Human inflammatory dendritic cells induce Th17 cell differentiation.

Dendritic cells (DCs) are critical regulators of immune responses. Under noninflammatory conditions, several human DC subsets have been identified. Little is known, however, about the human DC compartment under inflammatory conditions. Here, we characterize a DC population found in human inflammatory fluids that displayed a phenotype distinct from macrophages from the same fluids and from steady-state lymphoid organ and blood DCs. Transcriptome analysis showed that they correspond to a distinct DC subset and share gene signatures with in vitro monocyte-derived DCs. Moreover, human inflammatory DCs, but not inflammatory macrophages, stimulated autologous memory CD4(+) T cells to produce interleukin-17 and induce T helper 17 (Th17) cell differentiation from naive CD4(+) T cells through the selective secretion of Th17 cell-polarizing cytokines. We conclude that inflammatory DCs represent a distinct human DC subset and propose that they are derived from monocytes and are involved in the induction and maintenance of Th17 cell responses.

HLA-B27 alters BMP/TGFß signalling in Drosophila, revealing putative pathogenic mechanism for spondyloarthritis.

OBJECTIVES: The human leucocyte antigen (HLA)-B27 confers an increased risk of spondyloarthritis (SpA) by unknown mechanism. The objective of this work was to uncover HLA-B27 non-canonical properties that could explain its pathogenicity, using a new Drosophila model. METHODS: We produced transgenic Drosophila expressing the SpA-associated HLA-B*27:04 or HLA-B*27:05 subtypes, or the non-associated HLA-B*07:02 allele, alone or in combination with human ß2-microglobulin (hß2m), under tissue-specific drivers. Consequences of transgenes expression in Drosophila were examined and affected pathways were investigated by the genetic interaction experiments. Predictions of the model were further tested in immune cells from patients with SpA. RESULTS: Loss of crossveins in the wings and a reduced eye phenotype were observed after expression of HLA-B*27:04 or HLA-B*27:05 in Drosophila but not in fruit flies expressing the non-associated HLA-B*07:02 allele. These HLA-B27-induced phenotypes required the presence of hß2m that allowed expression of well-folded HLA-B conformers at the cell surface. Loss of crossveins resulted from a dominant negative effect of HLA-B27 on the type I bone morphogenetic protein (BMP) receptor saxophone (Sax) with which it interacted, resulting in elevated mothers against decapentaplegic (Mad, a Drosophila receptor-mediated Smad) phosphorylation. Likewise, in immune cells from patients with SpA, HLA-B27 specifically interacted with activin receptor-like kinase-2 (ALK2), the mammalian Sax ortholog, at the cell surface and elevated Smad phosphorylation was observed in response to activin A and transforming growth factor ß (TGFß). CONCLUSIONS: Antagonistic interaction of HLA-B27 with ALK2, which exerts inhibitory functions on the TGFß/BMP signalling pathway at the cross-road between inflammation and ossification, could adequately explain SpA development.

OX40L blockade protects against inflammation-driven fibrosis.

Treatment for fibrosis represents a critical unmet need, because fibrosis is the leading cause of death in industrialized countries, and there is no effective therapy to counteract the fibrotic process. The development of fibrosis relates to the interplay between vessel injury, immune cell activation, and fibroblast stimulation, which can occur in various tissues. Immunotherapies have provided a breakthrough in the treatment of immune diseases. The glycoprotein OX40-OX40 ligand (OX40L) axis offers the advantage of a targeted approach to costimulatory signals with limited impact on the whole immune response. Using systemic sclerosis (SSc) as a prototypic disease, we report compelling evidence that blockade of OX40L is a promising strategy for the treatment of inflammation-driven fibrosis. OX40L is overexpressed in the fibrotic skin and serum of patients with SSc, particularly in patients with diffuse cutaneous forms. Soluble OX40L was identified as a promising serum biomarker to predict the worsening of lung and skin fibrosis, highlighting the role of this pathway in fibrosis. In vivo, OX40L blockade prevents inflammation-driven skin, lung, and vessel fibrosis and induces the regression of established dermal fibrosis in different complementary mouse models. OX40L exerts potent profibrotic effects by promoting the infiltration of inflammatory cells into lesional tissues and therefore the release of proinflammatory mediators, thereafter leading to fibroblast activation.

Exploring antibody-dependent adaptive immunity against aortic extracellular matrix components in experimental aortic aneurysms.

BACKGROUND: Recent evidence suggests that adaptive immunity develops during abdominal aortic aneurysm evolution. Uncertainties remain about the antigens implicated and their role in inducing rupture. Because antigens from the extracellular matrix (ECM) have been suspected, the aim of this experimental study was to characterize the role of adaptive immunity directed against antigens from the aortic ECM. METHODS: In a first step, an experimental model of abdominal aortic aneurysm rupture based on adaptive immunity against the ECM was developed and characterized. Forty 4-week-old male Lewis rats were divided into two groups. In the ECM group (n = 20), rats were presensitized against the guinea pig aortic ECM before implantation of a decellularized aortic xenograft (DAX). In the control group (n = 20), rats were not presensitized before DAX implantation. In each group, half the rats were sacrificed at day 3 to analyze early mechanisms involved after DAX implantation. In a second step, we aimed to assess which ECM component was most efficient in inducing rupture. For this purpose, the nonfibrillar and fibrillar ECM components were sequentially extracted from the guinea pig aortic wall. Forty Lewis rats were then divided into four groups. Each group was presensitized against one ECM component (structural glycoproteins and proteoglycans, collagen, elastin alone, and elastin-associated glycoproteins) before DAX implantation. Apart from those that experienced rupture, rats were sacrificed at day 21. Xenografts were harvested for histologic, immunofluorescence, and conditioned medium analyses. RESULTS: In total, early aortic rupture occurred in 80% of the ECM group vs 0% of the control group (P < .001). In the ECM group, major circumferential immunoglobulin deposits were observed in combination with the C3 complement fraction, without cell infiltration. Conditioned medium analysis revealed that matrix metalloproteinase 9 and myeloperoxidase levels and elastase activities were significantly increased in this group. Immunofluorescence analysis demonstrated that myeloperoxidase co-localized with tissue-free DNA and histone H4, highlighting local neutrophil activation and formation of neutrophil extracellular traps. Following differential presensitization, it appeared that rats presensitized against structural glycoproteins and proteoglycans were significantly more susceptible to rupture after DAX implantation. CONCLUSIONS: Stimulating adaptive immunity against the aortic ECM, especially structural glycoproteins and proteoglycans, triggers rupture after DAX implantation. Further studies are needed to assess the precise proteins involved.

Faecal microbiota study reveals specific dysbiosis in spondyloarthritis.

OBJECTIVE: Altered microbiota composition or dysbiosis is suspected to be implicated in the pathogenesis of chronic inflammatory diseases, such as spondyloarthritis (SpA) and rheumatoid arthritis (RA). METHODS: 16S ribosomal RNA gene sequencing was performed on faecal DNA isolated from stool samples in two consecutive cross-sectional cohorts, each comprising three groups of adult volunteers: SpA, RA and healthy controls (HCs). In the second study, HCs comprised a majority of aged-matched siblings of patients with known HLA-B27 status. Alpha and beta diversities were assessed using QIIME, and comparisons were performed using linear discriminant analysis effect size to examine differences between groups. RESULTS: In both cohorts, dysbiosis was evidenced in SpA and RA, as compared with HCs, and was disease specific. A restriction of microbiota biodiversity was detected in both disease groups. The most striking change was a twofold to threefold increased abundance of Ruminococcus gnavus in SpA, as compared with both RA and HCs that was significant in both studies and positively correlated with disease activity in patients having a history of inflammatory bowel disease (IBD). Among HCs, significant difference in microbiota composition were also detected between HLA-B27+ and HLA-B27 negative siblings, suggesting that genetic background may influence gut microbiota composition. CONCLUSION: Our results suggest that distinctive dysbiosis characterise both SpA and RA and evidence a reproducible increase in R. gnavus that appears specific for SpA and a marker of disease activity. This observation is consistent with the known proinflammatory role of this bacteria and its association with IBD. It may provide an explanation for the link that exists between SpA and IBD.

A family-based genome-wide association study reveals an association of spondyloarthritis with MAPK14.

OBJECTIVE: More than 40 loci have been associated with ankylosing spondylitis (AS), but less is known about genetic associations in spondyloarthritis (SpA) as a whole. We conducted a family-based genome-wide association study (GWAS) to identify new non-major histocompatibility complex (MHC) genetic factors associated with SpA. METHODS: 906 subjects from 156 French multiplex families, including 438 with SpA, were genotyped using Affymetrix 250K microarrays. Association was tested with Unphased. The best-associated non-MHC single nucleotide polymorphisms (SNPs) were then genotyped in two independent familial cohorts (including 215 French and 294 North American patients with SpA, respectively) to replicate associations. RESULTS: 43 non-MHC SNPs yielded an association signal with SpA in the discovery cohort (p<1×10-4). In the extension studies, association was replicated at a nominal p value of p<0.05 for 16 SNPs in the second cohort and for three SNPs in the third cohort. Combined analysis identified an association close to genome-wide significance between rs7761118, an intronic SNP of MAPK14, and SpA (p=3.5×10-7). Such association appeared to be independent of HLA-B27. CONCLUSIONS: We report here for the first time a family-based GWAS study on SpA and identified an associated polymorphism near MAPK14. Further analyses are needed to better understand the functional basis of this genetic association.

Prevention of autoimmunity and control of recall response to exogenous antigen by Fas death receptor ligand expression on T cells.

Mice with mutations in the gene encoding Fas ligand (FasL) develop lymphoproliferation and systemic autoimmune diseases. However, the cellular subset responsible for the prevention of autoimmunity in FasL-deficient mice remains undetermined. Here, we show that mice with FasL loss on either B or T cells had identical life span as littermates, and both genotypes developed signs of autoimmunity. In addition, we show that T cell-dependent death was vital for the elimination of aberrant T cells and for controlling the numbers of B cells and dendritic cells that dampen autoimmune responses. Furthermore, we show that the loss of FasL on T cells affected the follicular dentritic cell network in the germinal centers, leading to an impaired recall response to exogenous antigen. These results disclose the distinct roles of cellular subsets in FasL-dependent control of autoimmunity and provide further insight into the role of FasL in humoral immunity.

Modulatory role of the anti-apoptotic protein kinase CK2 in the sub-cellular localization of Fas associated death domain protein (FADD).

The Fas associated death domain protein (FADD) is the key adaptor molecule of the apoptotic signal triggered by death receptors of the TNF-R1 superfamily. Besides its crucial role in the apoptotic machinery, FADD has proved to be important in many biological processes like tumorigenesis, embryonic development or cell cycle progression. In a process to decipher the regulatory mechanisms underlying FADD regulation, we identified the anti-apoptotic kinase, CK2, as a new partner and regulator of FADD sub-cellular localization. The blockade of CK2 activity induced FADD re-localization within the cell. Moreover, cytoplasmic FADD was increased when CK2ß was knocked down. In vitro kinase and pull down assays confirmed that FADD could be phosphorylated by the CK2 holoenzyme. We found that phosphorylation is weak with CK2α alone and optimal in the presence of stoichiometric amounts of CK2α catalytic and CK2ß regulatory subunit, showing that FADD phosphorylation is undertaken by the CK2 holoenzyme in a CK2ß-driven fashion. We found that CK2 can phosphorylate FADD on the serine 200 and that this phosphorylation is important for nuclear localization of FADD. Altogether, our results show for the first time that multifaceted kinase, CK2, phosphorylates FADD and is involved in its sub-cellular localization. This work uncovered an important role of CK2 in stable FADD nuclear localization.

Whole-genome single nucleotide polymorphism-based linkage analysis in spondyloarthritis multiplex families reveals a new susceptibility locus in 13q13.

OBJECTIVE: Spondyloarthritis (SpA) is a chronic inflammatory disorder with high heritability but with complex genetics. Apart from HLA-B27, most of the underlying genetic components remain to be identified. We conducted a whole-genome high-density non-parametric linkage analysis to identify new genetic factors of susceptibility to SpA. METHODS: 914 subjects including 462 with SpA from 143 multiplex families were genotyped using Affymetrix 250K microarrays. After quality control, 189 368 single nucleotide polymorphisms (SNPs) were kept for further analyses. Both non-parametric and parametric linkage analyses were performed using Merlin software. Association was tested with Unphased. RESULTS: Non-parametric linkage analysis identified two regions significantly linked to SpA: the major histocompatibility complex (LODmax=24.77) and a new 13q13 locus (LODmax=5.03). Additionally, eight loci achieved suggestive LOD scores, including the previously identified SPA2 locus at 9q33 (LODmax=3.51). Parametric analysis supported a codominant model in 13q13 with a maximum heterogeneity LOD, 'HLOD' score of 3.084 (α=0.28). Identification of meiotic recombination events around the 13q13 linkage peak in affected subjects from the 43 best-linked families allowed us to map the disease interval between 38.753 and 40.040 Mb. Family-based association analysis of the SNPs inside this interval in the best-linked families identified a SNP near FREM2 (rs1945502) which reached a p value close to statistical significance (corrected p=0.08). CONCLUSION: We report here for the first time a significant linkage between 13q13 and SpA. Identification of susceptibility factor inside this chromosomal region through targeted sequencing in linked families is underway.

Synergy of chemotherapy and immunotherapy revealed by a genome-scale analysis of murine tuberculosis.

OBJECTIVES: Although TB immunotherapy improves the results of conventional drug treatment, the effects of combining chemotherapy and immunotherapy have never been systematically evaluated. We used a comprehensive lung transcriptome analysis to directly compare the activity of combined chemotherapy and immunotherapy with that of single treatments in a mouse model of TB. METHODS: Mycobacterium tuberculosis-infected mice in the chronic phase of the disease (day 30) received: (i) isoniazid and rifampicin (drugs) daily for 30 days; (ii) DNA immunotherapy (DNA), consisting of four 100 µg injections at 10 day intervals; (iii) both therapies (DNA + drugs); or (iv) saline. The effects were evaluated 10 days after the end of treatment (day 70 post-infection). RESULTS: In all groups a systemic reduction in the load of bacilli was observed, bacilli became undetectable in the drugs and DNA + drugs groups, but the whole lung transcriptome analysis showed 867 genes exclusively modulated by the DNA + drugs combination. Gene enrichment analysis indicated that DNA + drugs treatment provided synergistic effects, including the down-regulation of proinflammatory cytokines and mediators of fibrosis, as confirmed by real-time PCR, ELISA, histopathology and hydroxyproline assay. CONCLUSIONS: Our results provide a molecular basis for the advantages of TB treatment using combined chemotherapy and DNA immunotherapy and demonstrate the synergistic effects obtained with this strategy.

Prevalence of spondyloarthritis in reference to HLA-B27 in the French population: results of the GAZEL cohort.

OBJECTIVE: To estimate the prevalence of spondyloarthritis (SpA) in reference to HLA-B27 in the French population. METHODS: In 1989, 20 625 employees of the French national gas and electricity company aged 35-50 years were enrolled in the GAZEL cohort. In 2010, 18 757 still active participants were screened by a questionnaire validated for the detection of SpA. Responders with available DNA were retained for further studies. Pelvic radiograph and HLA-B27 typing were performed in all the self-reported cases of SpA or psoriatic arthritis. Self-reported diagnosis was verified by a qualified rheumatologist. HLA-B27 determination was also performed in subjects without any SpA feature. RESULTS: The target population consisted of 6556 responders with available DNA. Their male:female ratio was 3.6 and their mean age was 65.5±3.3 years. A diagnosis of SpA was confirmed in 32 of the 72 self-reported cases, 75% of them being HLA-B27 positive. Estimated SpA prevalence adjusted for sex was 0.43% (95% CI 0.26% to 0.70%). HLA-B27 positivity rate in 2466 healthy controls was 6.9% (95% CI 5.9% to 7.9%). The relative risk of SpA in HLA-B27 positive individuals was 39 (95% CI 17 to 86). CONCLUSIONS: We estimated the prevalence of SpA in the French population in 2010 to 0.43%. With an estimated prevalence of 75.0% in SpA and 6.9% in healthy controls, HLA-B27 increased the disease risk 39-fold, as compared with HLA-B27 negative subjects.

Identification of secreted phosphoprotein 1 gene as a new rheumatoid arthritis susceptibility gene.

OBJECTIVE: To evaluate the contribution of the SPP1 rs11439060 and rs9138 polymorphisms, previously reported as autoimmune risk variants, in the rheumatoid arthritis (RA) genetic background according to anti-citrullinated protein antibodies (ACPAs) status of RA individuals. METHODS: We analysed a total of 11,715 RA cases and 26,493 controls from nine independent cohorts; all individuals were genotyped or had imputed genotypes for SPP1 rs11439060 and rs9138. The effect of the SPP1 rs11439060 and rs9138 risk-allele combination on osteopontin (OPN) expression in macrophages and OPN serum levels was investigated. RESULTS: We provide evidence for a distinct contribution of SPP1 to RA susceptibility according to ACPA status: the combination of ≥3 SPP1 rs11439060 and rs9138 common alleles was associated mainly with ACPA negativity (p=1.29×10(-5), ORACPA-negative 1.257 (1.135 to 1.394)) and less with ACPA positivity (p=0.0148, ORACPA-positive 1.072 (1.014 to 1.134)). The ORs between these subgroups (ie, ACPA-positive and ACPA-negative) significantly differed (p=7.33×10(-3)). Expression quantitative trait locus analysis revealed an association of the SPP1 risk-allele combination with decreased SPP1 expression in peripheral macrophages from 599 individuals. To corroborate these findings, we found an association of the SPP1 risk-allele combination and low serum level of secreted OPN (p=0.0157), as well as serum level of secreted OPN correlated positively with ACPA production (p=0.005; r=0.483). CONCLUSIONS: We demonstrate a significant contribution of the combination of SPP1 rs11439060 and rs9138 frequent alleles to risk of RA, the magnitude of the association being greater in patients negative for ACPAs.

Upholding the T cell immune-regulatory function of CD31 inhibits the formation of T/B immunological synapses in vitro and attenuates the development of experimental autoimmune arthritis in vivo.

CD31, a trans-homophilic inhibitory receptor expressed on both T- and B-lymphocytes, drives the mutual detachment of interacting leukocytes. Intriguingly, T cell CD31 molecules relocate to the immunological synapse (IS), where the T and B cells establish a stable interaction. Here, we show that intact CD31 molecules, which are able to drive an inhibitory signal, are concentrated at the periphery of the IS but are excluded from the center of the IS. At this site, were the cells establish the closest contact, the CD31 molecules are cleaved, and most of the extracellular portion of the protein, including the trans-homophilic binding sites, is shed from the cell surface. T cells lacking CD31 trans-homophilic binding sites easily establish stable interactions with B cells; at the opposite, CD31 signaling agonists inhibit T/B IS formation as well as the ensuing helper T cell activation and function. Confocal microscopy and flow cytometry analysis of experimental T/B IS shows that the T cell inhibitory effects of CD31 agonists depend on SHP-2 signaling, which reduces the phosphorylation of ZAP70. The analysis of synovial tissue biopsies from patients affected by rheumatoid arthritis showed that T cell CD31 molecules are excluded from the center of the T/B cell synapses in vivo. Interestingly, the administration of CD31 agonists in vivo significantly attenuated the development of the clinical signs of collagen-induced arthritis in DBA1/J mice. Altogether, our data indicate that the T cell co-inhibitory receptor CD31 prevents the formation of functional T/B immunological synapses and that therapeutic strategies aimed at sustaining CD31 signaling will attenuate the development of autoimmune responses in vivo.

Revisiting MHC genes in spondyloarthritis.

Spondyloarthritis (SpA) refers to a variety of inflammatory rheumatic disorders with strong heritability. Shared genetic predisposition, as shown by familial aggregation, is largely attributable to the major histocompatibility complex (MHC) locus, which was estimated to account for approximately half of the whole disease heritability. The first predisposing allele identified more than 40 years ago is HLA-B27, which is a major gene predisposing to all forms of SpA. However, despite intensive research, its pathogenesis remains uncertain. Other MHC alleles belonging to the class I and class II regions have been identified to exert additional effect. Candidate-gene approaches and genome-wide studies have recently allowed identification of several new loci residing outside of the MHC region that are involved in the predisposition to SpA. Interestingly, some of those new genes, such as ERAP1, ERAP2, and NPEPPS, code for aminopeptidases that are involved in MHC class I presentation and were shown to interact with HLA-B27.

Brief report: the IL23R nonsynonymous polymorphism rs11209026 is associated with radiographic sacroiliitis in spondyloarthritis.

OBJECTIVE: Spondyloarthritis (SpA) is a group of inflammatory articular disorders sharing a genetic background. The nonsynonymous single-nucleotide polymorphism (SNP) rs11209026 (Arg381Gln) in the IL23R gene has reproducibly been shown to be associated with ankylosing spondylitis (AS). We undertook this study to examine the association between rs11209026 and SpA as a whole, with particular attention devoted to genotype/phenotype correlation. METHODS: The SNP rs11209026 was genotyped in a French cohort of 415 patients/372 controls, with replication analysis performed in 383 "trios," each consisting of 1 patient with SpA and both parents. Association analysis was carried out in SpA as a whole group and then separately in AS and non-AS patients. Phenotype/genotype correlations were examined using logistic regression analysis. RESULTS: A significant association between rs11209026 and SpA overall was identified only in the familial data set (odds ratio 0.57, P=0.028). Strong association with AS was observed in both the case-control and familial data sets (P=4.5×10(-4) and P=4.0×10(-3), respectively). In contrast, such association was not detected in the non-AS group. Furthermore, rs11209026 frequency was significantly different between AS and non-AS patients (P=1.5×10(-3)). Phenotype/genotype correlation study revealed that both radiographic sacroiliitis and early age at onset were independently associated with a lower frequency of the rare protective rs11209026 allele A in patients (P=9×10(-3) and P=8×10(-3), respectively). CONCLUSION: Our study replicated the robust association between rs11209026 and AS in the French population. However, such association was restricted to AS, as compared to SpA without radiographic sacroiliitis. The fact that it was independently conditional on radiographic sacroiliitis and age at onset suggests that rs11209026 could affect disease severity rather than susceptibility.

A regulatory variant in CCR6 is associated with susceptibility to antitopoisomerase-positive systemic sclerosis.

OBJECTIVE: Recognition of the well-known pleiotropism of autoimmune genes supports the concept of a shared pathogenesis across autoimmune diseases such as rheumatoid arthritis (RA) and systemic sclerosis (SSc). Studies have reproducibly demonstrated an association between susceptibility to RA and polymorphisms of the CCR6 gene, a surface marker for Th17 cells, and the causal variant was recently identified. The present study was thus undertaken to investigate whether CCR6 polymorphisms could also be associated with susceptibility to SSc. METHODS: Twelve tag single-nucleotide polymorphisms (SNPs) of CCR6, including the known RA-associated SNP rs3093023, were genotyped in a total of 2,411 SSc patients and 7,084 healthy individuals from 3 European populations (France, Italy, and Germany). Meta-analyses of the data were performed to assess whether an association exists between CCR6 polymorphisms and susceptibility to SSc or its main subtypes. Direct sequencing of DNA was performed to ascertain whether the functional dinucleotide polymorphism of CCR6 previously identified in RA (CCR6DNP) was also present in SSc. RESULTS: Combined analyses revealed an association between the rs10946216 SNP and SSc susceptibility (odds ratio [OR] 1.13, 95% confidence interval [95% CI] 1.05-1.21, adjusted P [P(adj)] = 0.026). The rs3093023 A allele and rs10946216 T allele were in high linkage disequilibrium, and both were found to confer disease susceptibility in the antitopoisomerase-positive subset of SSc patients (OR 1.27, 95% CI 1.13-1.42, P(adj) = 1.5 × 10(-3) and OR 1.32, 95% CI 1.17-1.48, P(adj) = 9.0 × 10(-5), respectively, relative to healthy controls). Direct sequencing of the DNA of 78 individuals supported the hypothesis that the regulatory dinucleotide CCR6DNP could be the causal variant in SSc. CONCLUSION: The results of this study establish CCR6 as a new susceptibility factor for antitopoisomerase-positive SSc, as demonstrated in 3 European Caucasian populations, confirming the notion that SSc and RA could conceivably share autoimmune risk alleles. The results also suggest a potential role of the interleukin-17 pathway in SSc.

Investigating the genetic association between ERAP1 and spondyloarthritis.

OBJECTIVE: A robust association between polymorphisms in the non-major histocompatibility complex gene ERAP1 and ankylosing spondylitis (AS) in several populations was recently identified. The aim of the current study was to determine the level of association of ERAP1 polymorphisms with spondyloarthritis (SpA) in French/Belgian populations with particular attention to genotype-phenotype correlations. METHODS: We studied 734 independent SpA cases and 632 controls from two European cohorts. Five single-nucleotide polymorphisms (SNPs), rs27044, rs17482078, rs10050860, rs30187 and rs2287987 were genotyped, and case-control association analyses were carried using PLINK 1.07 software. Linkage disequilibrium and haplotypes were estimated with Haploview. Analysis was first carried out in SpA as a whole group, and then separately in AS and non-radiographic SpA (non-AS) patients. RESULTS: Consistent with previous studies conducted in AS, rs30187 was the most significantly associated SNP with SpA (p=0.008 in the French, and p=6.46×10(-4) in the Belgian cohorts). In the combined cohorts, this SNP was associated with both AS and non-AS (P(combined)= 3.9×10(-5) and P(combined)= 0.005, respectively). A similar trend was observed with other SNPs. The rs17482078/rs10050860/rs30187-CCT haplotype was significantly associated with increased risk of SpA in both cohorts (P(combined)= 9.08×10(-4)), including AS and non-AS (P(combined)=6.16×10(-4) and P(combined)=0.049, respectively), whereas the -TTC haplotype was associated with reduced risk of SpA, including AS and non-AS (P(combined)=2.36×10(-7), P(combined)= 5.69×10(-6) and P(combined)= 2.13×10(-4), respectively). CONCLUSIONS: This is the first study to show an association between several polymorphisms located in ERAP1 and SpA as a whole. Our findings demonstrate consistent association of the same SNPs and haplotypes with both AS and non-AS subtypes of SpA.

Critical role of the adhesion receptor DNAX accessory molecule-1 (DNAM-1) in the development of inflammation-driven dermal fibrosis in a mouse model of systemic sclerosis.

OBJECTIVE: To investigate the contribution of the adhesion receptor DNAX accessory molecule-1 (DNAM-1) in the development of dermal fibrosis on gene inactivation and targeted molecular strategies. METHODS: Human skin expression of DNAM-1 was determined by immunohistochemistry. Mice deficient for DNAM-1 (dnam1-/-) and wild-type controls (dnam1+/+) were injected with bleomycin or NaCl. Infiltrating leucocytes, T cells, B cells and monocytes were quantified and inflammatory cytokines were measured in lesional skin of dnam1-/- and dnam1+/+ mice. The anti-fibrotic potential of a DNAM-1 neutralising monoclonal antibody (mAb) was evaluated in the mouse model of bleomycin-induced dermal fibrosis. RESULTS: Overexpression of DNAM-1 was detected in the skin of patients with SSc (systemic sclerosis). Dnam1-/- mice were protected from bleomycin-induced dermal fibrosis with reduction of dermal thickening (75±5%, p=0.03), hydroxyproline content (46±8%, p=0.04) and myofibroblast counts (39±5%, p=0.01). Moreover, the number of T cells was significantly decreased in lesional skin of dnam1-/- mice (69±15%, p=0.0007). Dnam1-/- mice also displayed decreased levels of TNF-α and IL-6 in lesional skin. Consistent with the gene inactivation strategy, treatment of mice with DNAM-1 neutralising mAb prevented dermal fibrosis induced by bleomycin with reduction of dermal thickness (64±6%, p=0.002), hydroxyproline content (61±8%, p=0.004) and myofibroblast counts (83±12%, p=0.002). CONCLUSIONS: An inactivation gene strategy showed that DNAM-1 exerts profibrotic effects by controlling T cell activation and cytokine release. A molecular targeted strategy confirmed that DNAM-1 neutralising mAb has potent antifibrotic properties, supporting the hypothesis that inhibition of DNAM-1 might be a promising new approach for the treatment of SSc and potentially other related fibrotic diseases.

FADD: a regulator of life and death.

FAS-associated protein with death domain (FADD) is the key adaptor protein transmitting apoptotic signals mediated by the main death receptors (DRs). Besides being an essential instrument in cell death, FADD is also implicated in proliferation, cell cycle progression, tumor development, inflammation, innate immunity, and autophagy. Recently, many of these new functions of FADD were shown to be independent of DRs. Moreover, FADD function is dictated by protein localization and phosphorylation state. Thus, FADD is a crucial and unique controller of many essential cellular processes. The full understanding of the networks dictating the ultimate function of FADD may provide a new paradigm for other multifaceted proteins.

Proinflammatory Th17 cells are expanded and induced by dendritic cells in spondylarthritis-prone HLA-B27-transgenic rats.

OBJECTIVE: HLA-B27/human ß2-microglobulin-transgenic (B27-transgenic) rats, a model of spondylarthritis (SpA), develop spontaneous colitis and arthritis under conventional conditions. CD4+ T cells are pivotal in the development of inflammation in B27-transgenic rats. This study was undertaken to characterize the phenotype of CD4+ T cells in this model and to determine whether dendritic cells (DCs) induce proinflammatory T cells. METHODS: The phenotype of CD4+ T cells from rat lymph nodes (LNs) draining the sites of inflammation was analyzed by flow cytometry. Immunostaining was used to detect interleukin-17 (IL-17)-producing cells in the rat joints. DCs from B27-transgenic or control rats (transgenic for HLA-B7 or nontransgenic) were cocultured with control CD4+ T cells and stimulated with anti-T cell receptor α/ß. RESULTS: IL-17A- and tumor necrosis factor α (TNFα)-producing CD4+ T cells were expanded in mesenteric and popliteal LNs from B27-transgenic rats. The accumulation of Th17 cells correlated with disease development, in contrast to Th1 or Treg cells. IL-17-positive mononuclear cells were detected in the arthritic joints of B27-transgenic rats but not in the joints of control rats. Finally, in vitro cocultures demonstrated that Th17 cells were preferentially induced and expanded by DCs from B27-transgenic rats, by a process that may involve defective engagement of costimulatory molecules. CONCLUSION: Our findings indicate that expanded CD4+ T cells in B27-transgenic rats exhibit a proinflammatory Th17 phenotype characterized by IL-17A and TNFα production. Furthermore, this population is preferentially induced by DCs from B27-transgenic rats. These data point toward an induction of Th17 cells as a possible pathogenic mechanism in this model of SpA. However, their pathogenic role still needs to be shown.