In contrast, DP thymocytes that express a TCR specific for a self-antigenic peptide are negatively selected and die via apoptosis. To examine the effect of LAR deficiency on negative and positive selection, we crossed transgenic mice that carry a transgene encoding a TCR that recognizes male-specific peptides presented on H-2Db

molecules (HY-TCR-Tg mice) 21 with LAR−/− mice and compared the CD4/CD8 profile of LAR−/−HY-TCR-Tg mice with that of LAR+/+HY-TCR-Tg mice. In normal female HY-TCR-Tg mice, the differentiation of DP thymocytes is skewed toward CD8SP cells by positive selection. In LAR−/−HY-TCR-Tg female mice, thymocyte differentiation was less skewed toward CD8SP compared with normal HY-TCR transgenic female mice (Fig. 4). The https://www.selleckchem.com/products/PD-0332991.html average ratio of CD8SP thymocytes to CD4SP thymocytes was 2.31±1.01 and 1.54±0.61 in WT and LAR−/−HY-TCR-Tg female mice, respectively (p=0.04). In the periphery, the ratios of CD8/CD4 T cells in WT and HY-TCR-Tg

mice with or without the LAR−/− mutation were similar (Supporting Information Fig. 5) 6; the lack of difference in these ratios may be due to peripheral homeostatic mechanisms that compensate for LAR deficiency. In contrast to HY-TCR-Tg female mice, most DP thymocytes in the transgenic male mice die via apoptosis selleck products because the HY-TCR interacts with male peptides presented on H-2Db molecules expressed on thymic antigen-presenting cells during negative selection. Thus, the percentage of DP, CD4SP and CD8SP thymocytes was reduced. As shown

in Fig. 5, the sum of the DP, CD4SP and CD8SP thymocyte percentages from LAR−/−HY-TCR-Tg male mice was higher than the sum from normal HY-TCR-Tg male mice (p<0.01). Taken together, these results indicate that LAR deficiency may affect both the positive and negative selection of thymocytes. Next, we examined the effect of LAR deficiency on TCR-mediated thymocyte activation. Specifically, we examined the effect of LAR deficiency on GPX6 the alteration of the intracellular Ca2+ concentration that was induced following TCR-mediated stimulation. Thymocytes from WT and LAR−/− mice were loaded with a Ca2+ indicator, Fluo4 and stimulated with a CD3-specific antibody together with a hamster IgG-specific antibody. The intensity of Fluo-4 fluorescence was measured by flow cytometry following stimulation. The intensity increased after stimulation, reached a peak within 2–3 min and then decreased gradually in both groups of mice. However, compared with WT mice, the population that responded was significantly lower in thymocytes from LAR−/− mice (p<0.05) (Fig. 6). These results suggest that LAR is involved in TCR signaling in thymocytes. We also examined the effect of LAR deficiency on the proliferation of thymocytes following stimulation with CD3- and CD28-specific antibodies. The level of thymocyte proliferation was similar in both groups (Supporting Information Fig. 6).

The aim of this study was to determine the efficacy of intragraft inhibition of CIITA in attenuating liver transplant rejection. Three plasmids

containing small hairpin RNA (shRNA) against rat CIITA (pCIITA-shRNA) and one control plasmid of pHK-shRNA were constructed. In vitro dendritic cell (DC) transfection and liver transfection via portal vein in donor rats (n = 8) by shRNA plasmids were performed to confirm the inhibitory effect of pCIITA-shRNA on CIITA expression. It showed SAHA HDAC clinical trial that expressions of CIITA and MHC-II were significantly inhibited by pCIITA-shRNA in both DC in vitro and liver of donor rats in vivo (p < 0.05 vs. control pHK-shRNA treatment). pCIITA1-shRNA was proved to be the best inhibitor among three pCIITA-shRNAs and then used in high-responder rat liver transplantation model (DA donors-to-Lewis recipients). Transplant groups (n = 16/group) include untreated recipients transplanted with donor liver graft pretreated with either saline, or pHK-shRNA, or pCIITA1-shRNA. Cyclosporine-treated (10 mg/kg, im, day 0–7) recipients transplanted with unmodified liver grafts were used as no rejection control. The results showed that the recipient rats survived significantly longer in pCIITA1-shRNA-treated group with markedly attenuated liver graft rejection (p < 0.05 vs. saline and pHK-shRNA-treated groups). Furthermore,

significantly decreased intragraft expressions of CIITA, MHC-II, IL-2, and IFN-γ were found in pCIITA1-shRNA-treated group (p < 0.05 vs. saline Omipalisib clinical trial and pHK-shRNA-treated groups). This study suggests that intragraft inhibition of CIITA could be a novel strategy for attenuating graft rejection in liver transplantation. © 2014 Wiley Periodicals, Inc. Microsurgery, Bumetanide 2014. “
“Reconstruction of limb-threatening lower extremity defects presents unique challenges. The selected method must provide adequate coverage of exposed bone, joints, and tendons while maximizing function of the limb. The traditional workhorse flaps, the free latissimus

dorsi and rectus abdominis flaps, have been associated with donor site morbidity and bulkiness that can impair rehabilitation. We report a case series (n = 18) in which the free serratus anterior muscle flap and split thickness skin graft (STSG) was used for lower limb soft tissue coverage. Injuries were due to diabetes (9/18), trauma (7/18), and chronic venous stasis (2/18). A 94% flap survival rate was observed and all but one patient was ambulatory. No donor site morbidity was reported. Our series demonstrates that serratus anterior is an advantageous, reliable free flap with minimal donor site morbidity. © 2013 Wiley Periodicals, Inc. Microsurgery 34:183–187, 2014. “
“Microvascular free flaps continue to revolutionize coverage options in head and neck reconstruction.

For example, it was reported that pro-IL-16 suppresses Skp2 transcription by recruiting histone deacetylase 3 to the Skp2 promoter through interaction with a GA-binding protein [41]. Furthermore, HSC70, a chaperone for NF-κB, was identified as binding partner of pro-IL-16 via the PDZ domain [42]. In the study of Fujihara and Nadler, they reported that pro-IL-16 has

a nuclear localization sequence, and its PDZ domain acts not only as a nuclear scaffolding protein, but also functions as a nuclear chaperone to transport essential nuclear complex members with a role in transcriptional suppression into the nucleus. It was recently reported that HSC70 knockdown led to loss of nuclear translocation

by pro-IL-16 in T lymphocytes. More interestingly, loss of nuclear pro-IL-16 led Ponatinib solubility dmso subsequent increase in Skp2 level and decrease in p27kip, which ultimately enhanced T cell proliferation to facilitate the T cell transformation [43]. We initially hypothesized that pro-IL-16 would have a similar function in resting B cells as T lymphocytes, and that cell-cycle progression and proliferation would be inversely correlated with the level of pro-IL-16 in the nucleus. We therefore investigated the effects of pro-IL-16 on cellular signalling in resting B cells. Our western blot results revealed that pro-IL-16, rather than mature IL-16, LDE225 in vivo is the main form of IL-16 present in resting B cells; we assumed that the mature form was secreted as soon as it had been processed by caspase-3 (Fig. 1C). Pro-IL-16 was found both in the cytoplasm and nucleus (Fig. 2). Because pro-IL-16 was Exoribonuclease identified from immunoprecipitates using an anti-MHC class II antibody, this implies

that it is associated with MHC class II molecules, and we confirmed this assumption by Western blot analysis and confocal laser scanning microscopy (Figs 1B and 2B). More importantly, the nuclear level of pro-IL-16 was increased by treatment of cells with the corresponding anti-MHC class II antibody, consistent with the observation that the expression of pro-IL-16 is inhibited in activated T cells (Fig. 2A) [44, 45]. To confirm this inverse relationship between pro-IL-16 and B cell proliferation, we transfected pro-IL-16 cDNA into 38B9 cells and found that overexpression of pro-IL-16 suppressed B cell proliferation (Fig. 3A) and that the suppression was mediated by inhibition of the nuclear translocation of NF-κB subfamilies, p50, p52 and c-Rel (Fig. 3B). Our finding that p50, p52 and c-Rel are involved in pro-IL-16-mediated suppression of resting B cell proliferation is consistent with our previous observations that MHC class II-mediated negative signalling in resting B cell activation is closely related to the activation of the p50, p52 and c-Rel NF-κB subfamilies [16, 17].

Pharmacological inhibition of Uba1, levels of which are robustly reduced in SMA, was sufficient to induce accumulation of UCHL1 in primary neuronal cultures. Pharmacological inhibition of UCHL1 exacerbates rather than ameliorates disease

symptoms in a mouse model of SMA. Thus, pharmacological inhibition of UCHL1 is not a viable therapeutic target for SMA. Moreover, increased levels of UCHL1 in SMA likely represent a downstream consequence of decreased Uba1 levels, indicative of an attempted supportive compensatory response to defects in ubiquitin homeostasis caused by low levels of SMN protein. “
“Histone deacetylase 6 (HDAC6) plays a crucial role in aggresome formation, resulting in the clearance of misfolded proteins. Previous studies have shown that HDAC6 is concentrated in Lewy bodies (LBs) in Parkinson’s disease (PD) and dementia with LBs (DLB) (Cell 115: 727–738, 2003). We performed immunohistochemical and ultrastructural RGFP966 purchase investigations on the brains of patients Hormones antagonist with various neurodegenerative disorders. Anti-HDAC6 antibody faintly immunostained the cytoplasm of neuronal and glial cells in control subjects. In PD and DLB, almost all of the cortical, brainstem-type and peripheral LBs were intensely immunolabeled with anti-HDAC6. In multiple system atrophy (MSA), the vast majority of glial cytoplasmic inclusions (GCIs) were also positive for

HDAC6. Immunoelectron microscopy revealed that the reaction product was localized to the filamentous structures in LBs and GCIs.

Various neuronal and glial inclusions in neurodegenerative disorders other than LB disease and MSA were HDAC6-negative. These findings suggest that accumulation of HDAC6 is specific to α-synucleinopathy and that both LBs and GCIs may represent cytoprotective responses to sequester toxic proteins. “
“Motor neuron Cobimetinib price diseases, including amyotrophic lateral sclerosis (ALS), are devastating disorders and effective therapies have not yet been established. One of the reasons for this lack of therapeutics, especially in sporadic ALS (SALS), is attributed to the absence of excellent disease models reflecting its pathology. For this purpose, identifying important key molecules for ALS pathomechanisms and developing disease models is crucial, and omics approaches, including genomics, transcriptomics and proteomics, have been employed. In particular, transcriptome analysis using cDNA microarray is the most popular omics approach and we have previously identified dynactin-1 as an important molecule downregulated in the motor neurons of SALS patients from the early stage of the disease. Dynactin-1 is also known as a causative gene in familial ALS (FALS). Dynactin-1 is a major component of the dynein/dynactin motor protein complex functioning in retrograde axonal transport. In motor neuron diseases as well as other neurodegenerative diseases, the role of axonal transport dysfunction in their pathogenesis always draws attention, but its precise mechanisms remain to be fully elucidated.

The number of individuals without CCL3L or CCL4L is always below 5% in all continental regions [52,53]. The duplicated region encoding human CCL3L–CCL4L genes has an ancestral correlate in non-human primates. The CCL3L–CCL4L copy numbers are much higher in non-human primates than in human populations [53–55]. Gonzalez et al. determined the gene copy numbers of the chimpanzee (Pan troglodytes) CCL3L DNA Methyltransferas inhibitor orthologues from 83 animals. The CCL3L copies range from 6 to 17 per diploid genome (median 9; mean 9·3) [53]. Similarly, Degenhardt et al. observed extensive variation in copy number of the CCL3L region among 57 samples of rhesus macaque (Macaca mulatta):

copy number estimates range from 5 to 31 copies per diploid genome (median 10; mean 11·1) [54]. Currently, the official symbols of the genes included in the CCL3L–CCL4L cluster are based on the public human genome sequence which contains, by chance, three CCL3L copies and two CCL4L copies. CCL3L and CCL4L have been numbered based on their position from the more centromeric

BVD-523 manufacturer to the more telomeric. Thus the official symbols for CCL3L genes are CCL3L1 (GeneID: 6349), CCL3L2 (GeneID: 390788) and CCL3L3 (GeneID: 414062). The official symbols for CCL4L genes are CCL4L1 (GeneID: 9560) and CCL4L2 (GeneID: 388372). However, we believe that the nomenclature criterion should consider whether the genes are really different rather than solely their copy number. Although CCL3L1 and CCL3L3 are separate genes, both have three identical exons and encode identical proteins [42,47], and therefore they are denoted together here as CCL3L1 (Fig. 1). CCL3L2 (known previously as LD78γ or GOS19-3) was identified initially as a pseudogene, as it contains two exons that are homologous to exons 2 and 3 of the CCL3L1 Exoribonuclease gene and appeared to contain a 5′ truncation compared with CCL3L1[46].

However, Shostakovich-Koretskaya et al. recently identified novel 5′ exons for CCL3L2 which give rise to two alternatively spliced transcripts by bioinformatics and mRNA profiling (Fig. 1c) [51]. These alternatively transcribed mRNA species contain chemokine-like domains but are not predicted to encode classical chemokines (data not shown [51]). Regarding CCL4L genes, CCL4L1 and CCL4L2 share 100% sequence identity in the coding regions. However, a fixed mutation at the intron–exon boundary of some CCL4L genes results in the production of aberrantly spliced transcripts [48]. We proposed the name of the originally described gene (corresponding to GeneID: 388372) as CCL4L1 and CCL4L2 (GeneID: 9560) as the gene that contains the mutation at the intron–exon boundary [38,48,52,56]. We use this nomenclature in this review (view Fig. 1) and we note that the same concept has been applied recently by others [51].

We have demonstrated that early vaccination (at 7 days of life) with a live gE-deleted ADV vaccine, in the presence of high levels of MDA could be effective, but that the intensity and duration of the recall proliferative T-cell response depended on the moment of the second vaccination. Humoral as well cellular responses were most similar to results obtained in the group vaccinated following the manufacturer’s recommendation when the second vaccination was performed at 12 weeks of life. Future studies are required to evaluate the protective effects of vaccination with this protocol. Vaccination of pigs as young

as 7 days of age, from a practical point of view, could be more convenient for herd personnel. This work is supported by Project no. NN 308 275934 funded by Ministry MK-8669 manufacturer of Science and Higher

Education. The NIA-3 ADV strain was kindly provided by Dr Andrzej Lipowski from NVRI Pulawy. “
“The conventional acid fast AZD3965 bacilli (AFB) smear and Mycobacterium tuberculosis (M.tb) culture of pleural effusion and tuberculin skin test (TST) in tuberculous pleurisy are unable to meet clinical needs because of their low sensitivities and specificities. To evaluate the diagnostic accuracies of QuantiFERON TB Gold In-Tube test (QFT-GIT) and nested-PCR in tuberculous pleurisy, we conducted a cross-sectional study in regions of China with a high tuberculosis (TB) epidemic. Seventy-eight participants were enrolled: 58 TB patients with diagnosis of confirmed or probable tuberculous pleurisy and 20 non-TB patients with a diagnosis of other non-TB diseases. The positive rates of AFB smear and M.tb culture in the pleural effusion were 5.8% (2/42) and 10.6% (5/47), respectively. The sensitivity and specificity of QFT-GIT were 93.1% (54/58) and 90.0% (18/20), whereas those of TST were 68.5% (37/54) and 86.7% (13/15), respectively; the sensitivity of QFT-GIT was significantly higher NADPH-cytochrome-c2 reductase than TST (P = 0.013). The sensitivity and specificity of M.tb-specific nested-PCR in pleural effusion were 94.8% (55/58) and 90.0% (18/20), respectively, with a turnaround

time of 7 h. Furthermore, combined QFT-GIT and nested-PCR detection improves the specificity to 100% with a sensitivity of up to 90.0%. This combination of immunoassay and molecular detection holds promise for the clinical diagnosis of tuberculous pleurisy. Tuberculous pleurisy is the most common extrapulmonary tuberculosis (TB), accounting for c. 10–20% of all tuberculous patients and c. 10–30% of disease causing pleural effusions (Porcel, 2009). The conventional acid fast bacilli (AFB) smear and Mycobacterium tuberculosis (M.tb) culture in pleural effusion are unable to meet clinical needs because of their low sensitivities (Light, 2007). There is an overriding need for the development of highly sensitive, specific and rapid tools to aid in the diagnosis of tuberculous pleurisy.

The strongest response was induced by peptide 10–26 followed by peptides 289–306, 117–133/120–133, and 46–70, as determined by high levels of IFN-γ as well as the presence of IL-2 in culture supernatants (Fig. 2). The two peptides 117–133 and 120–133 led to a similar IFN-γ response, although the longer sequence selleck kinase inhibitor induced significantly more IL-2 (p=0.009). In addition, peptide 46–70 stimulated the production of higher amounts of IFN-γ and IL-2 compared to peptide 50–70 (Fig. 2), showing the importance of flanking residues for the induction of an optimal T-cell response. Of note, peptide 305–322, indicated as good binder to DR*0401 by TEPITOPE

(Table 1), did not bind in our assay (Fig. 1) nor did it induce a T-cell response in DR*0401-Tg mice (Fig. 2). Therefore, this peptide was not selected for analysis in RA patients. In conclusion, the four best binders to DR*0401, as determined by binding assays and TEPITOPE program (Table 1), were also the most

immunogenic ones in DR*0401-Tg mice (Fig. 2 and Table 1). We next assessed the potential of the selected peptides to induce production of IFN-γ in PBMC from RA patients and healthy individuals. Freshly isolated PBMC from 33 RA patients and 16 healthy controls were cultured with 13 individual hnRNP-A2 peptides (indicated in bold in Table Roscovitine 1) in ELISPOT plates pre-coated with an anti-IFN-γ mAb. In this assay, PBMC from 6 out of 33 (18%) patients showed an IFN-γ response to hnRNP-A2 peptides, five of them (15%) to a main determinant contained in peptide 117–133 (Fig. 3 and Table 2). The mean frequency of IFN-γ-producing cells specific for this dominant epitope was 21±9 out of 106 cells (mean/duplicate for each patient: 25, 15, 11, 20, 39, 15) compared to 2±2 out of 106 cells (3, 0, 0, 4, 5, 0) 3-mercaptopyruvate sulfurtransferase for the medium background. Remarkably, when retesting two of the six reactive patients 3 months after the first evaluation, the T-cell response to the peptide was sustained (Fig. 3 and Table 2). Conversely, PBMC from none of the healthy individuals reacted to hnRNP-A2

peptides (Table 2). Of note, T-cell reactivity to hnRNP-A2 peptides was independent of disease duration, which varied between 3 and 14 years, and immunosuppressive medication (Table 2 and Supporting Information Table 1). Importantly, all six patients with peptide reactivity presented with active disease (DAS28 > 3.2), and four out of five had bone erosions. We next thought to confirm these findings, to show that the responses to peptides 117/120–133 are mediated by CD4+ T cells, and to investigate whether they are selectively found in RA patients. To demonstrate MHC class II restriction, we incubated the cells with an anti-class II Ab together with peptides 117/120–133 and analyzed the proliferative response in 25 additional RA and 28 disease control (DC) patients with osteoarthritis (Supporting Information Table 2).

The other major advantage of ZFN is the speed of the procedure since KO rats can be generated

click here in about 4 months in both inbred and outbred strains 8, 9, 23. Finally, mutations are definitive and transmitted to the progeny. Our characterization of IgM KO and JH KO rats confirm the previous findings in μMT or J KO mice 11, 12 and immunodeficient human patients 24, 25 that the absence of membrane Ig expression results in the absence of B cells. On the contrary, IgM deletion 26 or truncation 13 in mice permitted expression of other heavy chains and allowed B-cell development and maturation due to replacement of IgM by IgD. Similarly to humans 24, 27, IgM KO rats showed only 5% of normal levels of BM pro–pre B cells, whereas μMT mice showed normal levels of BM pro–pre B cells 11. In this regard, deletion in mice of the Ig JH region resulted in a block of Ig gene expression and B-cell development selleck inhibitor at the pro-B-cell stage 12 as for JH KO rats. Thus, like μMT mice in which transcription and translation of μ-chain occurred but did not result in expression

of membrane-bound IgM and like JH KO mice, IgM KO rats showed a shortened μ transcript and the absence of Ig polypeptide production and therefore a very early B-cell block. As for mice and human cells, an enigma still persists

on how B-cell levels can be suppressed early or potentially, after rearrangement at the pre-BCR stage but before a fully functional μ polypeptide is expressed. An answer to this may be dependent on the level of early control of the IgH locus when chromatin is opening and antisense transcription will be initiated before D to J recombination 28. It is possible that strain-specific parameters as well as size and position of the removed or targeted region may determine the B-cell block. Another difference with IgM KO mice 11 is that these mice showed normal levels of IgA and absence of all other Ig isotypes 29, whereas IgM KO rats showed complete deficiency O-methylated flavonoid of all isotypes including IgA. Analogously to IgM KO rats, patients with deletion of the μ locus also result in the absence of Ig production for all isotypes including IgA 25. Since in contrast to mice, only 1% of cells recovered from the peritoneal cavity of rats are B cells 17, we did not analyze this compartment. In IgM or JH KO rats’ T-cell numbers in spleen but not in lymph nodes were decreased, as described for μMT mice 14, 15. In μMT mice, this was due to the lack of production of lymphotoxin α1β2 by B cells, required for CCL21 and stromal cell development, and as yet to be defined mechanism(s) for the promotion of T-cell numbers 14.

40 CDK4 and CDK6 were both induced upon CD3/CD28 costimulation. nIL-2 abrogated the up-regulation of CDK6, and partly inhibited CDK4 induction, while BMS-345541 and PS-1145 suppressed the induction of both kinases. Taken together, these results emphasize that an important effect of IKK activation on CDK4 and CDK6 expression relies on IL-2/IL-2R drug discovery signalling. However, as full CDK4 up-regulation requires the activation of IKK and IL-2 signalling, these data add new information about the mechanisms that govern CDK4 expression in human T cells. CDK2–cyclin E/A complexes are implicated in the

regulation of major processes governing the G1/S transition.5 In our experiments, CDK2 induction was detected in 24-hr costimulated cells, and was preserved in the presence of nIL-2, but abolished by BMS-345541 and PS-1145. We thus Epigenetics inhibitor conclude that, in activated T cells, CDK2

induction is independent of IL-2 signalling, and relies instead on IKK activation, which is a novel finding. To acquire catalytic activity, CDK2 must bind to cyclin E (G1/S phase transition) or cyclin A (S phase).5 We found that T-cell stimulation caused a significant increase in cyclin E and cyclin A gene expression. nIL-2 prevented cyclin A up-regulation but did not affect cyclin E, a clear indication that in activated human naïve CD4+ T cells only cyclin A expression is dependent on the IL-2/IL-2R signalling pathway, consistent with previous reports.3 Interestingly, BMS-345541 and PS-1145 prevented the expression not only of cyclin A, but also of cyclin E, providing compelling evidence for involvement of IKK in the regulation of cyclin E expression in human naïve CD4+ T cells. In light of the essential role played by the CDK2/cyclin E complex in initiating DNA replication,5 this finding underscores a critical function of IKK in the regulation of T-cell entry into S phase. Degradation of p27KIP1 by the ubiquitin–proteasome

pathway at the Palmatine G0/G1 transition results in activation of the cyclin E/CDK2 complex, and commitment of cells to S phase.41 In our results, stimulation of human naïve CD4+ T cells resulted in a considerable decrease in p27KIP1 that was prevented by nIL-2, or BMS-345541 or PS-1145. The degradation of p27KIP1 is a complex process that requires the formation of a ternary complex with cyclin D/CDK4, followed by p27KIP1 phosphorylation on Thr187 by cyclin E/CDK2.4 The RING finger-type ubiquitin ligase complex SCFSKP2-CKS1B recognizes phosphorylated p27KIP1 through the C-terminus of two of its subunits, SKP2 and CKS1B, resulting in targeting of p27KIP1 for ubiquitination and degradation.42 SKP2 and CKS1B levels periodically oscillate during the cell cycle: they are low or absent during G0 and early G1 phases, increase in late G1 phase, and peak in S phase, dropping as cells proceed through M and early G1 phases.

, 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 JNK inhibitor 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, Ibrutinib 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 Idelalisib chemical structure 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).