We discuss here important pro-inflammatory molecules and leucocyte populations that were identified as key players in the murine model of DENV-2 infection using the mouse-adapted strain P23085. The inflammatory response triggered by this model of DENV infection frequently leads to tissue damage and death. However, it is possible in this model to assess and distinguish mechanisms necessary for the host response

to deal with infection from those that cause unwanted, misplaced and uncontrolled inflammation and drive disease. RG-7388 solubility dmso By understanding where/how host–pathogen interactions lead to disease, we may be able to suggest novel strategies to restrain severe systemic and local inflammatory responses. Chemokines are members of a structurally related family of cytokines involved in leucocyte GSK1120212 chemical structure traffic during infection and inflammation. They are classified according to the relative position of conserved N-terminal cysteine residues, in which CC

chemokines represent the most abundant family and have the first two cysteines placed adjacently.[72] Chemokine receptors are expressed on the surface of leucocytes and are G protein-coupled receptors containing seven transmembrane domains.[73] Experimental and epidemiological evidence suggests an important role for chemokines, especially those from the CC family, and their receptors in infectious diseases such as HIV and herpes simplex virus 1.[74, 75] The expression of CC chemokines dominates over the expression of CXC chemokines during

viral infections, although this observation does not represent a general rule.[75] Among the CC chemokines, CCL3/MIP-1α and CCL5/regulated upon activation, normal T cell-expressed and secreted (RANTES) are widely associated with viral infections [74, 76] During intranasal influenza virus infection in mice, CCL2/monocyte chemotactic protein-1 (MCP-1) is detected in the lungs at various time-points post-infection, whereas other chemokines, including CCL3 and CCL5, are not expressed.[77] On the other hand, respiratory syncytial virus-infected mice display high levels of expression of numerous Carnitine palmitoyltransferase II chemokines in the lungs, including CCL3 and CCL5.[78] Among flaviviruses, CC chemokine receptors play an important role in leucocyte recruitment to the central nervous system.[79] Besides a deleterious pro-inflammatory role that CC chemokines could play in central nervous system, a well-studied example involves acute infection by West Nile virus in mice, in which the lack of CCR2 and CCR5 leads to decreased leucocyte recruitment, increased viral load in the central nervous system and enhanced mortality. West Nile virus infection induces high and continuous levels of CCL2 and CCL5, which are required for the local accumulation of NK cells, macrophages and T lymphocytes to control infection.

506). For SAP, C. albicans from NDOC showed the lower enzymatic activity (P < 0.001). There were no significant differences between isolates from HS and DOC (P = 0.7051). C. albicans isolates from NDOC and DOC patients showed an increased production of PL. "
“Candidaemia remains a relevant challenge in everyday patient care on intensive care units and general wards. Delays to adequate treatment

increase mortality rates and institutional standard operating procedures facilitate optimal treatment. A positive blood culture requires immediate treatment. Echinocandins are the first-line drugs AZD6738 of choice. Indwelling catheters have to be removed if feasible. Daily blood cultures until persistently negative exclude ongoing fungaemia. In case of Candida parapsilosis antifungal therapy should be switched to intravenous fluconazole. After 10 days of intravenous either echinocandin or fluconazole treatment, step-down to oral application of fluconazole simplifies antifungal therapy. Depending on organ involvement and clinical presentation of the patient antifungal treatment should be continued for at least 14 days after the last positive blood culture. We present our institutional management algorithm for candidaemia which is based on current guidelines and recommendations to improve patient outcome. “
“We prospectively observed 36 haematological

patients with mucormycosis from nine hospitals of St. Petersburg during 2004–2013. The most Tanespimycin manufacturer frequent underlying diseases were acute leukaemia (64%), and main risk factors were prolonged neutropenia (92%) and lymphocytopenia (86%). In 50% of the patients, mucormycosis was diagnosed 1–65 days after invasive aspergillosis. Main clinical form of mucormycosis was pulmonary (64%), while two or more organ involvement was noted

in 50% of the cases. The most frequent aetiological agents of mucormycosis were Rhizopus spp. (48%). Twelve-week survival rate was 50%. Combination therapy (echinocandins + amphotericin B forms) and recovery from the underlying disease significantly improved the survival rate. Mucormycosis (zygomycosis) is a severe opportunistic infection. At present, an increased frequency of mucormycosis is noted worldwide, particularly in patients with haematological malignancies. This is not only due to improvement of diagnostic methods for fungal infections, but rather because of more aggressive schemes of cytostatic therapy Rucaparib mw and more extensive use of haematopoietic stem cell transplantation. The range of underlying conditions in mucormycosis has changed. In the period 1980–1990, mucormycosis predominantly had developed in patients with decompensated diabetes mellitus. Over the last years, mucormycosis most frequently has been diagnosed in patients with haematological malignancies.[1, 2] We represent a clinical case of successful treatment of mucormycosis in a patient with acute myeloid leukaemia (AML), along with results of a prospective study of mucormycosis in haematological patients in St.

1d. There was no statistically significant difference in CCL2 liver expression between cirrhotic patients [4·4 102 (26·5-1·1 104) mRNA copies/106 copies HPRT; n = 62] and those without cirrhosis [2·4 102 (3·5-3·1 103) mRNA copies/106 copies LY2606368 clinical trial HPRT; n = 12] (P = 0·071). Liver CCL2 mRNA expression also showed an association with parameters of disease severity (Table 2b). We studied plasma levels and hepatic CCL2 expression according to short-term prognosis defined by 90-day survival. We did not find higher plasma levels in patients who died within 90 days [2·1 102 (90·5–1·6

103) pg/ml; n = 12] compared to those who survived [2·3 102 (20·4-1·4 103) pg/ml; n = 79] (P = 0·769). Nor was CCL2 liver expression higher in patients HDAC inhibitor who died within 90 days [3·5 102 (38·6-1·1 104) mRNA copies/106 copies HPRT; n = 11] than in those who survived [3·1 102 (3·5–4·3 103) mRNA copies/106 copies HPRT; n = 51] (P = 0·950). We sought to determine whether steroid therapy reduces CCL2 plasma levels, and we showed a trend towards decreased CCL2 plasma levels after 7 days of treatment (P = 0·056) (Supplementary

Fig. S1). To further unravel the role of CCL2 in the pathogenesis of ALD, we quantified inflammatory infiltrates of liver biopsy for which we had performed qRT–PCR for CCL2 (n = 74) (Fig. 2). Liver CCL2 mRNA levels in ALD patients were correlated specifically with neutrophil infiltrates (r = 0·411; P < 0·005), Fig. 3a, but neither with T lymphocyte nor with mononuclear cell infiltrates [(r = 0·226;

P = 0·058) and (r = −0·229; P = 0·055), respectively]. Moreover, we showed that liver CCL2 mRNA expression was correlated highly with liver IL-8 mRNA levels (r = 0·895; P < 0·001), Fig. 3b. As expected, IL-8 mRNA levels were correlated with neutrophil infiltration (r = 0·446; P = 0·002), Fig. 3c. To determine whether CCL2 plays a role in neutrophil recruitment, we analysed circulating neutrophils of ALD patients (alcoholic cirrhosis with or without AH) by flow cytometry and we found that these cells Flavopiridol (Alvocidib) did not express CCR2, Fig. 4. Because T helper type 17 (Th-17) cells play a role in neutrophil recruitment and express CCR2 [22], we evaluated, by immunohistochemistry, liver expression of IL-17 in patients for whom we had performed quantification of liver CCL2 mRNA. We found that CCL2 liver expression was associated with the number of IL-17+ cells (r = 0·339; P = 0·013). Moreover, Il-17+ cell infiltrates were correlated strongly with neutrophil infiltrates (r = 0·715; P < 0·001) and with IL-8 liver expression (r = 0·346; P = 0·038). CCL2 mRNA liver expression was not correlated with the degree of steatosis (r = 0·057 P = 0·637). We performed −2518 A > G CCL2 genotyping in 235 patients with ALD (109 cirrhosis without AH, 84 cirrhosis with AH, 13 steatofibrosis with AH and 29 steatofibrosis) and in 224 healthy controls.

Engagement of surface Compound Library datasheet receptors other than TCR also contributes to the induction or regulation of Vγ9Vδ2 T-cell responses. Vγ9Vδ2 T cells express several types of NK cell receptors (NKRs), which can increase (i.e. NKR-P1A) or decrease (i.e. ILT2 or NKG2A/CD94) TCR activation 21–23, whereas other surface receptors (i.e. CD16, CD224) can directly trigger a Vγ9Vδ2 T-cell response

24, 25. NKG2D, a homodimeric C-type lectin-like receptor, has this ability to directly trigger Vγ9Vδ2 T-cell functions 26. Particularly, anti-tumor cytotoxic activity of Vγ9Vδ2 T cells is triggered and regulated not only upon TCR-dependent antigen recognition but also through the ligation of NKG2D by its ligands 27. BGB324 purchase In humans, the ligands of NKG2D are the MHC class I-related chain proteins A and B (MICA/B) and UL16-binding proteins (ULBP) 1–4. Their expression is induced or upregulated on tumor and virus-infected cells 28. Nevertheless, while evidence of NKG2D contribution has been demonstrated in the anti-tumor response of Vγ9Vδ2 T cells 27, no direct role in their anti-infectious response has yet been reported. Moreover, the upregulation of MICA at the surface of Mycobacterium tuberculosis-infected DCs results

in a significant increase of the TCR-dependent Vγ9Vδ2 T-cell effector functions 29. However, the role played by the interaction of NKG2D with its ligands during anti-infectious activity of Vγ9Vδ2 T cells remains to be clarified. To further address this issue, we analyzed the role of NKG2D recruitment in a bacterial infection model. First, we demonstrated that NKG2D ligands (such as ULBP1 and ULBP2) trigger TNF-α and IFN-γ production and the release of lytic granules through their interaction with NKG2D and the triggering Cepharanthine of PI3K-dependent

intracellular signaling pathways. Moreover, concomitant TCR and NKG2D engagement led to stronger effector functions of Vγ9Vδ2 T cells. In vitro, the impairment of NKG2D recruitment decreases the anti-infectious activity of Vγ9Vδ2 T cells, leading to a higher development of Brucella in infected macrophages. ULBP1 is the main NKG2D ligand expressed by Brucella-infected macrophages and is involved in the impairment of intramacrophagic bacterial development. Altogether, these results provide evidence that NKG2D and its ligands are involved, at least in part, in the anti-infectious effect of Vγ9Vδ2 T cells. To study the role of NKG2D in Vγ9Vδ2 T-cell functions, we used two fusion proteins ULBP1-leucine zipper (LZ) and ULBP2-LZ composed of the ULBP1 or ULBP2 soluble part (two NKG2D ligands) and a LZ domain previously described in 30. While UL16-LZ, a control protein that is not a NKG2D ligand shows no detectable binding to Vγ9Vδ2 T cells, ULBP1-LZ and ULBP2-LZ do. This interaction is completely blocked by the presence of a blocking anti-NKG2D mAb (M585) (Fig. 1A).

Successful fluorochrome incorporation was confirmed by native polyacrylamide gel electrophoresis generating a single band at about 600 kDa corresponding to toxin A protein dimer [29, 30] under non-denaturing

conditions, which exhibited fluorescence during illumination with UV light. Toxin A488 was also shown to induce morphological changes in Vero cells and Caco-2 cells identical to that seen for unlabelled toxin A (treated as per labelled toxin A without the addition of selleck the label), confirming that labelling had not compromised receptor-binding ability. To confirm that fluorescence in flow cytometry was because of toxin A488 only, without any contribution from free label that may have either not been removed following the labelling procedure or become detached from the toxin during storage or binding studies, toxin A488 was preincubated with PCG-4-conjugated beads prior to incubation with Caco-2 cells. A complete loss of Caco-2 cell-associated fluorescence was seen after incubation with the toxin A-depleted preparation (Fig. 1), confirming that all fluorescence was toxin A specific. In initial studies, following incubation of PBMNCs with toxin A488 at 37 °C for up to 24 h, monocytes were distinguished from lymphocytes by their forward- and side-scatter characteristics. In contrast to toxin A488-exposed lymphocytes, toxin

A488-exposed monocytes showed significant fluorescence at all time points up to 5 h, with Bumetanide a peak at 1 h (Fig. 2A). Drop

in monocyte-associated fluorescence from 1 h onwards after exposure to check details toxin A488 was associated with loss of events in the monocyte gate (Fig. 2B). The fluorescence level of toxin A488-exposed lymphocytes remained low, with no significant change (compared with control lymphocytes) over the 24 h period of study (Fig. 2A). Thus, at 24 h, there was no significant difference in fluorescence between lymphocytes incubated (at 37 °C) in control medium, compared with those cultured with toxin A488. In contrast to monocytes, the number of events in the lymphocyte gate (in toxin A488-exposed PBMNCs) did not change significantly from cells exposed to control medium over the 24 h period of study (Fig. 2B). When studied after 48-h incubation at 37 °C, fluorescence of toxin A488-exposed lymphocytes was marginally, but significantly greater than lymphocytes cultured with control medium (5.35 versus 4.97; P < 0.01). By contrast, following incubation at 4 °C, the difference in fluorescence between toxin A488-exposed and control lymphocytes fell short of statistical significance (5.0 versus 4.85; P = 0.07). The ability of trypan blue to quench fluorescence of monocytes exposed to toxin A488 at 37 and 4 °C was subsequently investigated. Initial studies, using PBMNCs labelled with anti-CD45 antibody, followed by labelling with Alexa Fluor 488-conjugated anti-mouse antibody, showed that trypan blue quenched 87.27 (±4.7)% of cell surface–associated fluorescence.

We previously demonstrated that IC negatively regulates TLR4-triggered inflammatory

response in macrophages through FcγRIIb 27. We here demonstrate that in the presence of IC, FcγRIIb overexpression promotes resistance of immature DCs to TLR-triggered maturation induction and also increases DC tolerogenecity. Accordingly, IC-stimulated, FcγRIIb-overexpressing DCs (DC-FcγRIIb) can downregulate immune response more significantly both in vitro and in vivo, thus attenuating the progression of disease in lupus-prone mice. To investigate whether IC/Ig could inhibit TLR-induced maturation of DCs via FcγRIIb, Trametinib datasheet immature DCs derived from WT or FcγRIIb−/− mice were incubated with IC (OVA plus anti-OVA)/Ig (anti-OVA) for 24 h before these DCs were stimulated with LPS or CpG ODN for another 24 h. As for WT DCs, IC alone (whereas not Ig) slightly upregulated the expression of I-Ab, CD40, CD80

and CD86. Interestingly, IC pretreatment significantly inhibited the LPS or CpG ODN-induced upregulation www.selleckchem.com/products/GDC-0980-RG7422.html of I-Ab, CD40, CD80 and CD86 expression, and Ig pretreatment significantly inhibited the LPS-induced upregulation of the four molecules and the CpG ODN-induced upregulation of CD80 and CD86 expressions (Fig. 1A). In contrast, neither IC nor Ig pretreatment had significantly inhibitory effect on the LPS or CpG ODN-induced upregulation of I-Ab, CD40, CD80 and CD86 expression on FcγRIIb−/− DCs (Fig. 1A). These data indicate that FcγRIIb mediates the inhibitory effect of IC/Ig above. IC alone could stimulate WT DCs as well as FcγRIIb−/− DCs to secrete TNF-α and IL-1β to some degree; however, IC pretreatment obviously suppressed LPS or CpG ODN-induced TNF-α and IL-1β secretion from WT DCs (Fig. 1B). In contrast, IC pretreatment could not suppress TNF-α and IL-1β secretion by LPS-stimulated FcγRIIb−/− DCs, and even promoted TNF-α and IL-1β secretion by CpG ODN-stimulated

FcγRIIb−/− DCs (Fig. 1B). OVA alone had no effect on the phenotype and cytokine Thiamine-diphosphate kinase secretion of WT DCs and FcγRIIb−/− DCs with or without stimulation with LPS, CpG ODN (data not shown). Considering that the Ig (anti-OVA mAb) we used, also exhibited inhibitory effect on LPS-induced DC maturation (in a similar manner to IC), we speculated that anti-OVA mAb might have some aggregated Ig, because aggregated Ig has a higher binding affinity to FcγRIIb than monomeric Ig. Therefore, we compared the differences between monomeric and aggregated Ig. As expected, the aggregated Ig significantly inhibited WT DCs to express I-Ab and CD40 and to secrete TNF-α in response to LPS stimulation, whereas monomeric Ig did not. However, neither aggregated Ig nor monomeric Ig had such inhibitory effect on LPS-induced FcγRIIb−/− DC maturation (Supporting Information Fig.

The elevations were more modest (<1.5× upper normal EGFR inhibitor values here vs. 2- to 3-fold previously), not associated with symptoms, and not notably dose-related. We speculate that some bacteria may translocate the intestinal wall and be transported systemically, but at too low a level to generate strong systemic immune responses or be detected in blood cultures. No subject receiving BMB54 had abnormal transaminases, suggesting that as demonstrated in vitro (6), this organism may have decreased tropism for hepatic cells in humans. Other published murine studies in which the BMB54 parent strain vs. WT organisms

were injected i.v. showed that transaminases peaked approximately one and four days after intravenous administration, respectively (6). In that study, the BMB54 mutant caused much lower peak transaminase values, likely because of the lack of replication within the liver. After intravenous administration of a prfA-defective L. monocytogenes vaccine strain to humans, 1.5- to 3.5-fold elevations in both GGT and transaminases were reported eight days after administration, but these tests were apparently not performed during days 1 through 7 after i.v. administration (10). No clinical data suggest these transaminase elevations are harbingers of prolonged or serious hepatic dysfunction.

One murine cancer immunotherapy study using an inlB-positive L. monocytogenes Venetoclax solubility dmso strain exploited this hepatotropism. Hepatic metastases were more efficiently eliminated and survival was prolonged when attenuated L. monocytogenes were used as adjuvant/adjunctive therapy for an irradiated tumor cell vaccine expressing granulocyte-macrophage colony-stimulating factor (36), though that study did not include a comparator strain lacking inlB with decreased

liver tropism. We undertook this study to evaluate the physiological effect of two L. monocytogenes organisms as vectors for delivery of viral antigens. Oral delivery was hoped to stimulate or at least “prime” the mucosal immune system, as many viruses enter through mucosal sites. Bulk IFN-γ ELISpot studies performed on freshly isolated PBMC were chosen as a simple, for reproducible measure of systemic cellular immunity increasingly used in studies of viral vaccines. Our earlier human study was limited by a lack of immunological reagents, especially peptide reagents for ELISpots. Here we were able to test synthesized overlapping peptide pools for both listeriolysin and the foreign antigen. A recent study of PBMC derived from approximately 80 healthy human donors showed that bulk IFN-γ ELISpot responses to this same listeriolysin peptide pool also correlated in magnitude and incidence with IL-2 ELISpot responses to the pool (35), so this is likely a reasonable measure of cellular immunogenicity.

In addition, we note that sensitization alone, without adoptive transfer of iNKT cells, induces a partial but significant reconstitution of CS in Epigenetics inhibitor comparison with baseline, suggesting that iNKT cell–independent pathways may also exist (Groups B and E, Fig. 4A). We next asked whether CS is dependent upon any other trait of the hepatic environment other than CD1d-expressing cells. We explored the possibility of peripheral activation of iNKT cells following adoptive transfer. We investigated whether transferred hepatic iNKT cells exhibit tropism to

the livers of the recipient mice and again tested whether this might be dependent upon hepatocyte CD1d expression. We transferred activated iNKT cells into sensitized Jα18−/− and CD1d−/− mice (as mentioned earlier) and monitored by flow cytometry the percentage of hepatic T cells that were iNKT cells 1 day later. (This is the time point at which mice are challenged on the ears after adoptive transfer in our protocol.) We compared this to the percentage of iNKT cells in wild-type BALB/c mice, in which NKT cells comprised approximately 70% of hepatic T cells. In contrast, there is no evidence of re-population of donor iNKT cells into recipient livers: iNKT cells constituted <1% of total hepatic T cells in both iNKT cell–deficient strains following adoptive transfer (Fig. 4B). Staurosporine order Had donor iNKT cells migrated

to recipient livers, and if this had been dependent upon hepatocyte

CD1d expression, then a difference would have been seen between the Jα18−/− and CD1d−/− mice. Furthermore, there does not appear to be any essential component of the hepatic environment other than CD1d-expressing cells, as the result was equivalent in Jα18−/− and CD1d−/− mice following adoptive cell transfer. Although this experiment demonstrates that peripheral hepatocyte-independent activation of iNKT cells may Urocanase occur, it remains unclear whether the suggestion of extrahepatic iNKT cell activation via CD1d–lipid complexes is merely an artefact of the artificial experimental design or whether this finding is relevant to wild-type mice. It is clear that reconstituted iNKT cell–deficient mice, despite their equivalent CS reactions, differ in the distribution of iNKT cells. The livers of reconstituted mice are not equivalent to those of wild-type mice. Certainly, in wild-type mice, iNKT cells may interact with hepatocytes via CD1d; we simply show here that such an interaction is not critical in mounting a full CS reaction. We demonstrate here that soon after contact sensitization, stimulatory lipids accumulate in the liver and facilitate the activation of iNKT cells in a CD1d-dependent manner. Remarkably, a significant increase in stimulatory capacity was seen within 30 min of sensitization.

2 cells, using protein G columns according to standard protocols. Soluble TNFR1 fusion protein (sTNFR1-Ig) was a kind gift from Geoff Hale (Therapeutic Antibody Group, University of Oxford, UK). All fluorochrome-conjugated anti-mouse mAbs and secondary detection reagents used were purchased from BD Biosciences (Oxford, UK). Biotinylated anti-CD3ζ was from Upstate (Watford, UK), and purified polyclonal rabbit anti-mouse EP1, EP2, EP3 and EP4 were from Cayman Chemicals (Ann Arbor, MI). Bone marrow (BM) Mϕ were generated using a method adapted from Munder et al.21 Briefly, bone marrow cells were resuspended at 5 × 105

cells/ml see more in complete media supplemented with 5% v/v horse serum (Invitrogen), and 50 pg/ml macrophage colony-stimulating factor. The cell suspension was transferred to hydrophobic PTFE-coated tissue culture

bags (supplied by Dr M. Munder, University of Heidelberg, Heidelberg, Germany) and incubated for 8 days at 37° in 5% v/v CO2. Single-cell splenocyte suspensions were generated by grinding spleens through a 70-μm cell strainer (BD Biosciences) with a syringe plunger. When used as APCs, splenocytes were irradiated with 3000 Rads using a caesuim-137 source (Gravatom, Hants, UK). The OT-II CD4+ T cells were prepared by enriching CD4+ cells from single cell suspensions of C57BL/6 OT-II splenocytes, using anti-CD4 microbeads (Miltenyi Biotech, Bisley, UK) according to the manufacturer’s instructions. B cells were prepared from spleens using anti-B220 microbeads (Miltenyi Biotech). Doxorubicin chemical structure Dendritic cells were generated from cultures of bone marrow cells as previously described.22 The 1 × 105 APCs were co-cultured with CD4+ T cells at ratio of 1 : 1 in round-bottom 96-well plates in complete media. The OVA peptide was added at the indicated concentrations. To some cultures the arginine analogue, l-NG-monomethyl arginine, the NO donor S-nitroso-N-acetyl-l,l-penicillamine,

or the cyclo-oxygenase (COX) inhibitor indomethacin (all from Sigma) was added. In some experiments, recombinant IFN-γ (Peprotech, London, UK), or PGE2 (Sigma) was added. Cells were cultured in a humidified Amoxicillin environment at 37°, 5% v/v CO2. Proliferation was measured by pulsing with 18·5 kBq [3H]thymidine (GE Healthcare, Bucks, UK) per well for the final 8 hr of culture and determining thymidine uptake [measured in counts per minute (c.p.m.)]. Accumulated NO production was measured after 64 hr in culture supernatants using Griess reagent (Sigma) as previously described.23 Production of IFN-γ was assessed using a murine T helper type 1 (Th1)/Th2 Flow cytomix 10plex kit (Bender Medsystems, Vienna, Austria) according to the manufacturer’s instructions. Concentration of PGE2 was measured using an enzyme immunoassay competition enzyme-linked immunosorbent assay kit (Caymen Chemical, Ann Arbor, MI) according to the manufacturer’s instructions.

, 2004; Nobile et al., 2006, 2008). Candida complement receptor 3-related protein (CR3-RP) has been described to be a ‘mimicry’ antigen functionally comparative with the human CR3 protein expressed RXDX-106 concentration in neutrophils, macrophages and monocytes, with the ability to bind human complement fragment iC3b (Gilmore et

al., 1988; Hostetter et al., 1990; Hostetter, 1996). The human CR3 antigen can be detected via the monoclonal antibody (mAb) OKM1, which recognizes the α chain of CR3 and CD11b (Wright et al., 1983), but also cross-reacts with Candida CR3-RP (Heidenreich & Dierich, 1985; Bujdákováet al., 1997, 1999). The sequence of this antigen contains the DINGGG motif, which is characteristic of proteins belonging to the DING family (Bujdákováet al., 2008). This motif has already been mentioned in prokaryotic as well as in high eukaryotic organisms (Berna et al., 2009), but not in eukaryotic microorganisms. The CR3-RP has been recently reported to be a surface antigen participating in adherence to buccal epithelial cells as well as in in vitro biofilms. Moreover, Saracatinib chemical structure the immunomodulation properties of CR3-RP and the novel CR3-RP glycoconjugate effectively triggered an enhancement of immune responsiveness in the rabbit model (Bujdákováet al., 2008; Paulovičováet al., 2008). While many reports have reviewed the antifungal susceptibility/resistance of C. albicans in a mature biofilm (Henriques et al., 2005;

Seidler et al., 2006) only a few have mentioned inhibition during the adherence phase using antifungals or antibodies (Rodier et al., 2003; Cateau et al., 2007; Dorocka-Bobkowska et al., 2009; Maza et al., 2009). The lack of information about adherence and the possibility of decreasing biofilm production via a reduction in C. albicans adherence capability in the first stage of biofilm development was our motivation for searching the answer to two questions: (1) can a decrease in adherence (the first biofilm stage) affect the quantity of a mature biofilm? and (2) can blocking the C. albicans CR3-RP surface antigen by antibodies contribute Meloxicam significantly to a reduction in adherence during biofilm formation? In this study, the standard

C. albicans strain was used (CCY 29-3-162 from the CCY Culture Collection of Yeasts, Chemical Institute, Slovak Academy of Sciences, Slovakia), originally recovered from a patient with mycotic colpitis. This strain was selected because of its high CR3-RP expression (Bujdákováet al., 1997). For comparison, the clinical isolate C. albicans with a high ability to form biofilm obtained from the urinary catheter of a patient with candidiasis was tested. Different antibodies were applied: polyclonal anti-CR3-RP antibody, prepared as described by Bujdákováet al. (2008) and OKM1 mAb (hybridoma cell culture ATCC, CRL-8026), purchased as previously described by Bujdákováet al. (1999). Titers of the antibodies were determined by enzyme-linked immunosorbent assay (ELISA) in 96-well plates (Sarstedt, Germany) (Voller, 1978).