After experiments, the explants were snap frozen or embedded in p

After experiments, the explants were snap frozen or embedded in paraffin. Paraffin-embedded sections or cryostatic sections were incubated with Abs against phospho-STAT1 (Tyr701) (Santa Cruz Biotechnology), ICAM-1

(clone HA58, BD Pharmingen), HLA-DR (clone G46–6, BD Pharmingen), CXCL10 (C-19, Santa Cruz Biotechnology). Secondary biotinylated mAbs and staining kits (Vector Laboratories, Burlinagame, CA, USA) were used to develop immunoreactivities, and 9-ethyl-3-aminocarbazole Trichostatin A solubility dmso was used as substrate. Sections were counterstained with hematoxylin. Statistical significance was evaluated using Wilcoxon’s signed rank test (SigmaStat; Jandel, San Rafael, CA, USA). Values of p ≤ 0.05 were considered significant. This work was supported by the Italian Ministry of Health and by Ministero dell’Università e della Ricerca Scientifica (MIUR). The authors declare no financial or commercial conflict of interest. “
“There is debate over whether effective T-cell mediated protection against a second infection, or post-vaccination, is better done

by central memory cells or effector memory cells. The former may have greater powers of expansion, whereas the latter may be closer to the site of pathogen entry and faster to respond. This review focuses on memory T cells which are not recirculating but which remain at the peripheral Lumacaftor concentration site of initial pathogen or vaccine encounter, so-called tissue-resident memory cells. They may play key roles in protection against re-eruption of latent viral infections and at mucosal surfaces. After leaving the thymus, newly generated T cells have a few steps of continued maturation or polishing to undergo before they become fully

mature naïve T cells 1. As naïve cells, peripheral T cells migrate between the blood and the lymphoid structures in the spleen and lymph nodes in search of their cognate antigen. When T cells do encounter antigen on activated DCs in central lymphoid organs, they proliferate Sorafenib cost and differentiate into effector T cells. While some antigen-activated T cells, such as CD4+ follicular helper T cells, may remain in the central lymphoid organs to deliver help to B cells 2, those effector cells whose work is at the peripheral site of antigen entry must travel to this site via the bloodstream. Using cues from activated endothelial cells at sites of inflammation 3, T cells leave the blood vessels and enter tissues once more in search of antigen. When antigen-bearing cells are killed or accessory cells are activated to degrade or contain antigen, effector cells egress from the tissues via the afferent lymphatics. In some cases, a few effector cells remain behind; these tissue-resident memory T cells are the subject of this review 4. When antigen has been cleared, a contraction phase follows during which time the number of effector cells declines through apoptosis leaving behind some survivors that go on to differentiate into memory T cells.

3 In contrast, monocyte-derived DCs (MoDCs) are generated during

3 In contrast, monocyte-derived DCs (MoDCs) are generated during inflammation.4,5 Dendritic cells have been extensively characterized in a variety of species and protocols for obtaining DC subtypes range from in vitro culture methods to direct isolation of DCs from blood and tissues. Isolation, however, is complicated in humans and large animal species resulting in limited availability of functional studies. In pigs, blood

DCs (BDCs) have only been investigated in a few studies and very little is known about the function of these DCs in antigen presentation and T-cell activation. The objectives of the C59 wnt manufacturer present study were to compare directly isolated porcine BDCs with traditionally generated porcine MoDCs in terms of phenotype and functionality. Various porcine DCs have been described including bone marrow-derived (BM) DCs,6 Langerhans-type cells7 and MoDCs.6–11 The MoDCs are the most widely used subtype and can be phenotyped as CD1+, CD14+/−, CD16+, CD80/86+, CD172+, major histocompatibility complex (MHC) I+, MHC II+, CD4−, CD3−, and CD8−.6,7 Initially RAD001 MoDCs were generated by isolation of peripheral blood

mononuclear cells (PBMCs) followed by overnight plastic adherence. Non-adherent cells were then removed and the remaining monocytes were cultured in the presence of interleukin-4 (IL-4) and granulocyte–macrophage colony-stimulating factor (GM-CSF).6 More recent protocols, however, involve the isolation of monocytes using antibodies against CD1412,13 or CD172a,14 a porcine marker known as SWC3 that is present on myeloid cells15 including cDCs and pDCs.16 Porcine BDCs, on the other hand, comprising pDCs and cDCs, were originally described by Summerfield et al.,16 by flow cytometric analysis of PBMCs as being CD172a+, MHC II+, CD80/86+, CD1+/− and CD14− with pDCs being CD4+ and cDCs being CD4−. Subsequently,

this approach was further developed by isolating BDCs using antibodies against CD172a. However, because ASK1 CD172a is also expressed on monocytes, these enriched BDC populations contained not only different DC subtypes but monocytes as well.17 In the present study, we adapt previous protocols by initially depleting monocytes and subsequently enriching for CD172a to achieve a purer BDC population. These BDCs were compared with MoDCs in terms of antigen uptake, activation and maturation. DC maturation occurs upon recognition of microbe-associated molecule patterns and is characterized by up-regulation of co-stimulatory molecules such as CD80/86 and MHC II, various cytokines and the chemokine receptor CCR7.18,19 The process of maturation occurs as DCs migrate towards the lymph nodes where they encounter naive or primed T cells. In porcine MoDCs, stimulation with lipopolysaccharide (LPS) was demonstrated to decrease the expression of CD16, up-regulate the expression of CD80/866,20 and either increase7 or have no effect6,20 on expression of MHC II.

We measured and analyzed

parameters of lymphatic contract

We measured and analyzed

parameters of lymphatic contractility in isolated and pressurized rat MLVs under control conditions and after pharmacological blockade of NO by l-NAME (100 μM) selleck inhibitor or/and histamine production by α-MHD (10 μM). Effectiveness of α-MHD was confirmed immunohistochemically. We also used immunohistochemical labeling and Western blot analysis of the histamine-producing enzyme, HDC. In addition, we blocked HDC protein expression in MLVs by transient transfection with vivo-morpholino oligos. We found that only combined pharmacological blockade of NO and histamine production completely eliminates flow-dependent relaxation of lymphatic vessels, thus confirming a role for histamine as an EDRF in MLVs. We also confirmed the presence of HDC and histamine inside lymphatic endothelial cells. This study supports a role this website for histamine as an EDRF in MLVs. “
“Microcirculation (2010) 17, 21–31. doi: 10.1111/j.1549-8719.2009.00007.x Low birth weight is an indicator of exposure to unfavorable fetal environment and has been associated with the development of hypertension and cardiovascular disease in adulthood. There is now growing evidence suggesting that alterations in the microcirculation associated with exposure to a suboptimal in utero environment play a key role in the development of cardiovascular disease. Proposed

hypothetical mechanisms include: fetal circulatory redistribution, impaired synthesis of elastin, and endothelial dysfunction in response to antenatal and postnatal environment. More recent studies have shown associations of low birth weight with capillary rarefaction and narrowing retinal arteriolar caliber in both children and adults. This suggests that vascular adaptations in utero persist into maladaptive circulatory changes in adulthood, which may reflect an increased susceptibility to hypertension and cardiovascular disease later in life.

Therefore, the association between low birth weight and narrower retinal arteriolar caliber, together with associations between Farnesyltransferase narrower retinal arteriolar caliber and risk of hypertension and cardiovascular disease, suggest that retinal arteriolar narrowing may be a marker on the microvascular pathway and mechanisms linking early life exposures and subsequent cardiovascular disease. “
“Kv7 channels are considered important regulators of vascular smooth muscle contractility. The present study examined the hypotheses that 1. Kv7 channels are present in mouse cerebral and coronary arteries and regulate vascular reactivity, and 2. regional differences exist in the activity of these channels. PCR confirmed that basilar, Circle of Willis and left anterior descending (LAD) arteries express predominantly Kv7.1 and 7.4. Western blot analysis, however, showed greater Kv7.4 protein levels in the cerebral vessels. Relaxation to the Kv7 channel activator, retigabine (1-50μM) was significantly greater in basilar compared to LAD.

, 2003; Avonce et al , 2006; Cardoso et al ,

2007) It is

, 2003; Avonce et al., 2006; Cardoso et al.,

2007). It is tempting to speculate that A. baumannii trehalose production contributes to the organism’s ability to tolerate desiccation and thus may contribute to its transmission in the hospital setting. RT-PCR confirmed that members of the trehalose metabolic pathway are dramatically upregulated during stationary as opposed to exponential phase growth (Fig. 2). A hallmark of biofilm assembly is the transition from surface attachment to biofilm accumulation and maintenance. In that regard, our data also indicated that genes that are known to be associated with the initial stages of biofilm formation are Cilomilast in vivo predominantly expressed during exponential growth, whereas genes associated with biofilm maintenance are upregulated during stationary phase. More specifically, during the initial stages of A. baumannii biofilm formation, the csu operon is thought to modulate pili formation and, consequently, contribute to pilus-mediated attachment to abiotic surfaces (Tomaras et al., 2003). We found Stem Cell Compound Library that two members of the csu operon, csuA/B (A1S_2218) and csuC (A1S_2215), as well as a putative pili assembly chaperone (A1S_1509), were upregulated during exponential phase of growth. Conversely, during stationary

phase of growth, putative members of the second messenger cyclic diguanylate (c-di-GMP; A1S-1949) and exopolysaccharide (A1S_1987) synthesis machinery were upregulated. In Pseudomonas aeruginosa, a close A. baumannii relative, c-di-GMP is hypothesized to play a role in the latter stages of biofilm formation. C-di-GMP augments biofilm maturation in two ways:

(1) it activates extracellular polysaccharide production, leading to a thickening of biofilm matrices, BCKDHA and (2) it suppresses twitching motility and swimming (Tamayo et al., 2007). Collectively, these results indicate that exponential and stationary phase-induced ORFs would allow A. baumannii to initiate attachment to a surface, produce exopolysaccharide, and then mature into a hardy biofilm. Gram-negative bacterial secretion systems are responsible for the translocation of proteins across the double membrane. During exponential phase, a putative general secretion pathway protein (A1S_0269), with homology to type II secretion system (T2SS) proteins, was upregulated. Additionally, five loci from the Sec pathway were also induced; this pathway is essential in transporting proteins across the inner membrane before they can be excreted by the T2SS. In several bacterial species, including Vibrio cholerae and P. aeruginosa, the T2SS secretes toxins, proteases, phospholipases, and other virulence-associated proteins (Sandkvist, 2001). A putative type III effector protein (A1S_0390) was also induced during exponential phase of growth.

Urine levels of TGF-β1 and connective

tissue growth facto

Urine levels of TGF-β1 and connective

tissue growth factor increase with the progression of CKD;63–65 however, TGF-β1 is mostly Wnt signaling excreted as an inactive complex, which requires brief acidification to permit activation and detection. Some profibrotic molecules that are induced by TGF-β1, such as TGF-β-inducible gene H3 (βig-H3) and plasminogen activator inhibitor-1, are also detectable in urine and can act as surrogate markers of renal TGF-β1 activity. Urine levels of βig-H3 are about approximately 1000 times greater than TGF-β1 in diabetic patients and can be detected before the onset of albuminuria,66 indicating that βig-H3 is an early and sensitive marker of renal fibrosis during diabetes. Urine excretion of plasminogen activator inhibitor-1 has been shown to correlate with renal injury and fibrosis in patients with diabetic nephropathy and progressive chronic glomerulonephritis.67,68 Collagen type IV is a major component of kidney extracellular matrix, which is increased during the progression of renal fibrosis. Urine excretion of collagen IV is elevated in patients with IgA nephropathy and diabetic nephropathy and correlates with declining renal function.69,70 In addition, urine levels of collagen IV correlate AP24534 order with glomerular matrix accumulation and declining renal function in animal models of kidney disease.71 In contrast, serum levels of collagen IV are not associated with the development

of renal injury or loss of kidney Acetophenone function.72 Although reliable ELISA exists for most of the recently described renal biomarkers in serum and urine, this technique is limited to measuring a single marker per assay, which makes assessment of multiple biomarkers time-consuming and expensive. Recently, multiplex assay systems have been developed by Luminex ( and

BD Biosciences (, which uses the principles of both ELISA and flow cytometry to simultaneously quantitate multiple antigens in biological fluids. In the Luminex assays, microspheres with unique spectral signatures are coupled with primary antibodies. The antigens binding to these microspheres are then labelled with biotinylated secondary antibodies and streptavidin coupled to another fluorochrome (phycoerythrin). The microspheres and antigens labelled with phycoerythrin are excited with lasers at different wavelengths and the emission signals are used to identify the antigen and the amount of antigen bound to the microsphere. This technique is theoretically capable of assessing up to 100 different antigens and requires small volumes of biological fluid (30 µL). The Luminex assay system has been used to assess multiple biomarkers in the urine of patients with renal allograft rejection and lupus nephritis.51,73 The advantages and technical considerations for multiplex assays have been recently reviewed by Leng et al.

The plates were washed with PBS and blocked with 1% polyvinylpyrr

The plates were washed with PBS and blocked with 1% polyvinylpyrrolidone (Sigma, Munich, Germany) at room temperature for 1 h and then washed extensively with PBS at 37°C for 40 min. A total of 2.5 × 105 neutrophils in 500 μL of DPBS were pretreated with rmTNF (50 ng/mL at 37°C for 15 min) and then added to the wells for 40 min. Plates were then washed gently three times with prewarmed PBS and the remaining adherent cells were quantified by counting three microscopic fields at a 40× magnification. RNA was prepared as described [44]. Briefly, murine PMNs were isolated and RNA was immediately prepared with TriFast (Peqlab, Erlangen, Germany). Reverse transcription

was performed on 1 μg of RNA using random hexamers reverse transcriptase. A total of 200 nM of the resulting cDNA was then subjected to 40 cycles of PCR in a 25 μL reaction mixtures in a BioRad cell cycler (BioRad, Munich, Germany). The PCR products were subjected to agarose gel analysis; m24p3R fw: GGC GAT TTC TAC AGG GAA TGA rv: CTA TCA GCC ACC GTG CAG ACT; mMegalin fw: TGC HDAC assay ACG GAG GAA GTT GCT ATT rv: TCC ACT GTA GCC GCT AGA ACA. Rabbit polyclonal sera were raised against 24p3R. The sequences of the synthetic peptid used and

the location within the primary amino acid sequence was CDHVPLLATPNPAL (anti-24p3R: 507–520). Crude serum was affinity purified. Antibody production and affinity purification were performed by Eurogentec (LIEGE Science Park, Belgium). Protein extracts were prepared from freshly isolated human PMNs using cytoplasmic

lysis buffer (50 mM Tris-HCl, pH 7.6; 150 mM NaCl; 1% NP-40 with protease inhibitors). Ten micrograms of protein were resolved by SDS-PAGE (BioRad) and transferred to nitrocellulose membranes (Amersham Hybond-P; GE Healthcare, Buckinghamshire, UK). Membranes were blocked with 4% blocking milk/TBS/Tween and incubated with Abs against anti-human 24p3R (Eurogentec) and antiactin (Sigma). Oxyblots were performed using the ADP ribosylation factor SuperSignal West Dura Extended Duration Substrate (Thermo Scientific, Vienna, Austria) according to the manufacturer’s instructions. Freshly isolated murine blood was drawn by retroorbital puncture. Samples were stained with anti-mouse CD11b Alexa Fluor 488 (M1/70, Biolegend, Uithoorn, the Netherlands), anti-mouse Ly-6G/Ly-6C PerCP (RB6–8C5, Biolegend), anti-mouse Ly-6G FITC (1A8, Biolegend), anti-mouse CD182 PerCP/Cy5.5 (TG11/CXCR2, Biolegend), anti-mouse CD184 Alexa Fluor 647 (TG12/CXCR4, Biolegend), anti-mouse CD51-PE (RMV-7, Biolegend), anti-mouse CD62L Alexa Fluor 647 (MEL-14, Biolegend), or appropriate isotype IgGs. Cells were measured with BD FACS Calibur flow cytometer (BD Bioscience, Heidelberg, Germany) and analyzed with Kaluza Software (Beckman-Coulter, Vienna, Austria).

33 μM After 3 min of incubation, cells were vortexed at room tem

33 μM. After 3 min of incubation, cells were vortexed at room temperature and incubation continued for 2 additional minutes. One tenth of the volume of FCS click here was added and the cells were vortexed and incubated for 1 min. Samples were washed three times in complete media and used in experiments. Percent divided, division, and proliferation indices were determined by FlowJo software. Purified B cells (5 × 105) were used ex vivo or after 24–48 h of stimulation with media or 10 μg/mL anti-IgM in the presence of vehicle or dimedone.

Cells were harvested and surface stained with B220-allophycocyanin as described above. After surface staining, cells were resuspended in 7-amino-actinomycin D (7-AAD) and Annexin-V FITC (BD Pharmingen) for 15 min at room temperature according to the manufacturer’s protocol. Cells were acquired immediately on a FACSCalibur Instrument. All samples were analyzed using FlowJo Software. Purified (5 × 105) B cells were stimulated with 10 μg/mL anti-IgM in the presence of vehicle or dimedone. At 45 h, samples were pulsed with 10 μM BrdU (Sigma-Aldrich) for 3 h and labeling was performed as described previously [14]. Briefly, cells were harvested and resuspended in 1%

paraformaldehyde with 0.05% Igepal (Sigma-Aldrich), vortexed, and incubated at 4°C overnight. Samples were washed at room temperature two times with PBS at 1200 rpm for 6 min, resuspended in 1 mL PBS, and 4.2 mM MgCl2 containing 50 Kunitz U/mL DNase I (Sigma-Aldrich), and incubated at 37°C for 30 min. Following

two washes in wash buffer (PBS supplemented selleck compound with 5% FCS and 0.5% Igepal) at 1200 rpm for 6 min at 4°C, samples were resuspended in the same buffer containing 2% mouse serum and a 1/5 dilution of anti-BrdU FITC (BD Pharmingen). Samples were incubated on ice for 45 min. After two washes, cells were resuspended in 10 μL 7-AAD (BD Pharmingen) plus FACS buffer for 15 min on ice. Cells were acquired immediately on an FACSCalibur Instrument. Purified B cells (2 × 106) were stimulated in the presence or absence PLEKHM2 of dimedone with 10 μg/mL anti-IgM. After stimulation, cells were pelleted, washed with PBS, and lysed in buffer described previously [14]. Samples were boiled in the presence of reducing sample buffer, ran on a 7.5% precast SDS-PAGE gel (Bio-Rad), transferred to a PVDF membrane, and probed for phospho-Syk (Y525/526) (C87C1), Syk, phospho-p44/p42 MAPK (T202/Y204), or p44/p42 (Cell Signaling) according to the manufacturer’s protocol. For phospho-tyrosine detection, 2.5 × 106 purified B cells were stimulated in the presence or absence of dimedone with 10 μg/mL anti-IgM. Samples were lysed, ran on a SDS-PAGE, transferred to a membrane, and probed for tyrosine phosphorylated proteins (4G10-HRP, Millipore) as previously described by Fujimoto et al. [48]. After the blot was developed and imaged, it was stripped and probed with anti-actin as previously described [14].

BMDCs were resuspended 330 μL of PBS containing 3 μL of a proteas

BMDCs were resuspended 330 μL of PBS containing 3 μL of a protease inhibitor cocktail (Halt™ protease inhibitor-single-use cocktail; Thermo Scientific, Rockford, IL, USA) and 33 μL of Triton-X-114, and the mixture was alternatively vortexed and chilled for 5 min before being sedimented (10 000 × g for 10 min at 4°C) to remove the insoluble residue. The extraction was repeated twice. The pooled supernatants were incubated for 5 min at 37°C to allow the formation of micelles. Water phase and Triton phase containing the membrane proteins were separated after sedimentation at 10 000 × g for 5 min at room temperature. The water phase was removed. Membrane proteins in the Triton

phase were washed with 1 mL of PBS plus protease inhibitor cocktail (1%) and precipitated as described (22). The pellets were dried, resuspended in sample buffer for Selleck C59 wnt SDS–PAGE according to Laemmli (23) and stored at −20°C until use. Membrane protein fractions prepared as described above were heated at 65°C for 20 min before loading on a 12% gel. SDS–PAGE was performed as previously described (23), and the proteins were electrophoretically transferred onto a nitrocellulose sheet (Schleicher & Schüll, BA85). Blots were rinsed in PBS and incubated in blocking buffer, including PBS-Tween 20 (0·3%) plus 5% fat-free milk powder, during 2 h at room temperature.

The nitrocellulose membrane was then washed three times and incubated RAD001 chemical structure with the first antibody, rat anti-mouse MHC class II (I-A/I-E) (eBioscience,

THP, Vienna, Austria) diluted (1 : 500) in PBS-Tween 20 (0·3%) during 1 h. The membrane was subsequently washed three times and incubated with the secondary antibody, anti-rat-IgG-alkaline phosphatase-conjugate (Sigma Chem. Co.) for 1 h at a dilution (1 : 1000) in PBS-Tween 20 (0·3%). To visualize bands, the membrane was incubated in 10 mL of substrate buffer [MgCl2 (10 mm) + NaCl (100 mm) + Tris (100 mm)] adjusted to pH 9·5 and supplemented with 66 μL BCIP and 66 μL NBT, prepared, respectively, in 100% and 70% of dimethyl ASK1 formamide (DMF). At the last step, the membrane was transferred into water to stop the enzymatic reactions and then dried on absorbent paper at room temperature. The influence of peritoneal DCs isolated from metacestode-infected mice and naïve mice on the activation of naïve CD4+ pe-T cells was studied using an assay described previously (24); 5 × 104 of naive CD4+pe-T cells were stimulated with Con A (2 μg/mL) in the presence of varying numbers of naive pe-DCs or AE-pe-DCs ranging from 5 × 103 to 5 × 104 cells. Cells were incubated in 200 μL total volume of complete medium (RPMI-1640 supplemented with 10% FCS (v/v), 2 mm l-glutamine, 1 mm sodium pyruvate, 1 mm nonessential amino acids, 0·05 mm mercaptoethanol, 100 U/mL penicillin per streptomycin) during 48 h at 37°C and 5% CO2. Cell proliferation was assayed using the colorimetric BrdU (5-bromo-2-deoxyuridine) cell proliferation ELISA kit (Calbiochem, Merck Chemicals, Switzerland).

, 2009) In addition, BCG is not recommended for vaccination of i

, 2009). In addition, BCG is not recommended for vaccination of immunocompromised subjects because, in such individuals, it may cause disease itself (Hesseling et al., 2006; Marchand et al., 2008). Furthermore, due to the presence of cross-reactive antigens, BCG is not ideal for the vaccination of individuals with antimycobacterial reactivity (Crampin et al., 2009), and hence this

vaccine is not recommended for booster vaccination (Primm this website et al., 2004; Crampin et al., 2009). Therefore, current TB control focuses on the prompt detection of the diseased subjects with improved methods of diagnosis, and their treatment with effective drugs to prevent further transmission of the organism to healthy people (Lönnroth & Raviglione, 2008; WHO Report, 2009). In spite of some success of this strategy in controlling TB in industrialized countries, TB is persistently endemic in most of the poor and developing countries of the world (WHO Report, 2009). Furthermore, recent analyses suggest that the impact of current strategies of improved diagnostic and curative efforts to reduce TB incidence is less than expected and therefore these efforts need to be combined with additional preventive efforts (Lönnroth & Raviglione,

2008). Thus, there is a pressing need to develop new second-generation or booster vaccine(s), without which the global control of TB may not be achieved (Smith, 2009). Such vaccines may be based on cross-reactive antigens of M. tuberculosis, which are present in BCG and other mycobacteria, for example antigens of Ag85 complex and hsp65 (Mustafa, check details 2005a; Skeiky & Sadoff, 2006). However, one of the explanations given for the failure of BCG to protect against TB in adults is their sensitization to cross-reactive antigens through exposure to environmental mycobacteria (Crampin et al., 2009). Therefore, it may be wise to look for M. tuberculosis-specific antigens as alternative vaccines. The search for alternative vaccines and diagnostic reagents based on M. tuberculosis-specific antigens has been encouraged by

comparative genomic studies, which have shown that 16 genomic regions [known as regions of difference (RD) with designations RD1–RD16] of M. tuberculosis were lacking in M. bovis and/or M. bovis BCG (Behr et al., 1999; Gordon et al., 1999). Among these RDs, RD15 was predicted to have 15 ORFs, Rv1963c–Rv1977 GNA12 (Table 1) (Behr et al., 1999; Brosch et al., 2000), and is of special interest because it is absent in both pathogenic M. bovis and all vaccine strains of M. bovis BCG (Behr et al., 1999; Gordon et al., 1999). Furthermore, genes belonging to the third operon of mammalian cell entry (Mce3) proteins are located in this region (Behr et al., 1999; Gordon et al., 1999). Mce3 proteins are expressed in M. tuberculosis (Ahmad et al., 2004) and have been suggested to facilitate the entry of the pathogen in mammalian cells (El-Shazly et al., 2007). Furthermore, M.

(Barns et al , 1991; Dujon et al , 2004; Dujon, 2006) Both S  ce

(Barns et al., 1991; Dujon et al., 2004; Dujon, 2006). Both S. cerevisiae and C. glabrata can produce biofilms as haploids (Whelan et al., 1984; Hawser & Douglas, 1994; Reynolds & Fink, 2001) ZD1839 and form a thin biofilm layer of budding yeasts (Seneviratne et al., 2009; Haagensen et al., 2011). Saccharomyces cerevisiae is genetically tractable and has several properties that make it a favoured model organism (Guthrie & Fink, 1991). Saccharomyces cerevisiae is rarely pathogenic (McCusker et al., 1994), has a high rate of homologous recombination and has a highly versatile DNA transformation system (Rothstein, 1983; Wach et al., 1994). Because of its use

in the food industry and as a cell biology model, it has been studied extensively. Saccharomyces cerevisiae was the first eukaryotic genome to be sequenced (Goffeau et al., 1996), making it amenable to global genetic and phenotypic analysis. In addition, both transcriptomic (DeRisi et al., 1997; Velculescu et al., 1997) and proteomic (Zhu et al., 2001) studies were first applied in S. cerevisiae. Consequently, advanced genetic tools have been developed for this fungus. Ten years ago, Reynolds

Pexidartinib and Fink introduced S. cerevisiae as a model for yeast biofilm studies (Reynolds & Fink, 2001). Biofilm formation of S. cerevisiae and its regulation are conserved in opportunistic pathogenic Candida spp. (Rigden et al., 2004; Desai et al., 2011). Hence, understanding of adherence and its regulation in S. cerevisiae contributes to our understanding of the orthologous mechanisms in Candida spp. Other properties of yeast biofilms may also be conserved, such as quorum sensing (QS) mechanisms (Chen et al., 2004; Chen & Fink, 2006) and the presence of an ECM (Hawser & Douglas, 1994; Kuthan et al., 2003). Taken together, these make S. cerevisiae an attractive model for biofilm studies. In this review, we focus on the traits common to bacterial

and pathogenic yeast biofilms that are also found in S. cerevisiae, specifically adhesion, ECM, QS, drug resistance and evolution of cell surface variation. The knowledge of molecular mechanisms for cell–cell and cell–surface adherence in S. cerevisiae is detailed Protein tyrosine phosphatase and well reviewed (Brückner & Mösch, 2011). As adhesion is essential for biofilm, environmental cues and pathways regulating adhesion are also expected to affect biofilm development. Because less is known about the molecular mechanisms for matrix formation, QS and drug resistance, the last part of the review contains a discussion of novel microscopic techniques and state-of-the-art molecular genetics that can be applied to identify and investigating factors for S. cerevisiae biofilm development. Attachment of S. cerevisiae to foreign surfaces such as polystyrene is dependent on the cell surface protein Muc1/Flo11 (Reynolds & Fink, 2001). In S.