There is likely a functional significance for the duplication of metabolic genes in the genome of a parasite that has to convert between developmental stages under different micro-environmental

conditions during the asexual phase of its life cycle. It has been suggested, for instance, that stage-specific expression of different isoforms of metabolic enzymes such as lactate dehydrogenase and enolase (ENO) may be reflective of the different metabolic states of tachyzoites and bradyzoites, with tachyzoites being the more metabolically active. This assertion is supported by the fact that recombinant tachyzoite-specific enolase 2 (ENO2) displays higher activity in vitro than the bradyzoite-specific ENO1 (28–30). The differential expression of isoforms with varying activity levels between the two developmental stages is therefore consistent this website with their respective metabolic requirements (28). Ferguson et al. provide an alternate view highlighting the fact that early bradyzoites are just as metabolically active as tachyzoites and that the expression of bradyzoite-specific metabolic enzymes might be a feature that is adaptive to the different growth conditions

encountered by these developmental stages, with varying resource constraints (31,32). In another genome-wide search, the complement of genes encoding enzymes involved in metabolism of amylopectin has been identified in the Toxoplasma genome (33). It Docetaxel research buy is interesting to note that some of these genes also exhibit stage-specific expression profiles. R1 protein, α-glucan phosphorylase, α-glucosidase and α-amylase, which perform catabolic functions, learn more are preferentially expressed

in bradyzoites. On the other hand, enzymes involved in synthesis such as glycogenin, glycogen synthase and branching enzyme are predominantly expressed in tachyzoites (33). This expression pattern is consistent with the observation of amylopectin accumulation and subsequent turnover during differentiation (33,34). The use of microarrays in Toxoplasma studies has allowed for genome-wide queries of gene expression patterns and other genome-wide association studies that have had a significant impact on our understanding of the parasite’s biology. The first generation of Toxoplasma microarrays was designed to be used in the study of differential gene expression between the tachyzoite and bradyzoite stages of the asexual cycle (35). This array was constructed from a bradyzoite cDNA library, which represented a minimum of 600 genes. cDNAs were spotted onto glass slides and used to probe gene transcripts isolated from tachyzoites or bradyzoites. In spite of the inherent limitation of these arrays in terms of gene coverage (600 of approximately 8000 predicted genes), they have been very useful in identifying stage-specific genes that have proven to be important in differentiation (35–37).

Fourteen days after in vitro stimulation, cells were concentrated by removing half of the culture medium from each well. Then, 100 μl of the resulting cell suspension (100 000–250 000 cells) was stained using 2 μl DR0401 tetramer loaded with the corresponding peptide pool. After incubating at 37° for 1–2 hr, 5 μl anti-CD3-FITC, anti-CD4-PerCP and anti-CD25-APC was added www.selleckchem.com/products/iwr-1-endo.html at room temperature for 10 min. The cells were washed once in 1 ml PBS and analysed for tetramer positive responses using a FACS Calibur (BD Biosciences, San Jose, CA). Tetramer-positive responses were

decoded using tetramers loaded with the corresponding individual peptides. Our criterion for positivity was distinct staining that was more than two-fold above background (set to 0·2% and subtracted), which is consistent with our previous studies. After the initial round of tetramer screening (screening peptide pools), cells from positive wells were stained using sets of five tetramers, each loaded with one individual peptide from within the corresponding peptide pool. To isolate tetramer-positive T-cell lines, T cells were sorted by gating on tetramer positive CD4+ cells (at single-cell purity) using a FACS Vantage and expanded

in a 48-well plate in the presence of 2·5 × 106 irradiated allogeneic PBMC and 2 μg/ml phytohaemagglutinin (Remel Inc., Lenexa, KS). Sixteen days after expansion, T cells were stained with tetramers to evaluate the specificity of cloned T-cell lines. For peptide-stimulated proliferation assays, T-cell lines Ribociclib in vivo were stimulated using various concentrations of peptide (0, 0·4, 2 and 10 µg/ml), adding HLA-DR0401-positive monocytes as antigen-presenting cells. For protein-stimulated proliferation assays, CD14+ monocytes were isolated and used as antigen-presenting cells. Briefly, 150 × 106 PBMC from HLA-DR0401+ donors were labelled with anti-CD14-microbeads (Miltenyi Biotec) and CD14+ monocytes were positively isolated according to the manufacturer’s

instructions. To load monocytes with GAD65 protein, bead-enriched monocytes (approximately 20 × 106) were resuspended in 200 μl T-cell medium containing 200 μg/ml recombinant GAD65 protein and incubated at 37° for 2–3 hr. These monocytes were then used as antigen-presenting cells to stimulate Montelukast Sodium tetramer-positive T-cell lines. To generate dose-dependent response curves, protein-loaded monocytes and non-loaded monocytes were irradiated (2000 rads), washed, resuspended and mixed at various ratios (e.g. 1 : 0, 1 : 4, 1 : 24 and 0 : 1). For all proliferation assays, sorted T-cell lines were seeded at 1 × 105 cells/well (triplicate wells) in round-bottom 96-well plates with an equal number of antigen-presenting cells (1 × 105 cells/well total). Forty-eight hours after stimulation, each well was pulsed for an additional 16 hr with 1 μCi [3H]thymidine (Amersham Biosciences, Piscataway, NJ).

The freed Bcl-2 presumably exerts a prosurvival function, which would enhance the efficacy of the IL-15-induced Bcl-2 increment. Being resident in the intestine epithelium, it may be beneficial for CD8αα+ iIELs to control Bim activity by phosphorylation and dephosphorylation rather than synthesis and degradation, as the former can be achieved in a timely manner in response to the complex environment of the intestinal mucosa. Further studies are needed to test these possibilities. Activation of the Jak3-Jak1-PI3K-Akt-ERK signaling pathway is essential for IL-15-mediated CD8αα+ iIEL survival (Fig. 1). Although ERK activation is downstream of PI3K-Akt,

it was obviously delayed

compared to the activation of PI3K-Akt (Fig. 1C, D and Supporting Information Fig. 6, left panel). Consistently, the reduction of Bcl-2 level by MEK inhibition ABT-737 occurred later than that induced by Jak3 or PI3K inhibitor (Fig. 2A). It is possible that ERK1/2 activation was secondary to IL-15 selleck inhibitor stimulation. However, our preliminary experiments using supernatant from 40 h IL-15-treated CD8αα+ iIELs did not support the possibility that IL-15-induced secretory factor(s) activated ERK1/2 in CD8αα+ iIELs (Supporting Information Fig. 6, right panel). Other possible causes for the delayed and sustained ERK1/2 activation includes prolonged activation of upstream kinase and diminished activation of phosphatase. In view of these findings, we propose a stepwise model for IL-15-mediated CD8αα+ iIEL survival (Supporting Information Fig. 7). IL-15 first upregulates prosurvival Bcl-2 and Mcl-1 via activation of the Jak3-Jak1-PI3K-Akt pathway. With elevated Bcl-2, IL-15 induces ERK1/2-mediated phosphorylation of Bim at Ser65 to release Bcl-2 from the Bcl-2-Bim complex and to keep Bim in a phosphorylated SPTLC1 state. Activated ERK1/2 also participates in the maintenance of Bcl-2 level. The increase of Bcl-2 abundance and freed Bcl-2 shift the balance of Bcl-2 and Bim function toward promoting CD8αα+ iIEL survival. C57BL/6J (B6) and B6 human (hu) BCL-2

transgenic (B6-Tg (BCL2) 36Wehi/J) mice were purchased from the Jackson Laboratories. Il15ra−/− mice were generated in our lab [43] and backcrossed to B6 for 24 generations. RNA polymerase II-driven huMCL-1 transgenic mice in the B6 background were generated in Dr. S.-F. Yang-Yen’s lab [44]; Bim−/− mice were kindly provided by Dr. Jeffery C. Y. Yen (Institute of Biomedical Science, Academia Sinica, Taiwan). All mice were raised in a specific pathogen-free facility at the Institute of Molecular Biology, Academia Sinica. The mice were used at 8–22 weeks of age. All mice experiments were approved by the Institutional Animal Care and Use Committee at Academia Sinica and conformed to the relevant regulations.

Anyway the combined inhibition of p38 and p44/42 had the greatest impact on the cytokine secretion and the TLR-APC phenotype. Blocking experiments show that STAT-3 and MAPKs are essential for

Lumacaftor molecular weight the TLR-APC phenotype. To connect the MAPK and STAT-3 findings, we checked STAT-3 activation after MAPK inhibition to find that after blocking p38/p44/42 almost no tyrosine phosphorylation of STAT-3 was detectable (Fig. 9A). This effect could be overcome by the addition of exogenous IL-6 and IL-10 (Fig. 9C). Thus, the TLR-APC phenotype is dependent on the p38 and p44/42 MAPK-induced cytokine production and the resulting STAT-3 activation. An involvement of p38 and p44/42 in the activation of STAT-3 after TLR stimulation

has been observed also from others 46. Xie et al. 7 suggest that MAPK p38 activity might be responsible for the impaired differentiation of monocytes into iDCs after LPS stimulation. One day after LPS stimulation, p38 is activated and p44/42 not. Due to the late time point (d1), the initial and short activation of p44/42 was not seen, thus the link between p44/42 MAPK, IL-6 production and STAT-3 activation was missed. Our results indicate that TLR agonists added at an early time point of iDC differentiation induce a shift from STAT-5 toward STAT-3 activation and thus critical determine the functional phenotype of the APCs. We have shown before, that the addition of LPS during www.selleckchem.com/products/MG132.html the differentiation of murine bone marrow cells into myeloid DCs led to a reduced CD11c expression 5. The effect on CD11c could be traced back to a SOCS-1 dependent blockade of STAT-5 phosphorylation. Additionally, we could show that SOCS-3 is also able to reduce STAT-5 phosphorylation 5. Since TLR-APC upregulate preferentially SOCS3

(data not shown) we suppose that in the human system the block of STAT-5 might be SOCS-3-dependent. Hence, two different mechanisms seem to balance STAT-5/STAT-3 and thus regulate the expression of CD14, PD-L1 and CD1a. During infection, pathogen-derived TLR-agonists might bypass conventional iDCs differentiation and induce PD-L1-expressing tolerogenic APCs in a STAT-3-dependent manner. Studies investigating organs and tissues with close contact to microbial TLR agonists provide O-methylated flavonoid indications of the in vivo relevance of TLR-APC. For example, the liver has to deal with gut-derived portal blood that contains high concentrations of bacterial products. It has been demonstrated that liver DCs have reduced T-cell stimulatory capacities 47, 48. The data of Lunz et al. 49 support these findings. They could show that gut-derived bacterial products induce IL-6/STAT-3 signaling and thereby inhibit the hepatic DC activation/maturation. In summary, we show here that STAT-3 is responsible for the regulation of PD-L1 expression, triggered via IL-6 and IL-10. TLR agonists potently induce STAT-3 activation and thus direct DC differentiation to tolerogenic APCs.

When the strength of activating signals is powerful over the sum of inhibitory signals. NK cells and CD8+T cells will respond and kill the target cells [13]. In this study, the levels of NKG2A expression on CD3−CD56+NK cells and CD8+T cells were elevated to further examine whether lower expression of NKG2D was associated with over-expression of NKG2A. The results showed that there was no difference between the KD patients and the healthy Ixazomib controls in the percentage of CD3−CD56+NKG2A+NK cells (56.55% ± 10.23% versus 55.89% ± 7.90%, t = 0.050, P > 0.05) and CD8+NKG2A+T cells (5.40% ± 2.10% versus 6.68% ± 2.30%, t = 0.922, P > 0.05)

(Fig. 5). As shown in Fig. 6, there was no obvious difference to be found between the patients with KD and the healthy controls in the percentage of CD14+MICA+MC (6.15% ± 2.44% versus 5.27% ± 1.73%, t = 1.838, P > 0.05) and CD14+ULBP-1+MC (4.58% ± 1.76% versus 3.81% ± 1.61%, t = 0.764, selleck products P > 0.05). Kawasaki disease is currently recognized as an acute vasculitis resulted from immune dysfunction. The proinflammatory cytokines (such as TNF-α) are obviously elevated during the acute phase of KD and might be involved in the pathogenesis vasculitis in KD, but the mechanism triggering the cascade response of proinflammatory cytokine production

needs further clarification. Recent work demonstrated that NKG2D is expressed on most human P-type ATPase NK cells and CD8+T cells and is upregulated upon activation and stimulation [4, 14]. NK cells and CD8+T cells kill a variety of tumour cells, virus-infected cells and allogeneic cells in a nonmajor histocompatibility complex restricted manner and provide the first line of immune defence, thus representing a

useful tool to maintain host integrity. It is becoming increasingly appreciated that NK cells or CD8+T cells may play an immunoregulatory role in limiting autoimmune responses. Elimination of activated immune cells is one mechanism by which NK cells perform this immunoregulatory role. NKG2D plays a key role in immune regulation by bridging the crosstalk between NK cells, T cells and APCs such as dendritic cells or monocytes. Moreover, a role for NKG2D-dependent NK cells and CD8+T cells killing of activated immune cells has been proposed as a mechanism to dampen immune responses. As previously mentioned, inappropriate or deregulated expression of NKG2D on NK cells or CD8+T cells can break the delicate balance between immune activation and tolerance and trigger aberrant immune response [15, 16]. It has been reported that several autoimmune diseases associated with deviant NKG2D signalling, including type I diabetes, coeliac disease, SLE and rheumatoid arthritis, which were characterized by the feature of presence and aberrantly activation of a certain population of autoreactive immune cells [13, 17, 18].

Swidler[5] states that ‘although dialysis is life-sustaining therapy and extends life, it may also create, increase or prolong suffering while not restoring or maintaining well-being, function or cognition’ … and ‘to address suffering it must first be realised’. The burden of suffering may not be realized by a consultant who sees the patient infrequently but will be borne greatly by dialysis nurses and registrars. This is an often neglected

ethical issue. Beneficence We are obliged to provide our patients with the greatest benefit; to this end we should do our utmost to select patients most likely to benefit from dialysis, not just in terms of prolongation of life but in maintenance of worthwhile QOL. Justice We are obliged to provide BMS-777607 supplier our patients equal opportunity and allocation of available resources; in general terms we are fortunate in Australia and New Zealand that this principle rarely comes into play when making decisions around dialysis. In summary,

nephrologists’ thinking about elderly patients with ESKD needs to shift from traditional markers of medical ‘success’ to focus on the patients’ JQ1 symptoms and function as much or more than survival. This will help make an appropriate decision about suitability for dialysis. We believe that in making the decision to embark upon or forgo dialysis, we should consider all the above principles and enhance ESKD patient & family education to ensure that

the option of non-dialysis heptaminol conservative RSC is at least an equal offer to dialysis. This is best done with a formal RSC programme in place in each unit. Importantly all elderly patients with ESKD who do not receive dialysis need to not feel abandoned and know that all ongoing ESKD treatment will continue with their nephrologist. Finally, we already have some guidelines that discuss when it is OK to forgo dialysis, including Caring for Australians with Renal Impairment (CARI) & Renal Physicians Association (RPA) USA guidelines, discussed in the section by Crail ‘Management guidelines for patients choosing the RSC pathway: Information and web-based treatment protocols available to all’. Rosemary Masterson and Celine Foote There is a disproportionate increase in the number of elderly patients, many with multiple comorbidities, commencing dialysis. Predictors of survival for elderly patients on dialysis include age, comorbidity score, malnutrition, poor functional status and late referral. Patients with high comorbidity scores may not gain a survival advantage with dialysis versus a non-dialysis pathway. Late referral and lack of dialysis access are independent predictors of mortality in elderly patients commencing dialysis. Hospital free survival may be similar in dialysis and non-dialysis treated groups.

They suggest that screening for microvascular dysfunction using a combination of the approaches described above may be advantageous for the early detection of microvascular disease, in aiding diagnosis, in monitoring disease progression and response to therapy. Furthermore, they demonstrate a co-linearity in the development

and progression of microvascular and macrovascular disease. Much effort has gone into establishing whether there is a causal effect in either direction or whether this co-linearity simply represents shared risk factors. It is most likely to be a complex combination of bidirectional interactions. While the techniques used to measure microcirculatory structure

and function Selleckchem PLX4032 Selleckchem C59 wnt have become more robust and better understood over the past few decades, their application to the study of large populations remains limited. Furthermore, their ability to provide a mechanistic understanding of the processes underlying the pathology of microvascular disease is restricted by the need to interrogate accessible microvascular beds influenced by a wide range of confounding factors. Also included in this volume of Microcirculation is a review article from Cheung and Daanen [2] in which they present longitudinal and laboratory studies investigating dynamic adaptation of the peripheral microvasculature to cold exposure and improved tolerance in those living in cold environments. Collectively, the clinical microcirculatory research evidenced

in these articles provide readers with a unique opportunity to gain further insight into the challenges facing out those working at the translational interface seeking new ways in which to interrogate the human microcirculation. “
“Microcirculation (2010) 17, 237–249. doi: 10.1111/j.1549-8719.2010.00026.x The mammalian transient receptor potential (TRP) superfamily consists of six subfamilies that are defined by structural homology: TRPC (conventional or canonical), TRPV (vanilloid), TRPM (melastatin), TRPA (ankyrin), TRPP (polycystin), and TRPML (mucoliptin). This review focuses on channels belonging to the vanilloid (V) and melastatin (M) TRP subfamilies. The TRPV subfamily consists of six members (TRPV1-6) and the TRPM subfamily has eight (TRPM1-8). The basic biophysical properties of these channels are briefly described. All of these channels except TRPV5, TRPV6, and TRPM1 are reportedly present in arterial smooth muscle from various segments of the vasculature. Studies demonstrating involvement of TRPV1, TRPV2, TRPV4, TRPM4, TRPM7, and TRPM8 in regulation of arterial smooth muscle function are reviewed. The functions of TRPV3, TRPM2, TRPM3, and TRPM6 channels in arterial myocytes have not been reported.

Cell culture and stimulation.  PBMCs were cultured in complete RPMI-1640 culture medium supplemented with 7.5% heat-inactivated foetal calf serum (Sigma-Aldrich, St. Louis, MO, USA) and plated on 24-well plates. For stimulation, cells were incubated with

anti-CD3/anti-CD28-coated beads (Invitrogen Dynal AS, Oslo, Norway) at a bead:cell ratio of 1.0. Proliferation assay.  For cell proliferation assay, PBMCs were labelled with carboxyfluorescein diacetate (CFSE) (Molecular Probes, Inc., Eugene, OR, USA) according to the manufacturers recommendations. At the end of the culture period, the CFSE labelled cells were stained with anti-CD4-APC, anti-CD8-PerCP and anti-CD25-APC-Cy7 monoclonal antibodies (mAbs) (BD Pharmingen, San Diego, CA, USA), Cisplatin nmr washed and then run immediately on the flow cytometer. The BD FACS Aria (Becton-Dickinson, Franklin Lakes,

NJ, USA) was used for all measurements of our study. Cell death assay.  Cells Selleck ACP-196 were stained with anti-CD4-PE-Cy7, anti-CD25-FITC and anti-CD8 APC-Cy7 mAbs (BD Pharmingen). After 10 min of incubation in dark, propidium iodide was added and samples were incubated for 10 more min. The samples were then analysed immediately on flow cytometer. Intracellular FoxP3 assay for the identification of Tregs.  For analysis of Foxp3, cells were first stained for the expression of CD4 and CD25 surface molecules with anti-CD4 APC and anti-CD25 FITC mAbs (both BD PharMingen). Cells were fixed and permeabilized based on the manufacturer’s recommendations (Fixation/Permeabilization solution,

Permeabilization solution, eBioscience, San Diego, CA, USA). Anti-Foxp3 PE mAb (eBioscience) was then used for intracellular staining (40 min at 4 °C), and corresponding isotype control was also included. Cells were washed once and analysed immediately on flow cytometer. Surface markers of T lymphocytes.  To determine C1GALT1 the activation, maturation markers and Th1/Th2 polarization of CD4+ and CD8+ lymphocytes, the following mAbs were used in combinations: anti-CD4 PE-Cy7, anti-CD8 APC-Cy7, anti-CXCR3 APC (Th1), anti-CCR4 PE (Th2), anti-CD62L PE-Cy5, anti-CD25 FITC, anti-CD69 APC, anti-CD45RO PE, anti-HLA-DR PerCP, anti-CD45RA FITC (all purchased from BD Pharmingen). The cells were incubated with mAbs for 20 min in dark, washed once and analysed on flow cytometer. Statistical analysis.  Median [range] of the variables is reported. Hettmansperger–Norton trend test was applied to investigate the trend of the changes with increasing hyperoxia time [18]. P values <0.05, two tailed, were considered significant. We did not correct for multiplicity. Mann–Whitney U test was used for comparison of two groups. Data are summarized in Table 1 for cell cultures without T cell stimulation and in Table 2 for experiments with anti-CD3/CD28 bead stimulation.

Mucormycosis often exhibit different clinical forms. A few types are primarily cutaneous and subcutaneous infections and may also happen in immunocompetent patients, with long course and no dissemination. Most types, however, are deep and rapidly progressive mycoses targeting immunocompromised patients. Characteristic features

like thrombosis and tissue necrosis at the site of infection[7, 8] coincide with mortality rates ranging from 30% to 90%.[9] The number of cases of mucormycosis has been increased in past few years, especially among diabetic, neutropenic, thrombocytopenic and immunocompromised patients.[10-15] Among the members of Mucorales; Rhizopus, Mucor and Lichtheimia species are the main causative agents for mucormycosis in 70–80% cases.[15-18] find more The route of infection is mainly via the respiratory tract due to its property of being highly airborne, followed by the skin and less commonly via the gut which is more often found in

case of neonates. The most common type of infection comprises the involvement of sinus (39%), pulmonary (24%) and lastly cutaneous (19%) with development of dissemination in 23% of all cases. Pulmonary infection is most commonly found PLX4032 cell line among malignant patients while the involvement of the sinuses is the most abundant among patients with diabetes.[13] Entomophthorales are pathogenic fungi for insects and humans. Like Mucorales, they are environmental saprophytic fungi, commonly found in decaying matters.

On the other hand, it is linked to areas with tropical climates hence commonly found in India, Africa, South America and Caribbean Islands. However, there are some rare cases emerging from the United States.[19-21] Its infection, summarised to entomophthoromycoses, can be divided into two types; basidiobolomycosis and conidiobolomycosis. Unlike Mucorales, the cases with Entomophthorales Rutecarpine are often associated with immunocompetent patients and it is not associated with rapid angio-invasive or disseminated infections. It is described as a chronic and slowly progressive infection.[20, 22] Basidiobolomycosis is caused by Basidiobolus ranarum and conidiobolomycosis is due to subcutaneous infection of Conidiobolus coronatus or C. incongruus. Common mode of transmission is via traumatic inoculation. Histological examination of infected lesions may display eosinophilic infiltration and Splendor-Hoeppli phenomenon (non-septate hyphae surrounded by an eosinophilic halo).[20, 23] Apart from those infectious diseases, Lichtheimia corymbifera; a close relative of Rhizopus oryzae and member of the zygomycetous order Mucorales, can lead to another non-infectious disease called farmer’s lung disease (FLD); one type of hypersensitivity pneumonitis.[24] This is due to the inhalation of spores from agricultural products (e.g. hay, grains etc.) leading to accumulation of inflammatory cells in the lung of the patients.

The important discovery that transforming growth factor (TGF)-β and IL-6 could promote Th17 differentiation from naive T cells [10] prompted studies

that confirmed that Treg can also be generated in vitro by stimulation with TGF-β in the absence of IL-6 [11,12]. The remarkable balancing act of adaptive immunity to facilitate the targeted destruction of pathogens without excessive collateral damage to self is nowhere better exemplified than in the shared use of TGF-β in controlling the newly described Th17 effector lineage and adaptive Treg development. Probiotic bacteria can be potent inducers of cytokines, for example Gram-positive bacteria, have been found to stimulate IL-12, while Gram-negative bacteria tend to stimulate IL-10 production [13]. Several studies have demonstrated that selected probiotics are able to induce the production of proinflammatory cytokines Selleckchem Palbociclib by macrophages and Th1 cytokines by peripheral blood monocytes [14,15]. However, little is known about the effects of exposure time and bacterial state on the stimulation

of cytokine production. As such, the aim of this study was to profile pro- and anti-inflammatory cytokines secretion from human peripheral blood mononuclear cells (PBMCs)and the CRL-9850 cell line and the differentiation of Th17 or induced Treg cells following exposure to various strains of live, heat-killed or gastrointestinal tract (GIT)-simulated bacteria. Lb. acidophilus LAVRI-A1, Bifidobacterium (B.) lactis B94 PAK5 and Lb. rhamnosus GG (LGG) were kindly provided by DSM Food Specialties (Moorebank, NSW, Australia), and Vaalia Parmalat buy KPT-330 Australia Ltd (South Brisbane, Queensland, Australia), respectively. Exopolysaccharides-producing Streptococcus (S.) thermophilus St1275, B. longum BL536 and pathogenic Escherichia

(E.) coli TG1 used as a Gram-negative control strain were supplied by the culture collection of Victoria University (Melbourne, Australia). Strains were stored at −80°C in 40% glycerol. Sterile 10 ml aliquots of de Man Rogosa and Sharpe (MRS) broth (Sigma Chemical Co., St Louis, USA) were inoculated with 1% (v/v) LAVRI-A1 and LGG. Additionally, sterile 10 ml aliquots of MRS were supplemented with 0·05% L-cystein.HCl and inoculated with 1% (v/v) B94 and BL536 and incubated at 37°C for 18 h. For the propagation of E. coli and St1275, 1% (v/v) of either strain was used to inoculate 10 ml tryptic soy broth (BHI; Difco Laboratories, Sparks, MD, USA) or M17 broth (Amyl Media, Dandenong, Australia), respectively [16]. Following two successive transfers to fresh 10-ml broth preparations, bacteria were grown for 18 h log phase growth. Cultures were harvested at 1360 g for 30 min at 4°C. To heat kill, samples were incubated at 80°C for 30 min. GIT-simulated samples were treated as described below. Following these manipulations, preparations were centrifuged and the pellet resuspended in phosphate-buffered saline (PBS).