg/ml of DQ-OVA for 30 min, 1 hr, or 2 hr at 37°. After each time-point, cells were extensively washed using PBS and then fixed with 2% paraformaldehyde (diluted from Cytofix; BD Pharmingen, San Jose, CA) for 10 min at room temperature. Fixed cells were washed twice using ice-cold FACS buffer, then examined using a FACS Canto (BD Biosciences).

However,

unlike NFAT and AP-1 factors that interact and collaborate in binding to DNA, NFAT, and NF-κB seem neither to interact nor to collaborate. We show here that NF-κB1/p50 and c-Rel, the most prominent NF-κB proteins in BCR-induced splenic B cells, control the induction of NFATc1/αA, a prominent short NFATc1 isoform. In part, this is mediated through two composite κB/NFAT-binding sites in the inducible Nfatc1 P1 promoter that directs the induction of NFATc1/αA by BCR signals. In concert with coreceptor signals that induce NF-κB factors, BCR signaling induces a persistent generation of NFATc1/αA. These data suggest a tight connection between NFATc1 and NF-κB induction in B lymphocytes contributing to the effector function of peripheral B cells. “
“Ficolins are soluble molecules of the innate immune system that recognize carbohydrate molecules on microbial pathogens, apoptotic and necrotic

HIF-1 pathway cells. They act through two distinct routes: initiating the lectin pathway of complement activation and mediating a primitive opsonophagocytosis. In this study, we measured plasma levels of ficolin-2 and ficolin-3 in 60 pre-eclamptic patients, 60 healthy LY2606368 price pregnant women and 59 healthy non-pregnant women by enzyme-linked immunosorbent assay (ELISA). Circulating levels of complement activation products (C4d, C3a, SC5b9), angiogenic factors (soluble fms-like tyrosine kinase-1, placental growth factor) and markers of endothelial activation (von Willebrand factor antigen), endothelial injury (fibronectin) and trophoblast debris (cell-free fetal DNA) were also determined. Plasma levels Cyclin-dependent kinase 3 of ficolin-2 were significantly lower in healthy pregnant than in healthy non-pregnant women, while ficolin-3 levels did not differ significantly between the two groups. Furthermore, pre-eclamptic patients had significantly lower ficolin-2 and ficolin-3 concentrations than healthy non-pregnant and pregnant women. In the pre-eclamptic group,

plasma ficolin-2 levels showed a significant positive correlation with serum placental growth factor (PlGF) concentrations and significant inverse correlations with serum levels of soluble fms-like tyrosine kinase-1 (sFlt-1), blood urea nitrogen and creatinine, serum lactate dehydrogenase activities, as well as with plasma VWF:antigen, fibronectin and cell-free fetal DNA concentrations. In conclusion, circulating levels of ficolin-2 are decreased in the third trimester of normal pregnancy. There is a further decrease in plasma ficolin-2 concentrations in pre-eclampsia, which might contribute to the development of the maternal syndrome of the disease through impaired removal of the trophoblast-derived material released into the maternal circulation by the hypoxic and oxidatively stressed pre-eclamptic placenta.

In contrast to colonic IFN-γ release, caecal IFN-γ was maximal at day 7 (Fig. 1). No significant changes in cytokine production were

noted in small intestinal tissues (data not shown). The results shown are derived from experiments with 129/SvEv mice; however, results indistinguishable from these were also produced with Swiss Webster mice. The imbalance in intestinal GSK-3 activation cytokine release with a maximal production of proinflammatory cytokines prior to production of anti-inflammatory cytokines was associated subsequently with a transient intestinal histopathological injury at day 7 post-faecal slurry exposure (Fig. 2a). The increase in intestinal injury scores was seen in both colonic and caecal tissues and involved mainly an influx in lamina propria mononuclear cells (Fig. 2b). However, not all mice developed colonic or caecal injury; the injury score among individual mice ranged from 1 to 8 in colon and from 1 to 7 in the caecum. ABT-199 purchase Higher scores were found primarily among the Swiss Webster

mice, whereas 129/SvEv mice scored generally lower. However, even those mice that were found to be microscopic disease-limited (i.e. histopathological injury score of 1 at day 7) demonstrated increased proinflammatory mucosal cytokine production. Colonic and caecal injury had subsided in most mice by day 14 (Fig. 2) and returned to base levels by day 28 (data not shown). Colonic epithelial permeability was not altered significantly in these mice when tested at days 3, 7 and 14 post-faecal slurry exposure. In fact, we observed a slight reduction in mannitol flux in colonic tissue when subjected to Ussing chamber analysis (Fig. 3). Thus, despite the temporary cytokine imbalance and brief inflammatory response in the large bowel, the intestinal epithelial barrier function appeared to be intact. To investigate systemic immune responses to ingestion of faecal slurry in these

axenic mice we assessed cytokine release in unseparated splenocytes stimulated with faecal lysates derived from specific pathogen-free (SPF)-raised mice. Maximal release of IFN-γ, IL-17 and IL-10 was measured at day 7 post-bacterial treatments (Fig. 4a, shaded bars). No increase in either TNF-α or IL-4 production Oxymatrine was noted in any of these antigen-stimulated spleen cell cultures. As expected, cytokine release following spleen cell stimulation with lysates from axenic mice that are devoid of bacterial components remained at baseline level (Fig. 4a, solid bars). Consistent with these results from stimulation with faecal lysates, we observed a similar increase in production of IFN-γ and IL-10 at day 7 in cultures stimulated with sonicates derived from pure cultures of three endogenous bacterial strains: Bacteroides vulgatus, Enterobacter cloacae and Lactobacillus reuteri (Fig. 4b).