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Copyright © 2021 Gonçalves et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license., Activation of immune cells in response to fungal infection involves the reprogramming of their cellular metabolism to support antimicrobial effector functions. Although metabolic pathways such as glycolysis are known to represent critical regulatory nodes in antifungal immunity, it remains undetermined whether these are differentially regulated at the interindividual level. In this study, we identify a key role for 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) in the immunometabolic responses to Aspergillus fumigatus. A genetic association study performed in 439 recipients of allogeneic hematopoietic stem cell transplantation (HSCT) and corresponding donors revealed that the donor, but not recipient, rs646564 variant in the PFKFB3 gene increased the risk of invasive pulmonary aspergillosis (IPA) after transplantation. The risk genotype impaired the expression of PFKFB3 by human macrophages in response to fungal infection, which was correlated with a defective activation of glycolysis and the ensuing antifungal effector functions. In patients with IPA, the risk genotype was associated with lower concentrations of cytokines in the bronchoalveolar lavage fluid samples. Collectively, these findings demonstrate the important contribution of genetic variation in PFKFB3 to the risk of IPA in patients undergoing HSCT and support its inclusion in prognostic tools to predict the risk of fungal infection in this clinical setting. IMPORTANCE The fungal pathogen Aspergillus fumigatus can cause severe and life-threatening forms of infection in immunocompromised patients. Activation of glycolysis is essential for innate immune cells to mount effective antifungal responses. In this study, we report the contribution of genetic variation in the key glycolytic activator 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) to the risk of invasive pulmonary aspergillosis (IPA) after allogeneic hematopoietic stem cell transplantation. The PFKFB3 genotype associated with increased risk of infection was correlated with an impairment of the antifungal effector functions of macrophages in vitro and in patients with IPA. This work highlights the clinical relevance of genetic variation in PFKFB3 to the risk of IPA and supports its integration in risk stratification and preemptive measures for patients at high risk of IPA., This work was supported by the Fundação para a Ciência e Tecnologia (FCT) (PTDC/SAU-SER/29635/2017, PTDC/MED-GEN/28778/2017, UIDB/50026/2020, and UIDP/50026/2020), the Northern Portugal Regional Operational Program (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (ERDF) (NORTE-01-0145-FEDER-000039), the Institut Mérieux (Mérieux Research Grant 2017), the European Society of Clinical Microbiology and Infectious Diseases (ESCMID Research Grant 2017), the European Union’s Horizon 2020 research and innovation program under grant agreement no. 847507, and the “la Caixa” Foundation (ID 100010434) and FCT under the agreement LCF/PR/HR17/52190003. Individual support was provided by FCT (SFRH/BD/136814/2018 to S.M.G., PD/BD/137680/2018 to D.A., CEECIND/04058/2018 to C.C., and CEECIND/03628/2017 to A.C.). M.G.N. was supported by an ERC Advanced Grant and a Spinoza Grant of the Netherlands Organization for Scientific Research.

The application of innovative three-dimensional (3D) spheroids cell culture strategy to Parasitology offers the opportunity to closely explore host&ndash, parasite interactions. Here we present a first report on the application of 3D hepatic spheroids to unravel the immune response of canine hepatocytes exposed to Leishmania infantum. The liver, usually considered a major metabolic organ, also performs several important immunological functions and constitutes a target organ for L. infantum infection, the etiological agent of canine leishmaniasis (CanL), and a parasitic disease of major veterinary and public health concern. 3D hepatic spheroids were able to sense and immunologically react to L. infantum parasites, generating an innate immune response by increasing nitric oxide (NO) production and enhancing toll-like receptor (TLR) 2 and interleukin-10 gene expression. The immune response orchestrated by canine hepatocytes also lead to the impairment of several cytochrome P450 (CYP450) with possible implications for liver natural xenobiotic metabolization capacity. The application of meglumine antimoniate (MgA) increased the inflammatory response of 3D hepatic spheroids by inducing the expression of Nucleotide oligomerization domain (NOD) -like receptors 1 and NOD2 and TLR2, TLR4, and TLR9 and enhancing gene expression of tumour necrosis factor &alpha, It is therefore suggested that hepatocytes are key effector cells and can activate and orchestrate the immune response to L. infantum parasites.

We review state-of-the-art in translational and clinical studies focusing on the tumor microenvironment (TME) with a focus on tumor-infiltrating B cells (TIBs). The TME is a dynamic matrix of mutations, immune-regulatory networks, and distinct cell-to-cell interactions which collectively impact on disease progress. We discuss relevant findings concerning B cells in pancreatic cancer, the concepts of "bystander" B cells, the role of antigen-specific B cells contributing to augmenting anticancer-directed immune responses, the role of B cells as prognostic markers for response to checkpoint inhibitors (ICBs), and the potential use in adoptive cell tumor-infiltrating lymphocyte (TIL) products.

Highlights • In addition to ethnicity, socio-economic factors, prior vaccinations and exposure to other coronaviruses, other factors need to be considered to explain geographical and regional variations in susceptibility, severity of clinical expression of COVID-19 disease and outcomes. • Differences in peptide binding of SARS-CoV-2 variants to MHC class II, but not to MHC class I alleles frequent in individuals with African, Asian or Caucasian descent could be identified. • Single mutations in the wildtype of SARS-CoV-2, the so called B strain or L strain impact on MHC presentation • Most likely there is selective pressure from MHC class II alleles in regard to binding of the ORF8 (L84S) variants assuming that this mutation may be biologically relevant • MHC 4 – digit typing should be considered in studies examining clinical outcomes in patients with COVID-19., As the 2019 (COVID-19) pandemic caused by the novel coronavirus, SARS-CoV-2 spreads globally, differences in adverse clinical management outcomes have been associated with associated with age >65years, male gender, and co-morbidities such as smoking, diabetes, hypertension, cardiovascular comorbidity and immunosuppression. Ethnicity has been the focus of attention after data from the United Kingdom showed a disproportionate number of deaths among healthcare workers from black, Asian and other ethnic minority backgrounds (1). In addition to ethnicity, socio-economic factors, prior vaccinations and exposure to other coronaviruses, other factors need to be considered to explain geographical and regional variations in susceptibility, severity of clinical expression of COVID-19 disease and outcomes. In the United States there have been disproportionate COVID-19 death rates among African Americans at around 2.6 times higher than that of other groups. Although these data could be due to multiple cultural and socioeconomic factors an underlying genetic susceptibility to SARS-CoV-2 infection may be a factor.

We review state-of-the-art in translational and clinical studies focusing on the tumor microenvironment (TME) with a focus on tumor-infiltrating B cells (TIBs). The TME is a dynamic matrix of mutations, immune-regulatory networks, and distinct cell-to-cell interactions which collectively impact on disease progress. We discuss relevant findings concerning B cells in pancreatic cancer, the concepts of "bystander" B cells, the role of antigen-specific B cells contributing to augmenting anticancer-directed immune responses, the role of B cells as prognostic markers for response to checkpoint inhibitors (ICBs), and the potential use in adoptive cell tumor-infiltrating lymphocyte (TIL) products.

Hepatocytes constitute the majority of hepatic cells, and play a key role in controlling systemic innate immunity,viapattern-recognition receptors (PRRs) and by synthesizing complement and acute phase proteins.Leishmania infantum, a protozoan parasite that causes human and canine leishmaniasis, infects liver by establishing inside the Kupffer cells. The current study proposes the elucidation of the immune response generated by dog hepatocytes when exposed toL. infantum. Additionally, the impact of adding leishmanicidal compound, meglumine antimoniate (MgA), to parasite-exposed hepatocytes was also addressed.L. infantumpresents a high tropism to hepatocytes, establishing strong membrane interactions. The possibility ofL. infantuminternalization by hepatocytes was raised, but not confirmed. Hepatocytes were able to recognize parasite presence, inducing PRRs [nucleotide oligomerization domain (NOD)1, NOD2 and Toll-like receptor (TLR)2] gene expression and generating a mix pro- and anti-inflammatory cytokine response. Reduction of cytochrome P 450s enzyme activity was also observed concomitant with the inflammatory response. Addition of MgA increased NOD2, TLR4 and interleukin 10 gene expression, indicating an immunomodulatory role for MgA. Hepatocytes seem to have a major role in coordinating liver's innate immune response againstL. infantuminfection, activating inflammatory mechanisms, but always balancing the inflammatory response in order to avoid cell damage.

Kupffer cells (KC) are the liver macrophage population that resides in the hepatic sinusoids and efficiently phagocyte pathogens by establishing an intimate contact with circulating blood. KC constitute the liver host cells in Leishmania infection, nevertheless little is described about their role, apart from their notable contribution in granulomatous inflammation. The present study aims to investigate how canine KC sense and react to the presence of Leishmania infantum promastigotes and amastigotes by evaluating the gene expression of specific innate immune cell receptors and cytokines, as well as the induction of nitric oxide and urea production. Complementarily, the impact of a leishmanicidal drug - meglumine antimoniate (MgA) - in infected KC was also explored. KC revealed to be susceptible to both parasite forms and no major differences were found in the immune response generated. L. infantum parasites seem to interact with KC innate immune receptors and induce an anergic state, promoting immune tolerance and parasite survival. The addition of MgA to infected KC breaks the parasite imposed silence and increased gene expression of Toll-like receptors (TLR) 2 and TLR4, possibly activating downstream pathways. Understanding how KC sense and react to parasite presence could bring new insights into the control or even elimination of canine leishmaniasis.

Programmed cell death-1 protein (PD-1) is an immune checkpoint that has gained popularity in the treatment of several advanced cancers. Inhibiting this checkpoint is known to enhance immune response, but is also known to diminish immune tolerance and to increase autoimmune toxicity. We discuss a case of rapid onset fulminant Type 1 diabetes induced by treatment with anti-programmed cell death-1 monoclonal antibody, nivolumab, in a patient with late-stage non-small-cell lung adenocarcinoma. The patient had no history of previous diabetes but did reveal a high-risk genotype for Type 1 diabetes development (DR3-DQ2; DR4-DQ8). This finding supports that acute Type 1 diabetes can be an important adverse effect of immunotherapies targeting T-cell activation regulation. Because of the severity of this adverse effect, physicians should be aware of it, and studies directed to the detection of new biomarkers for early risk stratification (e.g., HLA) should be sought.

In the light of poor management outcomes of antibiotic-resistant respiratory tract infection (RTI)-associated sepsis syndrome and multidrug-resistant tuberculosis (MDR-TB), new management interventions based on host-directed therapies (HDTs) are warranted to improve morbidity, mortality and long-term functional outcomes. We review developments in potential HDTs based on precision cancer therapy concepts applicable to RTIs including MDR-TB.Immune reactivity, tissue destruction and repair processes identified during studies of cancer immunotherapy share common pathogenetic mechanisms with RTI-associated sepsis syndrome and MDR-TB. T-cell receptors (TCRs) and chimeric antigen receptors targeting pathogen-specific or host-derived mutated molecules (major histocompatibility class-dependent/ major histocompatibility class-independent) can be engineered for recognition by TCR γδ and natural killer (NK) cells. T-cell subsets and, more recently, NK cells are shown to be host-protective. These cells can also be activated by immune checkpoint inhibitor (ICI) or derived from allogeneic sources and serve as potential for improving clinical outcomes in RTIs and MDR-TB.Recent developments of immunotherapy in cancer reveal common pathways in immune reactivity, tissue destruction and repair. RTIs-related sepsis syndrome exhibits mixed immune reactions, making cytokine or ICI therapy guided by robust biomarker analyses, viable treatment options.

Systemic inflammation has been implicated in the pathogenesis of chronic obstructive pulmonary disease (COPD) systemic effects. However, most COPD patients do not suffer from persistent systemic inflammation even after exacerbations and exercise and scientific evidence has provided conflicting results. Our aim is to evaluate inflammatory gene expression at rest and at 1 and 24 h after strenuous exercise in COPD patients and study the patient variables associated with inflammatory expression.A cross-sectional study was conducted in COPD patients who were recruited on entry to a pulmonary rehabilitation (PR) program. Demographic, clinical and functional data were collected. Blood samples were collected and gene expression was analyzed by reverse transcriptase polymerase chain reaction for IFNg, IL1b, IL6, IL8, TNFa, TGFb1 and iNOS.The study included 21 patients (15 men, 71.4%), mean age 66.1 years old (SD = 8.27), mean FEV1 46.76% (SD 20.90%), 67% belonging to GOLD grade D, mean BODE index of 3.9, 90.5% with smoking history, mean BMI 25.81 (SD = 4.87), median of 1.29 exacerbations in the previous year.There was no statistical significant difference between inflammatory expression at rest and at 1 h and 24 h after the maximal exercise test for all tested genes.We found an association between BMI and inflammatory expression at all the points of time checked, a slight inverse association occurs with low BMI for mRNA IL1b, IL6, TNFa, TGFb1 and iNOS, and there was a more pronounced positive association for obese patients for all tested genes.This preliminary study did not show an enhanced inflammatory gene expression from rest to 1 h and 24 h after short-term exercise, but did show an increased inflammatory gene expression in both BMI extremes, both at rest and after exercise, suggesting not only malnourishment, but also obesity as potential links between COPD and systemic inflammation. Studies with larger samples and designed to definitely exclude OSA or OHS as confounding factors in obese patients are required. Keywords: COPD, Inflammation, Exercise, Obesity, Reverse transcriptase polymerase chain reaction

The diversity of killer-cell immunoglobulin-like receptors (KIR) genes was evaluated in Portuguese and the observed genotypic profiles were found related to the ones reported in European populations. The KIR repertoire after hematopoietic stem cell transplantation is determined by these gene frequencies and the KIR group B motifs are the less common. We estimated donor–KIR/recipient–ligand interactions in transplants with related donors and unrelated donors found in a local registry or from abroad. A large fraction of transplants had all three ligands of inhibitory receptors, and therefore, in theory were not prone to natural killer cell (NK) mediated alloreactivity. Furthermore, the distribution of KIR alloreactive interactions was found independent of the donor–recipient genetic proximity, probably because of different gene segregation and comparable KIR frequencies in the donor pools.

Copyright © 2019 Campos, Leite, Pereira, Vaz, Branca, Campilho, Freitas, Ligeiro, Marques, Torrado, Silvestre, Lacerda, Campos, Cunha and Carvalho. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms., Background: Reactivation of latent human cytomegalovirus (CMV) in patients undergoing allogeneic stem-cell transplantation (HSCT) predisposes to several clinical complications and is therefore a major cause of morbidity and mortality. Although pentraxin-3 (PTX3) has been previously described to bind both human and murine CMV and mediate several host antiviral mechanisms, whether genetic variation in the PTX3 locus influences the risk of CMV infection is currently unknown. Methods: To dissect the contribution of genetic variation within PTX3 to the development of CMV infection, we analyzed described loss-of-function variants at the PTX3 locus in 394 recipients of HSCT and their corresponding donors and assessed the associated risk of CMV reactivation. Results: We report that the donor, but not recipient, h2/h2 haplotype in PTX3 increased the risk of CMV reactivation after 24 months following transplantation, with a significant effect on survival. Among recipients with h2/h2 donors, CMV seropositive patients as well as those receiving grafts from unrelated donors, regardless of the CMV serostatus, were more prone to develop viral reactivation after transplantation. Most importantly, the h2/h2 haplotype was demonstrated to display an influence toward risk of CMV reactivation comparable to that conferred by the unrelated status of the donor alone. Conclusions: Our findings demonstrate the important contribution of genetic variation in donor PTX3 to the risk of CMV reactivation in patients undergoing HSCT, highlighting a promising prognostic value of donor PTX3 to predict risk of CMV reactivation in this clinical setting., This work was supported by the Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER) (NORTE-01-0145-FEDER-000013), and the Fundação para a Ciência e Tecnologia (FCT) (SFRH/BPD/96176/2013 to CC, IF/01390/2014 to ET, IF/00021/2014 to RS, and IF/00735/2014 to AgC).

Background The glycan moieties sialyl-Lewis-X and/or -A (sLeX/A) are the primary ligands for E-selectin, regulating subsequent tumor cell extravasation into distant organs. However, the nature of the glycoprotein scaffolds displaying these glycans in breast cancer remains unclear and constitutes the focus of the present investigation. Methods We isolated glycoproteins that bind E-selectin from the CF1_T breast cancer cell line, derived from a patient with ductal carcinoma. Proteins were identified using bottom-up proteomics approach by nanoLC-orbitrap LTQ-MS/MS. Data were curated using bioinformatics tools to highlight clinically relevant glycoproteins, which were validated by flow cytometry, Western blot, immunohistochemistry and in-situ proximity ligation assays in clinical samples. Results We observed that the CF1_T cell line expressed sLeX, but not sLeA and the E-selectin reactivity was mainly on N-glycans. MS and bioinformatics analysis of the targeted glycoproteins, when narrowed down to the most clinically relevant species in breast cancer, identified CD44 glycoprotein (HCELL) and CD13 as key E-selectin ligands. Additionally, the co-expression of sLeX-CD44 and sLeX-CD13 was confirmed in clinical breast cancer tissue samples. Conclusions Both CD44 and CD13 glycoforms display sLeX in breast cancer and bind E-selectin, suggesting a key role in metastasis development. Such observations provide a novel molecular rationale for developing targeted therapeutics. General significance While HCELL expression in breast cancer has been previously reported, this is the first study indicating that CD13 functions as an E-selectin ligand in breast cancer. This observation supports previous associations of CD13 with metastasis and draws attention to this glycoprotein as an anti-cancer target.

info:eu-repo/grantAgreement/FCT/5876/147283/PT Funding for this work was provided by the Portuguese Foundation for Science and Technology (FCT) with co-participation of European Union Fund (FEDER) through the research project UID/CVT/00276/2013. The authors present special thanks to Sandra Carvalho and Rosario Luis (Histology Dept., FMV) and Gloria Nunes (Immunology Dept., FCM) for technical assistance. Post-parturient behavior of mammalian females is essential for early parent–offspring contact. After delivery, lambs need to ingest colostrum for obtaining the related immunological protection, and early interactions between the mother and the lamb are crucial. Despite visual and auditory cues, olfactory cues are decisive in lamb orientation to the mammary gland. In sheep, the inguinal sinus is located bilaterally near the mammary gland as a skin pouch (IGS) that presents a gland that secretes a strong-smelling wax. Sheep IGS gland functions have many aspects under evaluation. The objective of the present study was to evaluate sheep IGS gland functional aspects and mRNA transcription and the protein expression of several hormone receptors, such as progesterone receptor (PGR), estrogen receptor 1 (ESR1), and 2 (ESR2) and prolactin receptor (PRLR) present. In addition, another aim was to achieve information about IGS ultrastructure and chemical compounds produced in this gland. All hormone receptors evaluated show expression in IGS during the estrous cycle (follicular/luteal phases), pregnancy, and the post-partum period. IGS secretion is rich in triterpenoids that totally differ from the surrounding skin. They might be essential substances for the development of an olfactory preference of newborns to their mothers. © 2017 by the authors. Licensee MDPI, Basel, Switzerland. publishersversion published

Background Targeting humoral immunity has been proved effective in several inflammatory rheumatic diseases (IRD). Though clinical trials have shown some efficacy of B-cell depletion in ankylosing spondylitis (AS), results are less convincing. Other studies have revealed an association between mutations and expression of immune regulatory genes suggesting B-cell dysfunction in the development and progression of AS. Yet, there is still lack of data describing B-cell subsets in AS, how these compare to other IRD and an evaluation of B cell compartment homeostasis in the pathophysiology of this disease. Objectives To assess and compare the immature, naive and antigen differentiated subsets of peripheral B-cell compartment in AS with those in healthy controls (HC) and other IRD Methods Patients (pts) with AS, RA and SLE according to respective classification criteria were included in this study. Pts under biologic DMARDS were not included. Sociodemographic and clinical variables were recorded. Blood samples were collected for quantification of inflammatory markers (ESR and CRP), immunoglobulin serum levels and assessment of B-cell immature transitional stages and mature subsets by flow cytometry (figure). Mann-Whitney and Fisher9s exact test were used for comparison of AS with other groups Results Overall, 60 pts and 12 HC were included (Table). All patient groups presented similar and rather low levels of inflammation, as measured by CRP, ESR and immunoglobulins, in addition to a decreased lymphocyte count by comparison with HC. There were no differences in the B-cell counts between AS pts and HC, with both groups having higher B-cell counts than RA and SLE pts. Regarding B-cell subsets, the immature transitional compartment of AS pts was found in normal range, but not in the RA and SLE groups. In fact, the latter presented a significant decrease in all transitional cell maturity stages (T1-T3). The next step in B-cell differentiation is mature naive cells, also found in normal levels in AS and decreased in RA and in particular in SLE. AS pts presented slightly higher counts of CD27+IgD+ MZ-like and class able to switch memory cells with reference to HC and these cell numbers were found to be low in RA and even lower in SLE pts. Switched memory CD27+IgD- B-cells were reduced in all patient groups, however, only SLE pts presented highly decreased cell levels. Conclusions We found that while a severe dysfunction is present in the homeostasis of the B-cell compartment in RA and in particular SLE pts, which are lymphopenic in both immature and mature B-cell compartments, it appears that AS pts are not affected in the same way. At this stage, functional studies appear to be necessary in order to identify differences in key mechanisms of B cell development and differentiation that may play a role in the aetiology and progression of these inflammatory rheumatic diseases. Our first results, however, establish that pathophysiological mechanisms involving B-cells clearly differentiate AS from RA and SLE Disclosure of Interest None declared

Within the framework of the EU-funded HLA-NET action, an analysis of three G-group alleles, HLA-B*44:02:01G, DRB1*14:01:01G and DQB1*03:01:01G, was undertaken in 12 European populations. Ambiguities were resolved by polymerase chain reaction-sequence-specific amplification (PCR-SSP) or PCR-sequence-based typing (PCR-SBT) in a total of 5095 individuals. The results of the DRB1*14:01/14:54 ambiguity showed high relative ratios (24–53%) of DRB1*14:01 in Bulgarians, Croatians, Greeks, Italians and Slovenians, contrasting with low ratios (6–13%) in Austrians, Finnish, French, Hungarians, Norwegians and Swiss. Resolution of the B*44:02/44:27 ambiguity showed that B*44:27 had a high relative ratio in Slovenians (25.5%) and Bulgarians (37%) and low in French and Swiss (0.02–1%), and was not observed in Greeks and Italians. The highest relative ratio of DQB1*03:19 was found in Portuguese (11%), by contrast with low ratios (0–3%) in the other five populations. Analysis of the A, B, DRB1 phenotypes and family-derived haplotypes in 1719 and 403 individuals positive for either HLA-B*44:02G or DRB1*14:01G ambiguities, respectively, showed some preferential associations, such as A*26∼DRB1*14:01, B*35∼DRB1*14:01, B*38∼DRB1*14:01 and B*44:27∼DRB1*16. Because these ambiguities are located outside the peptide-binding site, they may not be recognized by alloreactive T-cells. However, because of strong linkage disequilibrium (LD), the DRB1*14:01 vs DRB1*14:54 and the B*44:02 vs B*44:27 mismatches are associated to DRB3-, and C-mismatches, respectively. These results are informative for algorithms searching unrelated hematopoietic stem cell donors. For B*44:27-positive patients, searches are expected to be more successful when requesting donors from Southeastern-European ancestry. Furthermore, the introduction of human leukocyte antigen (HLA)-typing strategies that allow resolving exon 4 (for class I) and exon 3 (for class II) polymorphisms can be expected to contribute significantly to population genetics studies.

Treg infusion for graft-versus-host-disease treatment has been increasingly investigated. Yet, Treg of unknown specificity may suppress graft-versus-leukemia. To ascertain the factors affecting Ag-sp Treg potency and specificity of suppression, we assessed Ag-sp Treg phenotype and function within different expansion conditions and suppression assay (SA) milieus. Ex vivo Treg were co-cultured with monocyte-derived dendritic cells (DC) presenting allogeneic Ag directly (allo-DC) or indirectly (self-DC loaded with allo-lysate). Suppression of effector proliferation and cytokine secretion was assessed in SA with the same DC as in expansion (original) or 3rd party DC (3P) from various donors. Ag-sp Treg significantly suppressed Tcon and CD8+ T cell responses to original DC. Directly expanded cells suppressed responses to Ag presented direct and indirectly; however, indirectly expanded cells were less suppressive of responses to allo-DC. More potent Treg were less specific in their suppression of Tcon. Still, responses to 3P donors with MHC matches to original DC donor were significantly more suppressed than responses to unmatched 3P donors. Potency of suppression was also negatively correlated to expression of CD86 and CD83 in DC. Finally, FlowSOM analysis showed minimal overlap between Ag-sp Treg and activated Tcon phenotypes, suggesting Ag-sp Treg were pure. K-means clustering of Ag-sp Treg expression of CD137, CD154, CTLA-4, PD-1, CTLA-4 and HLA-DR identified clusters that may be correlated to Treg function. We propose the specificity of Ag-sp Treg function in the inflammatory milieu is a result of Treg phenotype, type of responders, DC co-stimulation and degree of MHC match between expansion DC and DC present in the milieu.

Human leukocyte antigen (HLA)-B27 is the mostly known major histocompatibility complex (MHC) gene associated with ankylosing spondylitis (AS). Nonetheless, there is substantial evidence that other MHC genes appear to be associated with the disease, although it has not yet been established whether these associations are driven by direct associations or by linkage disequilibrium (LD) mechanisms. We aimed to investigate the contributions of HLA class I and II alleles and B27-haplotypes for AS in a case-control study. A total of 188 HLA-B27 AS cases and 189 HLA-B27 healthy controls were selected and typed for HLA class I and II by the Luminex polymerase chain reaction-sequence specific oligonucleotide probe (PCR-SSOP) method. Allelic and haplotypic distributions were estimated by maximum likelihood method using Arlequin v3.11 and statistical analysis were performed by Stata10.1. No associations were found between non-HLA-B27 loci and AS susceptibility, but several associations were observed for phenotypic features of the disease. DRB1*08 was identified as a risk factor for uveitis and DQB1*04 seems to provide protection for AS severity (functional, metrological and radiological indexes). A*02/B27/C*02/DRB1*01/DQB1*05 [P

The Zucker diabetic fatty (ZDF) rat is an obesity and type 2 diabetes model. Progression to diabetes is well characterised in ZDF rats, but only in the fasted state. We evaluated the mechanisms underlying postprandial insulin resistance in young ZDF rats. We tested the hypothesis that the overall postprandial action of insulin is affected in ZDF rats as a result of impairment of the hepatic parasympathetic-nitric oxide (PSN-NO) axis and ‡ or glutathione (GSH), resulting in decreased indirect (PSN-NO axis) and direct the action of insulin. Nine-week-old male ZDF rats and lean Zucker rats (LZR, controls) were used. The action of insulin was assessed in the fed state and after parasympathetic antagonism (atrophine). Basal hepatic NO and GSH were measured, as well as NO synthase (NOS) and c-glutamyl-cysteine synthethase (GCS) activity and expression. ZDF rats presented postprandial hyperglycaemia (ZDF, 201.4 � 12.9 mg ‡ dl; LZR, 107.7 � 4.3 mg ‡ dl), but not insulinopaenia (ZDF, 5.9 � 0.8 ng ‡ ml; LZR, 1.5 � 0.3 ng ‡ ml). Total postprandial insulin resistance was observed (ZDF, 78.6 � 7.5 mg glucose ‡ kg; LZR, 289.2 � 24.7 mg glucose ‡ kg), with a decrease in both the direct action of insulin (ZDF, 54.8 � 7.0 mg glucose ‡ kg; LZR, 173.3 � 20.5 mg glucose ‡ kg) and the PSN-NO axis (ZDF, 24.5 � 3.9 mg glucose ‡ kg; LZR, 115.9 � 19.4 mg glucose ‡ kg). Hepatic NO (ZDF, 117.2 � 11.4 lmol ‡ g tissue; LZR, 164.6 � 4.9 lmol ‡ g tissue) and GSH (ZDF, 4.9 � 0.3 lmol ‡ g; LZR, 5.9 � 0.2 lmol ‡ g) were also compromised as a result of decreased NOS and GCS activity, respectively. These results suggest a compromise of the mechanism responsible for potentiating the action of insulin after a meal in ZDF rats. We show that defective PSN-NO axis and GSH synthesis, together with an impaired direct action of insulin, appears to contribute to postprandial insulin resistance in this obesity model.

Objective.Unconfirmed reports describe association of ankylosing spondylitis (AS) with several candidate genes includingANKH. Cellular export of inorganic pyrophosphate is regulated by the ANK protein, and mutant mice (ank/ank), which have a premature stop codon in the 3’ end of theankgene, develop severe ankylosis. We tested the association between single-nucleotide polymorphisms (SNP) in these genes and susceptibility to AS in a population of patients with AS. We investigated the role of these genes in terms of functional (BASFI) and metrological (BASMI) measures, and the association with radiological severity (mSASSS).Methods.Our study was conducted on 355 patients with AS and 95 ethnically matched healthy controls. AS was defined according to the modified New York criteria. Four SNP inANKH(rs27356, rs26307, rs25957, and rs28006) were genotyped. Association analysis was performed using Cochrane-Armitage and linear regression tests for dichotomous and quantitative variables. Analyses of Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), BASFI, and mSASSS were controlled for sex and disease duration.Results.None of the 4 markers showed significant single-locus disease associations (p > 0.05), suggesting thatANKHwas not a major determinant of AS susceptibility in our population. No association was observed between these SNP and age at symptom onset, BASDAI, BASFI, BASMI, or mSASSS.Conclusion.These results confirm data in white Europeans thatANKHis probably not a major determinant of susceptibility to AS.ANKHpolymorphisms do not markedly influence AS disease severity, as measured by BASMI and mSASSS.

Human cytomegalovirus (HCMV) is the most frequent cause of congenital infection. Despite the fact that 5-17% of asymptomatic infected babies will develop late sequelae and, therefore, should be closely followed, most of these children will remain undetected, as screening of all newborns by viral culture is too expensive and no valid alternative has been widely accepted. The aim of this work was to demonstrate that pool testing can be used to screen HCMV congenital infection in newborns. For this purpose, a real-time PCR technique was tested in urine pools. This pool method was applied to all urine specimens from children received in the virology laboratory of the Centro Hospitalar Cova da Beira for diagnosis of HCMV infection for a period of 14 months. Ten out of the 160 urine samples were tested positive by shell-vial culture and were also detected by this pool method. Additionally, 100 urine specimens, collected in 2004 and culture negative for HCMV were included to test the specificity of this methodology, all of which were negative. In conclusion, these results suggest that urine pools can be used to detect HCMV-positive urines in children, with similar sensitivity and specificity when compared with the standard method. Because of the very significant reduction both in terms of labour and cost of testing materials, this methodology may represent a valid option for screening the HCMV congenital infection in newborns.

Gene expression studies demand that data be normalized to an endogenous control gene. The selection of this control is not straightforward, in particular for tumor tissue samples. We have measured the

Introduction. This study aims to compare the molecular gene expression during ischemia reperfusion injury. Several surgical times were considered: in the beginning of the harvesting (T0), at the end of the cold ischemia period (T1), and after reperfusion (T2) and compared with graft dysfunction after liver transplant (OLT). Methods. We studied 54 patients undergoing OLT. Clinical, laboratory data, and histologic data (Suzuki classification) as well as the Survival Outcomes Following Liver Transplantation (SOFT) score were used and compared with the molecular gene expression of the following genes: Interleukin (IL)-1b, IL-6, tumor necrosis factor-a, perforin, E-selectin (SELE), Fas-ligand, granzyme B, heme oxygenase-1, and nitric oxide synthetase. Results. Fifteen patients presented with graft dysfunction according to SOFT criteria. No relevant data were obtained by comparing the variables graft dysfunction and histologic variables. We observed a statistically significant relation between SELE at T0 (P ¼ .013) and IL-1b at T0 (P ¼ .028) and early graft dysfunction. Conclusions. We conclude that several genetically determined proinflammatory expressions may play a critical role in the development of graft dysfunction after OLT.

© 2015 Xavier et al.; licensee BioMed Central. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated., Introduction: The aim of this study was to characterize the association of human leukocyte antigen (HLA) B alleles and major histocompatibility complex (MHC) single nucleotide polymorphisms (SNPs) with Behçet's disease (BD) in an Iranian dataset. Methods: The association of three SNPs in the MHC region previously identified as the most associated in high-density genotyping studies was tested in a case-control study on 973 BD patients and 825 controls from Iran, and the association of HLA-B alleles was tested in a subset of 681 patients and 414 controls. Results: We found that HLA-B*51 (P = 4.11 × 10(-41), OR [95% CI] = 4.63[3.66-5.85]) and B*15 confer risk for BD (P = 2.83 × 10(-2), OR [95% CI] = 1.75[1.08-2.84]) in Iranian, and in B*51 negative individuals, only the B*15 allele is significantly associated with BD (P = 2.51 × 10(-3), OR [95% CI] = 2.40[1.37-4.20]). rs76546355, formerly known as rs116799036, located between HLA-B and MICA (MHC class I polypeptide-related sequence A), demonstrated the same level of association with BD as HLA-B*51 (P adj = 1.78 × 10(-46), OR [95% CI] = 5.46[4.21-7.09], and P adj = 8.34 × 10(-48), OR [95% CI] = 5.44[4.20-7.05], respectively) in the HLA-B allelotyped subset, while rs2848713 was less associated (P adj = 7.14 × 10(-35), OR [95% CI] = 3.73[2.97-4.69]) and rs9260997 was not associated (P adj = 1.00 × 10(-1)). Additionally, we found that B*51 genotype-phenotype correlations do not survive Bonferroni correction, while carriers of the rs76546355 risk allele predominate in BD cases with genital ulcers, positive pathergy test and positive BD family history (2.31 × 10(-4) ≤ P ≤ 1.59 × 10(-3)). Conclusions: We found that the HLA-B*51 allele and the rs76546355/rs116799036 MHC SNP are independent genetic risk factors for BD in Iranian, and that positivity for the rs76546355/rs116799036 risk allele, but not for B*51, does correlate with specific demographic characteristics or clinical manifestations in BD patients., This work was supported by the Portuguese Fundação para a Ciência e a Tecnologia (grants PTDC/SAU-GMG/098937/2008, PTDC/IIM-GES/5015/2012 and CMUP-ERI/TPE/0028/2013, fellowships SFRH/BD/43895/2008 to JMX, SFRH/BPD/35737/2007 to PA, SFRH/BPD/70008/2010 to IS, a Ciência and an Investigator-FCT contract to SAO), and the Research Committee of the Tehran University of Medical Sciences (grant 132/714).

© 2017 American Academy of Allergy, Asthma & Immunology, Proinflammatory immune responses are critically required for antimicrobial host defenses; however, excessive inflammation has the potential to damage host tissues thereby paradoxically contributing to the progression of infection. A central negative regulator of inflammatory responses is IL-10, an immunosuppressive cytokine with a wide variety of functions across multiple cell types. Although the role of IL-10 during infection appears to vary for different microorganisms, a largely detrimental role has been attributed to this cytokine during fungal disease. Given the variable risk of infection and its outcome among patients with comparable predisposing factors, susceptibility to invasive aspergillosis (IA) is thought to rely largely on genetic predisposition. The initial investigation of genetic variability at the IL10 locus led to the identification of single nucleotide polymorphisms (SNPs) influencing its transcriptional activity; thus, IL-10 may be a reasonable candidate for the genetic regulation of susceptibility to IA in high-risk patients., Supported by the Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER) (NORTE-01-0145-FEDER-000013), the Fundação para a Ciência e Tecnologia (FCT) (contracts IF/00735/2014 to A.C., IF/01390/2014 to E.T., IF/00021/2014 to R.S., and SFRH/BPD/96176/2013 to C.C.), the Conselho de Reitores das Universidades Portuguesas (CRUP), Portugal (Ações Integradas Luso-Alemãs A-43/16), the Deutscher Akademischer Austauschdienst (DAAD) (project-ID 57212690), the Fondo de Investigaciones Sanitarias (Madrid, Spain) (grant #PI12/02688) and the ERA-NET PathoGenoMics (grant #0315900A).

This study aimed to compare the histologic and molecular gene expression at several surgical times (beginning of harvesting, T0; end of cold ischemia period, T1; and after reperfusion, T2) to characterize the ischemia-reperfusion injury (IRI) in deceased-donor liver grafts harvested from patients with familial amyloidotic polyneuropathy (FAP). For this purpose, 54 patients undergoing liver transplantation were studied and divided into 3 groups: deceased donor to cirrhotic recipient (group 1; n = 27), deceased donor to FAP recipient (group 2; n = 15), and FAP donor to cirrhotic recipient (group 3; n = 12). The main comparison was performed between a histologic score (Suzuki score, adding steatosis and neutrophil infiltration), and molecular gene expression of the following genes: interleukin (IL) 1β, IL-6, E-selectin, Fas-ligand, granzyme B, heme oxygenase 1 (HO1), and nitric oxide synthetase (iNOS2A). We observed less neutrophil infiltration levels in group 3 in sample T0 (P = .0082), which was associated with gene expression of HO1 in the biopsies at T2 (P = .022). In group 3, the molecular expression of genes related to attenuated proinflammatory reaction during IRI, iNOS2A at T0 and HO1 at T2, was detected. We conclude that FAP liver grafts express differently the genes associated with an attenuated proinflammatory reaction, presenting less neutrophil infiltration at harvesting. These findings add more knowledge about the better short-term outcomes in patients receiving this type of liver graft.

This work was supported by the Portuguese Foundation for Science and Technology (FCT) - PTDC/SAU-MII/67561/2006 and Premio Santander Totta - UNL (Paula A. Videira), LPCC/Pfizer2011 (Mylene A. Carrascal), SFRH/BPD/21619/2005 (M. Guadalupe Cabral), SFRH/BD/81860/2011 (Mariana Silva), SFRH/BD/45120/2008 (Paulo F. Severino) and SFRH/BPD/ 66288/2009 (Jose Alexandre Ferreira). FCT is co-financed by European Social Fund (ESF) under Human Potential Operation Programme (POPH) from National Strategic Reference Framework (NSRF).) and European Union, QREN, FEDER, COMPETE, for funding the Organic Chemistry Research Unit (QOPNA) (project PEst-C/QUI/UI0062/2013; FCOMP-01-0124-FEDER-037296). Despite the wide acceptance that glycans are centrally implicated in immunity, exactly how they contribute to the tilt immune response remains poorly defined. In this study, we sought to evaluate the impact of the malignant phenotype-associated glycan, sialyl-Tn (STn) in the function of the key orchestrators of the immune response, the dendritic cells (DCs). In high grade bladder cancer tissue, the STn antigen is significantly overexpressed and correlated with the increased expression of ST6GALNAC1 sialyltransferase. Bladder cancer tissue presenting elevated expression of ST6GALNAC1 showed a correlation with increased expression of CD1a, a marker for bladder immature DCs and showed concomitant low levels of Th1-inducing cytokines IL-12 and TNF-α. Invitro, human DCs co-incubated with STn+ bladder cancer cells, had an immature phenotype (MHC-IIlow, CD80low and CD86low) and were unresponsive to further maturation stimuli. When contacting with STn+ cancer cells, DCs expressed significantly less IL-12 and TNF-α. Consistent with a tolerogenic DC profile, T cells that were primed by DCs pulsed with antigens derived from STn+ cancer cells were not activated and showed a FoxP3high IFN-γlow phenotype. Blockade of STn antigens and of STn+ glycoprotein, CD44 and MUC1, in STn+ cancer cells was able to lower the induction of tolerance and DCs become more mature.Overall, our data suggest that STn-expressing cancer cells impair DC maturation and endow DCs with a tolerogenic function, limiting their capacity to trigger protective anti-tumour T cell responses. STn antigens and, in particular, STn+ glycoproteins are potential targets for circumventing tumour-induced tolerogenic mechanisms. publishersversion published

Many species that belong to Artiodactyls order show an interdigital sinus (IS), as it occurs in sheep, in all four extremities. These are considered to be scent glands responsible for sexual communication having strong attractiveness to mature males at the peak of the breed- ing season. The aim of this study was to evaluate, in IS in cyclic ewes, the microscopic and ultra- structure (scanning and transmission electron microscopy) anatomy, secretion composition, and mRNA and protein expression of estrogen receptors a and b and progesterone receptors. Glandu- lar sebaceous structures occupy a superficial area of the pouch. The other glands present in the IS show a coiled tubular structure and tall and polyhedral secretory cells with irregular luminal surface resulting from the secretory process. Protein and mRNA gene transcription studies were performed to determine the presence of ER (a and b) and P4r in IS. At the follicular phase, IS cell populations analyzed using flow cytometry expressed higher levels of ERb compared with ERa (P < 0.05), whereas no difference was observed between them in the luteal phase. The IS amount of secretion was the highest in the follicular phase compared with luteal phase (P < 0.05) or pregnancy (P < 0.001).To the best of our knowledge, for the first time, the presence of ER (a and b) within the IS was demonstrated. As estrogen action is mediated by specific receptors in target cells, the presence of these receptors in IS might be needed to trigger signaling pathways involved in conspecific chemical (sexual) communication attributed to this area. Microsc. Res.

Genes in human leucocyte antigens (HLA) System are important in the study of autoimmune diseases and responsible for the rejection of transplants of organs and tissues. HLA genes are part of the human major histocompatibility complex (MHC) which is characterized by the presence of several multigene families, extensive polymorphism at many loci and significant linkage disequilibrium between alleles at particular loci. We analysed HLA-A,-B,-DRB1 locus phenotypes through a sample of 1,021 subjects that were randomly selected among the volunteers recruited by the Portuguese Bone Marrow Donors Registry (Cedace) in order to evaluate allele, gene, haplotype and phenotype frequencies. Allelic frequencies in each of the studied locus were obtained by direct counting. Maximum-likelihood haplotype frequencies were estimated using an expectation-maximization (EM) algorithm [2]. Locus phenotype and gene relative frequencies were estimated according to Baur and Danilov [1]. Hardy–Weinberg equilibrium were tested. The data presented is a definition of HLA genetic repertoire of Cedace with relevance on the strategic management for the increase of a more diverse register with clinical utility. info:eu-repo/semantics/publishedVersion

© 2013 American Society for Blood and Marrow Transplantation., Haploidentical hematopoietic stem cell transplantation (HSCT) constitutes an important alternative for patients lacking a human leukocyte antigen (HLA)-matched donor. Although the use of haploidentical donors is increasingly common, the long-term impact of generating a donor-derived immune system in the context of an HLA-mismatched thymic environment remains poorly characterized. We performed an in-depth assessment of immune reconstitution in a group of haploidentical HSCT recipients 4 to 6 years posttransplantation, in parallel with the respective parental donors and age-matched healthy control subjects. Our data show that the proportion of naive and memory subsets in the recipients, both within CD8(+) and CD4(+) T cells, more closely resembled that observed in age-matched control subjects than in the donors. HSCT recipients displayed relatively high signal-joint T cell-receptor excision circle levels and a high frequency of the recent thymic emigrant-enriched CD31(+) subset within naive CD4(+) and naive regulatory T cells. Moreover, CD8(+), CD4(+), and regulatory T cells from HSCT recipients displayed a diverse T cell repertoire. These results support a key role for thymic output in T cell reconstitution. Nevertheless, HSCT recipients had significantly shorter telomeres within a naive-enriched CD4(+) T cell population than age-matched control subjects, despite the similar telomere length observed within the most differentiated CD8(+) and CD4(+) T cell subsets. Overall, our data suggest that long-term immune reconstitution was successfully achieved after haploidentical HSCT, a process that appears to have largely relied on de novo T cell production., Supported by grant PTDC/SAU-MII/67662/2006 from Fundação para a Ciência e Tecnologia (FCT) and by Programa Operacional Ciência e Inovação 2010 (POCI2010) to M.V.D.S. In addition, R.I.A., A.S.A., R.T., and R.S.S. received scholarships from FCT cofinanced by POCI 2010 and FSE.

Dendritic cells (DCs) play an essential role in immunity against bacteria by phagocytosis and by eliciting adaptive immune responses. Previously, we demonstrated that human monocyte-derived DCs (MDDCs) express a high content of cell surface α2,6-sialylated glycans. However, the relative role of these sialylated structures in phagocytosis of bacteria has not been reported. Here, we show that treatment with a sialidase significantly improved the capacity of both immature and mature MDDCs to phagocytose Escherichia coli. Desialylated MDDCs had a significantly more mature phenotype, with higher expression of MHC molecules and interleukin (IL)-12, tumour necrosis factor-α, IL-6 and IL-10 cytokines, and nuclear factor-κB activation. T lymphocytes primed by desialylated MDDCs expressed more interferon-γ when compared with priming by sialylated MDDCs. Improved phagocytosis required E. coli sialic acids, indicating a mechanism of host-pathogen interaction dependent on sialic acid moieties. The DCs harvested from mice deficient in the ST6Gal.1 sialyltransferase showed improved phagocytosis capacity, demonstrating that the observed sialidase effect was a result of the removal of α2,6-sialic acid. The phagocytosis of different pathogenic E. coli isolates was also enhanced by sialidase, which suggests that modifications on MDDC sialic acids may be considered in the development of MDDC-based antibacterial therapies. Physiologically, our findings shed new light on mechanisms that modulate the function of both immature and mature MDDCs, in the context of host-bacteria interaction. Hence, with particular relevance to DC-based therapies, the engineering of α2,6-sialic acid cell surface is a novel possibility to fine tune DC phagocytosis and immunological potency.

Forkhead box N1 (FOXN1) is a transcription factor crucial for thymic epithelium development and prevention of its involution. Investigation of a patient with a rare homozygous FOXN1 mutation (R255X), leading to alopecia universalis and thymus aplasia, unexpectedly revealed non-maternal circulating T-cells, and, strikingly, large numbers of aberrant double-negative αβ T-cells (CD4negCD8neg, DN) and regulatory-like T-cells. These data raise the possibility that a thymic rudiment persisted, allowing T-cell development, albeit with disturbances in positive/negative selection, as suggested by DN and FoxP3+ cell expansions. Although regulatory-like T-cell numbers normalized following HLA-mismatched thymic transplantation, the αβDN subset persisted 5 years post-transplantation. Involution of thymus allograft likely occurred 3 years post-transplantation based on sj/βTREC ratio, which estimates intrathymic precursor T-cell divisions and, consequently, thymic explant output. Nevertheless, functional immune-competence was sustained, providing new insights for the design of immunological reconstitution strategies based on thymic transplantation, with potential applications in other clinical settings.

HLA-NET (a European COST Action) aims at networking researchers working in bone marrow transplantation, epidemiology and population genetics to improve the molecular characterization of the HLA genetic diversity of human populations, with an expected strong impact on both public health and fundamental research. Such improvements involve finding consensual strategies to characterize human populations and samples and report HLA molecular typings and ambiguities; proposing user-friendly access to databases and computer tools and defining minimal requirements related to ethical aspects. The overall outcome is the provision of population genetic characterizations and comparisons in a standard way by all interested laboratories. This article reports the recommendations of four working groups (WG1-4) of the HLA-NET network at the mid-term of its activities. WG1 (Population definitions and sampling strategies for population genetics' analyses) recommends avoiding outdated racial classifications and population names (e.g. 'Caucasian') and using instead geographic and/or cultural (e.g. linguistic) criteria to describe human populations (e.g. 'pan-European'). A standard 'HLA-NET POPULATION DATA QUESTIONNAIRE' has been finalized and is available for the whole HLA community. WG2 (HLA typing standards for population genetics analyses) recommends retaining maximal information when reporting HLA typing results. Rather than using the National Marrow Donor Program coding system, all ambiguities should be provided by listing all allele pairs required to explain each genotype, according to the formats proposed in 'HLA-NET GUIDELINES FOR REPORTING HLA TYPINGS'. The group also suggests taking into account a preliminary list of alleles defined by polymorphisms outside the peptide-binding sites that may affect population genetic statistics because of significant frequencies. WG3 (Bioinformatic strategies for HLA population data storage and analysis) recommends the use of programs capable of dealing with ambiguous data, such as the 'gene[rate]' computer tools to estimate frequencies, test for Hardy-Weinberg equilibrium and selective neutrality on data containing any number and kind of ambiguities. WG4 (Ethical issues) proposes to adopt thorough general principles for any HLA population study to ensure that it conforms to (inter)national legislation or recommendations/guidelines. All HLA-NET guidelines and tools are available through its website http://hla-net.eu. © 2012 Blackwell Publishing Ltd., SCOPUS: re.j, FLWOA, info:eu-repo/semantics/published

Introduction A number of genetic-association studies have identified genes contributing to ankylosing spondylitis (AS) susceptibility but such approaches provide little information as to the gene activity changes occurring during the disease process. Transcriptional profiling generates a 'snapshot' of the sampled cells' activity and thus can provide insights into the molecular processes driving the disease process. We undertook a whole-genome microarray approach to identify candidate genes associated with AS and validated these gene-expression changes in a larger sample cohort. Methods A total of 18 active AS patients, classified according to the New York criteria, and 18 gender- and age-matched controls were profiled using Illumina HT-12 whole-genome expression BeadChips which carry cDNAs for 48,000 genes and transcripts. Class comparison analysis identified a number of differentially expressed candidate genes. These candidate genes were then validated in a larger cohort using qPCR-based TaqMan low density arrays (TLDAs). Results A total of 239 probes corresponding to 221 genes were identified as being significantly different between patients and controls with a P-value SPOCK2 (osteonectin) and EP300, which modulate cartilage and bone metabolism. Conclusions We have validated a gene expression signature for AS from whole blood and identified strong candidate genes that may play roles in both the inflammatory and joint destruction aspects of the disease.

Human monocyte-derived dendritic cells (mo-DCs) express highly sialylated structures, with recognized but poorly understood function in maturation, immunogenicity and endocytosis capacity. We have previously shown that mo-DCs surface sialylation is changeable upon different stimuli, which led us to hypothesise the existence of cell surface (non-intracellular) sialyltransferases, rapidly restoring or altering mo-DC surface sialylation, thus modulating specific functions. Here, we demonstrate that, in the presence of exogenous CMP-Neu5Ac, mo-DCs incorporate considerable amounts of sialic acids into cell surface, predominantly when mo-DCs were previously desialylated or matured. This is a genuine sialyltransferase activity, confirmed by specific inhibition assays, which is not influenced by secreted enzymes. Functionally, the ecto-sialyltransferase activity causes a significant down-regulation of mo-DCs endocytic capacity, without affecting the maturation state. These findings suggest that ecto-sialyltransferases participate in a dynamic control of mo-DC sialylation, with functional repercussions. This activity is possibly related with specific physiological and pathological conditions, as inflammation and infection, contributing to protection and homeostasis regulation.

Background The T antigen is a tumor-associated structure whose sialylated form (the sialyl-T antigen) involves the altered expression of sialyltransferases and has been related with worse prognosis. Since little or no information is available on this subject, we investigated the regulation of the sialyltransferases, able to sialylate the T antigen, in bladder cancer progression. Methods Matched samples of urothelium and tumor tissue, and four bladder cancer cell lines were screened for: ST3Gal.I, ST3Gal.II and ST3Gal.IV mRNA level by real-time PCR. Sialyl-T antigen was detected by dot blot and flow cytometry using peanut lectin. Sialyltransferase activity was measured against the T antigen in the cell lines. Results In nonmuscle-invasive bladder cancers, ST3Gal.I mRNA levels were significantly higher than corresponding urothelium (p < 0.001) and this increase was twice more pronounced in cancers with tendency for recurrence. In muscle-invasive cancers and matching urothelium, ST3Gal.I mRNA levels were as elevated as nonmuscle-invasive cancers. Both non-malignant bladder tumors and corresponding urothelium showed ST3Gal.I mRNA levels lower than all the other specimen groups. A good correlation was observed in bladder cancer cell lines between the ST3Gal.I mRNA level, the ST activity (r = 0.99; p = 0.001) and sialyl-T antigen expression, demonstrating that sialylation of T antigen is attributable to ST3Gal.I. The expression of sialyl-T antigens was found in patients' bladder tumors and urothelium, although without a marked relationship with mRNA level. The two ST3Gal.I transcript variants were also equally expressed, independently of cell phenotype or malignancy. Conclusion ST3Gal.I plays the major role in the sialylation of the T antigen in bladder cancer. The overexpression of ST3Gal.I seems to be part of the initial oncogenic transformation of bladder and can be considered when predicting cancer progression and recurrence.

To ascertain the role and prognostic value of antigen-presenting molecules and chemokines in the prophylactic effect of intravesical bacille Calmette-Guérin (BCG) in tumor recurrence. We compared its gene expression in urothelium biopsy and tumor specimens from patients who had undergone BCG immunotherapy.Patients with nonmuscle-invasive bladder cancer were divided into 3 groups, according to the cancer recurrence status: group 1, primary cancer without recurrence for a minimal period of 12 months; group 2, primary cancer with subsequent recurrence; and group 3, recurrent cancer at study entry. From each patient, cancerous bladder tissue and biopsy specimens of the urothelium (before and 3 months after transurethral resection of the bladder) were collected. The RNA levels of the antigen-presenting molecules CD1a, CD1b, CD1c, CD1d, CD1e, and major histocompatability complex-I, class I (MHC-I) and the chemokines macrophage inflammatory protein-1alpha, monocyte chemoattractant protein-1 and -2, interferon-inducible protein 10 kD (IP10), and monokine induced by gamma-interferon (MIG) were evaluated using real-time polymerase chain reaction on all samples.Generally, BCG treatment increased the urothelium expression of antigen-presenting molecules and chemokines. However, the differences for CD1a (P = .005), CD1b (P.000), CD1c (P = .03), CD1e (P = .007), MHC-I (P.000), MIG (P.0001), and IP10 (P.0001) were significantly superior in the BCG-treated urothelium of group 1 compared with the other groups. Tumor tissue from group 1 also had increased expression of MHC-I (P = .04) and contrasted with tumor tissue from group 3 with decreased expression of CD1c (P = .007) and CD1e (P = .02).Patients without recurrence had greater increased urothelium expression of antigen-presenting molecules and chemokines after BCG treatment. These parameters might, therefore, serve to predict and monitor the efficacy of BCG immunotherapy.

The pharmacogenetic relevance of the tumour necrosis factor α (TNFα) gene has just begun to be investigated. There is only preliminary evidence that the –238 GG genotype is associated with unresponsiveness to conventional disease modifying antirheumatic drugs1 and that a combination of alleles (−308 TNF1/TNF1 and –1087 GG interleukin 10) is associated with good responsiveness to etanercept.2 In addition, Mugnier et al recently suggested that patients with rheumatoid arthritis (RA) with a TNFα −308 GG genotype might, in the short term, be better infliximab responders than patients with AA or AG genotypes.3 To evaluate the influence of the polymorphism at position −308 of the TNFα gene in the long term response to infliximab, we performed a prospective study of 22 consecutive patients with RA who were given infliximab treatment. …