Th17 cells are implicated in a variety of inflammatory disorders including Spondyloarthritis (SpA). Ankylosing spondylitis (AS), the most common SpA, is genetically associated with HLA-B27 and IL-23R polymorphisms, although the link remains unexplained. In addition to classically folded heterotrimers, HLA-B27 can form β2m-free homodimers (B272). B272, as well as free heavy chains, are able to interact with KIR3DL2. I have shown an expansion of KIR3DL2+ CD4+ T cells in peripheral blood of AS patients and HLA-B27+ healthy controls. Although KIR3DL2+ CD4+ T cells comprise a minority of CD4+ T cells in peripheral blood, KIR3DL2+ CD4+ T cells account for the majority of peripheral blood CD4+ T cell IL-23R expression. KIR3DL2+ CD4+ T cells from synovial fluid are also enriched for IL-23R expression compared to matched peripheral blood samples. Moreover, treatment exposure to rIL-1/23 significantly increases IL-17 production by KIR3DL2+ CD4+ T cells in AS patients but not RA patients and healthy controls. KIR3DL2+ CD4+ T cells from AS patients produce more IL-17 than KIR3DL2+ CD4+ T cells from HLA-B27- healthy controls. KIR3DL2+ CD4+ T cells from AS patients and healthy controls are also enriched for dual production of IL-17 and IFNγ, consistent with the theory that AS is a Th17 or a Th17/1 driven disease. I have shown that naïve CD4+ T cells express KIR3DL2 de novo after activation. Interaction with B272-expressing cells acts to maintain and/or further increase KIR3DL2 expression. Interaction with B272-expressing cells also increases expression of Bcl-2. I propose that interaction of KIR3DL2+ CD4+ T cells with B272 preferentially promotes the survival of these pro-inflammatory cytokine-producing KIR3DL2+ CD4+ T cells in SpA. HD6, a B272-specific monoclonal antibody, decreases IL-17 production by PBMCs from AS patients, making it an attractive candidate therapeutic reagent in the treatment of AS and other HLA-B27-associated SpA.

Psoriatic Arthritis (PsA) is an inflammatory arthritis associated with psoriasis that leads to progressive joint damage. Genetic variants in the Major Histocompatibility Complex (MHC) region on human chromosome 6p, especially Human Leucocyte Antigen (HLA), are the most important genetic risk variants associated with susceptibility to PsA. I aimed to investigate the heritability of PsA and to determine the association between HLA polymorphisms with susceptibility and severity of PsA. I first validated the new CASPAR classification criteria for PsA in patients in with both early and established disease. Subsequently, I demonstrated that PsA as defined by the CASPAR criteria has a high recurrence risk ratio. In a large case-control and family-based association study, I demonstrated that the class I HLA alleles, HLA-C*12/B*38, -B*27 and -C*06/B*57 are associated with increased susceptibility to PsA. HLA class I molecules biologically interact with Killer-cell Immunoglobulin-like Receptors (KIR) on Natural Killer (NK) to influence immune response. I demonstrated that KIR2DS2 and HLA C group 2 and HLA-B Bw4 were associated with PsA susceptibility. Further analyses of PsA cases with Type II psoriasis and with dactylitis suggested that HLA-C*02, -B*27, -B*38 and KIR2DS2 may be markers of musculoskeletal manifestations of PsA. Furthermore, using longitudinal data, I demonstrated that HLA-B*39, -B*27, -A*02 and KIR3DS1 are associated with peripheral joint damage progression whereas the alleles –DQB1*0604, -C*04 and –B*60 are associated with less damage progression. HLA-C*02, -C*12, -DQB1*0609 and KIR2DS1 are associated with higher risk of sacroiliitis, and HLA-B*27 with syndesmophytes. HLA-A*29 was associated with reduced risk of development of both sacroiliitis and syndesmophytes. These studies indicate that HLA alleles and KIR genes are important in PsA susceptibility and severity and suggest that CD8+ T cells and NK cells that modulate the innate and adaptive immune response play an important role in the susceptibility and severity of PsA.

Psoriatic Arthritis (PsA) is an inflammatory arthritis associated with psoriasis that leads to progressive joint damage. Genetic variants in the Major Histocompatibility Complex (MHC) region on human chromosome 6p, especially Human Leucocyte Antigen (HLA), are the most important genetic risk variants associated with susceptibility to PsA. I aimed to investigate the heritability of PsA and to determine the association between HLA polymorphisms with susceptibility and severity of PsA. I first validated the new CASPAR classification criteria for PsA in patients in with both early and established disease. Subsequently, I demonstrated that PsA as defined by the CASPAR criteria has a high recurrence risk ratio. In a large case-control and family-based association study, I demonstrated that the class I HLA alleles, HLA-C*12/B*38, -B*27 and -C*06/B*57 are associated with increased susceptibility to PsA. HLA class I molecules biologically interact with Killer-cell Immunoglobulin-like Receptors (KIR) on Natural Killer (NK) to influence immune response. I demonstrated that KIR2DS2 and HLA C group 2 and HLA-B Bw4 were associated with PsA susceptibility. Further analyses of PsA cases with Type II psoriasis and with dactylitis suggested that HLA-C*02, -B*27, -B*38 and KIR2DS2 may be markers of musculoskeletal manifestations of PsA. Furthermore, using longitudinal data, I demonstrated that HLA-B*39, -B*27, -A*02 and KIR3DS1 are associated with peripheral joint damage progression whereas the alleles –DQB1*0604, -C*04 and –B*60 are associated with less damage progression. HLA-C*02, -C*12, -DQB1*0609 and KIR2DS1 are associated with higher risk of sacroiliitis, and HLA-B*27 with syndesmophytes. HLA-A*29 was associated with reduced risk of development of both sacroiliitis and syndesmophytes. These studies indicate that HLA alleles and KIR genes are important in PsA susceptibility and severity and suggest that CD8+ T cells and NK cells that modulate the innate and adaptive immune response play an important role in the susceptibility and severity of PsA.

BACKGROUND: The Spondyloarthritis Research and Treatment Network (SPARTAN), together with the American College of Rheumatology (ACR) and the Spondylitis Association of America (SAA), published treatment recommendations for ankylosing spondylitis (AS) and non-radiographic axial spondyloarthritis (nr-axSpA) in 2016. The extent to which these recommendations are being followed in clinical practice is unclear. We performed a retrospective chart review focusing on disease activity monitoring, back exercises and DXA scanning in adults with AS. METHODS: We reviewed electronic medical records at a single academic hospital network. The cohort for this study was derived from an existing population of 775 patients identified as having AS or nr-axSpA based on imaging studies, all of whom had 3 or more ICD- 9 or 10 codes for AS (720.x or M45.x). Patients were included if they had at least one rheumatology clinic visit for AS during a three-year study period from July 1, 2016 to June 30, 2019. Data were recorded for the first and last clinic visit within the study period. RESULTS: 358 patients met inclusion criteria. The study population was predominantly male (72.6%) with a mean age of 48.1 years (SD=15.5 years); 84.9% had AS based on review of pelvic or spine radiographs. A total of 661 clinic visits were analyzed. A clinical disease activity measure was recorded at 110/661 (16.6%) visits. RAPID3 was the most frequently used score (57.3%) followed by BASDAI (33.6%) and ASDAS (9.1%). CRP and/or ESR was ordered at 346/661 (52.3%) visits. AS-specific physical therapy (PT) or home exercises for the back were documented in 103/661 (15.6%) visits. A discussion of general exercise unrelated to the back was documented in an additional 153/661 (23.1%) visits. 303/358 patients had two or more visits during the study period. The use of disease activity measures or documentation of AS-specific PT or home exercises did not increase between the first and last visit. 65/358 (18.2%) patients had a DXA scan at any time prior to June 30, 2019. Patients with a DXA were significantly older than those without (58.7 vs. 45.7 years, p<0.001) and more likely to be female (31.6% vs. 13.1%, p<0.0001). 32/358 (8.9%) patients carried a diagnosis of osteoporosis, while 27/358 (7.5%) patients had a history of vertebral fracture. 48.1% of patients with a history of vertebral fracture had a DXA compared to 15.7% of patients without vertebral fracture (p<0.001). CONCLUSION: Monitoring with validated disease activity measures, treatment with AS- specific PT or home exercises, and osteoporosis screening with DXA were performed at low frequencies in our study population. Our data emphasize the need for more education of rheumatologists and the development of implementation strategies along with future updates of the AS/nr-axSpA treatment recommendations. / 2020-12-07T00:00:00Z

BACKGROUND: Spondyloarthritis is a family of related inflammatory diseases including psoriatic arthritis and ankylosing spondylitis. The cytokine IL-23 is known to play an important role in spondyloarthritis development. Overexpression of IL-23 using IL-23 minicircle DNA in B10.RIII mice results in the development of a spondyloarthritis-like disease. B10.RIII is a major histocompatibility complex (MHC) congenic mouse strain that is susceptible to a number of autoimmune and autoinflammatory diseases. Contaminating regions outside the congenic interval from the donor strain, RIII, have been previously detected. While B10.RIII mice develop collagen-induced arthritis (CIA) and collagen antibody induced arthritis (CAIA), the background strain, C57BL/10 (B10) does not. The MHC region is known to play a role in the susceptibility of B10.RIII mice to CIA, but not in CAIA development, so other regions must be involved in arthritis development as well. These contaminating RIII-derived regions may control susceptibility to IL-23 minicircle induced arthritis in B10.RIII mice. OBJECTIVE: This study aimed to identify RIII-derived regions in the genome of B10.RIII mice and begin to interrogate their effects on IL-23 minicircle induced arthritis. METHODS: A systematic literature review was conducted using Pubmed and Web of Science. Articles using B10.RIII mice to study inflammatory disease models were included and data about year of publication, inbred mouse strains used, disease model, and inbred strain notation were extracted. Genome sequences of B10.RIII(71NS)/Sn, C57BL/10SnJ, and C57BL/10J were compared to identify RIII-derived clusters of variation in the B10.RIII genome. B10.RIII mice were crossed with B10 mice and subsequently backcrossed to B10.RIII mice to introduce the B10 allele at chromosome 15. Chr15b/b, Chr15b/r, and Chr15r/r B10.RIII mice were hydrodynamically injected with IL-23 minicircles, and arthritis development was monitored every other day for two weeks. RESULTS: The systematic literature review yielded 8 studies that compared arthritis development in B10.RIII to RIII or B10. These studies identified three arthritis susceptibility regions: Cia5/Eae3 on chromosome 3, Eae2 on chromosome 15, and Eae39 on chromosome 5. Eae2 is further split into four sub-regions: Cia30, Cia31, Cia32, and Cia26. Genome sequence comparison identified RIII-derived clusters in B10.RIII mice on chromosomes 10, 14, 15, and 17. The chromosome 15 region overlaps with the Eae2 susceptibility region for approximately 17 Mbp and includes the arthritis susceptibility loci Cia26 and Cia32. When this region was interrogated in vivo, Chr15r/r and Chr15b/r B10.RIII mice developed IL-23 minicircle arthritis, while Chr15b/b mice did not. CONCLUSION: The B10.RIII(71NS)/Sn strain contains several large RIII/WySn-derived regions outside the congenic MHC region. One of these clusters on chromosome 15 includes the arthritis susceptibility loci Cia26 and Cia32 and appears to determine susceptibility to IL-23 minicircle induced arthritis. Future studies will interrogate the role of the chromosome 15 RIII-derived region in arthritis development in more detail and aim to identify the specific gene variants that control arthritis susceptibility. / 2023-01-28T00:00:00Z

Ankylosing spondylitis (AS) is associated with HLA-B*2704 and B*2705 but not with HLA-B*2706 and B*2709. Genome wide studies recently identified ERAP1 as an important genetic association in AS and could be the missing link in the pathogenesis of AS. I studied the implications of the two known actions of ERAP1 on AS pathogenesis. For assessing the peptide trimming function, surface HLA-B27 and MHC-I free heavy chain (FHC) expression on peripheral blood mononuclear cells of AS patients were studied. Subsequently, in an in vitro system of C1R cells expressing different AS-associated and AS-neutral HLA-B27 subtypes, I studied the effect of ERAP1 suppression on HLA-B27 and FHC expression. To assess the cytokine receptor shedding function, I studied serum cytokine receptor level variation with ERAP1 polymorphisms and its relationship to disease activity in AS patients. Finally, I studied the effect of variants of ERAP1 and other members of the antigen presentation machinery on radiographic severity in AS patients. AS patients with the major allele of the ERAP1 rs27044 polymorphism had higher FHC expression on monocytes. In C1R cells ERAP1 suppression led to an increase in intracellular FHC (IC-FHC) and B27-peptide complexes identified by a special MARB4 antibody, but only in C1R cells expressing the AS-associated subtypes HLA-B*2704 and B*2705. ERAP1 variants had no effect on serum cytokine receptor levels. Baseline radiographic severity was associated with ERAP1 polymorphism in univariate analysis only. LMP2 variants were associated with baseline radiographic severity in multivariate analysis. ERAP1 affects peptide presentation and FHC formation by HLA-B27 and could be the missing link in the pathogenesis of AS. ERAP1 through its differential HLA-B27 subtype interaction could explain why certain subtypes of HLA-B27 are associated with AS while others are not. Larger studies are required to look closely at the effect of ERAP1 on radiographic severity and progression in AS.

Ankylosing spondylitis (AS) is associated with HLA-B*2704 and B*2705 but not with HLA-B*2706 and B*2709. Genome wide studies recently identified ERAP1 as an important genetic association in AS and could be the missing link in the pathogenesis of AS. I studied the implications of the two known actions of ERAP1 on AS pathogenesis. For assessing the peptide trimming function, surface HLA-B27 and MHC-I free heavy chain (FHC) expression on peripheral blood mononuclear cells of AS patients were studied. Subsequently, in an in vitro system of C1R cells expressing different AS-associated and AS-neutral HLA-B27 subtypes, I studied the effect of ERAP1 suppression on HLA-B27 and FHC expression. To assess the cytokine receptor shedding function, I studied serum cytokine receptor level variation with ERAP1 polymorphisms and its relationship to disease activity in AS patients. Finally, I studied the effect of variants of ERAP1 and other members of the antigen presentation machinery on radiographic severity in AS patients. AS patients with the major allele of the ERAP1 rs27044 polymorphism had higher FHC expression on monocytes. In C1R cells ERAP1 suppression led to an increase in intracellular FHC (IC-FHC) and B27-peptide complexes identified by a special MARB4 antibody, but only in C1R cells expressing the AS-associated subtypes HLA-B*2704 and B*2705. ERAP1 variants had no effect on serum cytokine receptor levels. Baseline radiographic severity was associated with ERAP1 polymorphism in univariate analysis only. LMP2 variants were associated with baseline radiographic severity in multivariate analysis. ERAP1 affects peptide presentation and FHC formation by HLA-B27 and could be the missing link in the pathogenesis of AS. ERAP1 through its differential HLA-B27 subtype interaction could explain why certain subtypes of HLA-B27 are associated with AS while others are not. Larger studies are required to look closely at the effect of ERAP1 on radiographic severity and progression in AS.

Background: MicroRNAs (miRNAs) are small non-coding single-stranded RNAs involved in the posttranscriptional inhibition of gene expression and thereby regulating all cellular functions. Their dysregulation contributes to the pathophysiology of many diseases, including rheumatic diseases. MiRNAs can also be found extracellularly in body fluids and represent promising diagnostic and prognostic biomarkers. Our study aimed to investigate miRNAs as biomarkers of stage and activity and predictors of therapeutic response of two most common inflammatory rheumatic diseases: spondyloarthritis (SpA) and rheumatic arthritis (RA). Results: We found several circulating miRNAs differentially expressed in SpA patients reflecting the severity of axial involvement and/or disease activity. The decrease in circulating miR-145 in plasma of patients with ankylosing spondylitis 3 months of anti-TNF therapy predicted a good therapeutic response and low disease activity after a year of therapy. Circulating and intracellular expression of miR-125b in peripheral blood mononuclear cells (PBMC) was lower in treatment-naïve patients with early RA than in healthy controls. Baseline expression of miR-125 in PBMC predicted a (non)adequate therapeutic response. We also found the increased expression of miR-451 in PBMC in...

Die axiale Spondyloarthritis (axSpA) ist eine chronische entzündlich-rheumatische Erkrankung unbekannter Ursache, die durch Entzündungen in den Sakroiliakalgelenken (SIG) und an den Gelenken der Wirbelsäule gekennzeichnet ist. Darüber hinaus kann es im Krankheitsverlauf zu einer Ankylose in den SIG und zur Entwicklung von Syndesmophyten an den Wirbelkörpern kommen. Da ein Großteil der axSpA-Patienten auch sub-klinische mukosale Entzündungen aufweist, werden mukosale Antigene als Trigger der Entzündung diskutiert. Für die Diagnose und Prognose der axSpA existieren bisher wenige serologische Marker mit hoher Sensitivität. In dieser Arbeit konnte gezeigt werden, dass die Serumkonzentrationen von CTX-II, BMP-2 und LBP ein hohes diagnostisches Potenzial für die axSpA aufweisen, während die Serumkonzentrationen von BMP-2, PINP und VEGF als Marker für die Vorhersage röntgenolgischer Progression geeignet sein könnten. Weiterhin konnten erhöhte LPS-, LBP- und IL-6-Serum-Konzentrationen bei axSpA-Patienten nachgewiesen werden, die auf eine Translokation bakterieller Antigene hinweisen. Die Charakterisierung der Monoyzten zeigte erhöhte Frequenzen der CD14++CD16- und einen verminderten Anteil an CD14++CD16+ Monozyten in Patienten mit axSpA. Funktionell wiesen die Monozyten von axSpA-Patienten eine in vivo Präaktivierung mit erhöhter spontaner und durch suboptimale bakterielle Stimuli induzierter Freisetzung proinflammatorischer Zytokine bei gleichzeitig verminderter Reaktivität auf LPS in vitro auf. Diese Präaktivierung war bei Patienten unter Standardtherapie, nicht aber unter TNF-Blocker-Therapie, nachweisbar. Interessanterweise bestand bei Patienten unter Standardtherapie ein Zusammenhang zwischen der Krankheitsaktivität und der Frequenz zytokinproduzierender Monozyten. Somit konnten mit dieser Arbeit Biomarker mit diagnostischer und prognostischer Bedeutung für die axSpA identifiziert werden, deren Bedeutung in unabhängigen Kohorten weiter untersucht werden muss. / Axial Spondyloarthitis (axSpA) is a chronic inflammatory-rheumatic disease of unknown cause. It is characterized by inflammations of the sacroiliac joints (SIG) and the spinal joints. In addition an ankylosis in the SIG can develop in the progression of the disease as well as syndesmophytes at the vertebral body. Since the majority of axSpA-patients also have sub-clinical mucosal inflammations, mucosal anti-gens are discussed as triggers of the inflammation. For the diagnosis and prognosis of axSpA there are only a few serological markers with high sensitivity and specificity until now. In this thesis it could be shown that the serum concentration of CTX-II, BMP-2 and LBP exhibit a high diagnostic potential for axSpA, while the serum concentration of BMP-2, PINP and VEGF could be suitable markers for the forecast of radiographic progression. Furthermore increased LPS-, LBP- and IL-6-serum concentrations could be verified, which can be an indicator for the translocation of bacterial antigenes. The monocyte characterization showed increased frequencies of the pro-inflammatory CD14++CD16- sub population and a reduced portion of CD14++CD16+ monocytes in patients with axSpA. Functionally the monocytes of axSpA-patients showed an in vivo pre-activation with increased spontaneous and by suboptimal bacterial stimuli induced release of pro-inflammatory cytokines with simultaneously reduced reactivity towards LPS in vitro. This pre-activation could be detected for patients undergoing standard therapy, but not for those under TNF-blocker-therapy. Interestingly for the standard therapy patients there was a connection between the activity of the disease, meaning the BASDAI, and the frequency of the cytokine-producing monocytes. Therefore biomarkers with diagnostic and prognostic relevance could be identified in line with this thesis. The significance of these biomarkers has to be researched further in independent cohorts.

La Spondyloarthrite (SpA) est un rhumatisme inflammatoire chronique fréquent, avec une prévalence de 0,43 % en France. Elle consiste en une atteinte prédominante du squelette axial, mais aussi des articulations périphériques, et peut conduire à une immobilité du rachis et des articulations sacro-iliaques. Des atteintes extra-articulaires sont fréquentes, telles qu'une uvéite, un psoriasis ou une maladie inflammatoire chronique de l'intestin. Les traitements actuels ne sont que symptomatiques, ciblant principalement les manifestations inflammatoires. L'étiologie de la SpA est multifactorielle avec une composante génétique dominée par l'association forte et bien connue avec l'allèle HLA-B27. Cependant, ce facteur génétique n'est clairement pas suffisant pour induire le développement de la maladie. L'objectif de ce projet de thèse était donc d'identifier d'autres facteurs génétiques à l'origine du développement de la SpA.Mon travail a porté sur l'analyse de deux jeux de données complémentaires, dans une perspective de biologie des systèmes. Dans une première partie, j'ai conduit une analyse de liaison dans 210 familles atteintes de la maladie représentant 1310 personnes génotypées avec des puces Affymetrix 250k. Une nouvelle région significativement liée à la SpA a été détectée en 13q13, avec un intervalle de 1,3 Mb défini par des haplotypes recombinants chez les patients.Ensuite, une analyse transcriptomique des cellules dendritiques dérivées des monocytes de 23 patients HLA-B27+, 23 témoins sains HLA-B27+ et 21 témoins sains HLA-B27-, et stimulées ou non par du LPS, a tenté de distinguer les gènes dont l'expression est modifiée par la maladie de ceux influencés par l'allèle HLA-B27 seul. L'annotation fonctionnelle et une analyse par réseau de gènes ont mis en évidence l'inhibition chez les patients des étapes précoces de la biosynthèse du cholestérol. / Spondyloarthritis is a frequent chronic inflammatory rheumatism, with a prevalence of 0.43 % in France. This disease presents axial skeleton injuries, but also on peripheral joints, and can results in a total spinal and sacro-iliac motility loss. Extra-articular features including uveitis, psoriasis and inflammatory bowel disease are frequent. Current SpA treatments are only symptomatic, relieving inflammatory symptoms. SpA etiology is largely multifactorial with a genetic component dominated by the long-known strong association with the HLA-B27 allele. This allele, however, is not sufficient for the disease to occur. This thesis project objective was then to identify other genetic factors in the origin of SpA.My work was mainly divided in two complementary data analyses, in a way to get a systems biology approach. The first one consisted in proceed linking analyses on data from Affymetrix genotyping chips gathered from DNA of 1310 people grouped in 210 families. This study allowed notably to detect a new significantly linked region to SpA : 13q13, with an interval of 1.3 Mb. This part of genome is currently being sequenced to allow a better causal SNP identification.Secondly, an Affymetrix HumanGene 1.0 st transcriptomic chips analysis was performed on MD-DCs extracted from 68 people, stimulated or not by LPS during 6 or 24 hours. This cohort was grouped between 23 patients HLA-B27+, 23 healthy controls HLA-B27+ and 21 healthy controls HLA-B27-. I could notice that HLA-B27 allele is farly enough to considerably affect cell transcriptomic profiles, which encourages to include HLA-B27+ healthy controls. Otherwise, a gene network analysis allowed me to highlight on an inhibition of early steps of cholesterol biosyntthesis.