Is Ankylosing Spondylitis a single disease? A clinical and genetic perspective

doi:10.21203/rs.2.22687/v1

Download PDF

Research article

Is Ankylosing Spondylitis a single disease? A clinical and genetic perspective

https://doi.org/10.21203/rs.2.22687/v1

This work is licensed under a CC BY 4.0 License

Version 1

posted

You are reading this latest preprint version

Background:

The aim of the present study was to evaluate whether ankylosing spondylitis (AS) and non-radiographic axial spondyloartritis (nr-axSpA) are subsets of a single disease from a genetic perspective.

Methods:

We analyzed from a clinical and genetic perspective 62 patients from the University Hospital of Basurto (Bilbao, Spain) diagnosed with axial spondyloarthritis. Forty three SNPs previously associated with spondyloarthritis (SpA) were selected. The DNA samples were genotyped through SNP Type Assay, using the BioMark HD platform of Fluidigm.

Results:

Regarding clinical characteristic we found statistically significant differences between the patients with AS and nr-axSpA in the age at diagnosis, the disease duration, the presence of syndesmophytes and the BASMI index. In relation to genetic markers, we only found statistically significant differences in HLA-B27 allele frequency between the two groups. Regarding the frequencies of the SNPs, no statistically significant differences were found between the two groups. Despite the high genetic heterogeneity observed among the patients, it is worth highlighting that some of the most important risk SNPs associated with AS, located in genes (ERAP1, ERAP2, IL-23R, GPR25) and intergenic region (2p15), appeared at high frequencies in all the patients.

Conclusion:

We have observed that AS and nr-axSpA have a common genetic background associated with the pathogenic development of these diseases; therefore, we suggest that the two entities constitute two different expressions of the same disease. Among the genetic factors, the present study shows the importance of genes involved in the pathogenesis of AS, such as HLA-B27, ERAP1, ERAP2, IL-23R, GPR25 and intergenic region 2p15, whose role may influence the onset, development and severity of the disease. However, the pathogenesis of SpA is very complex, indicating the involvement of environmental factors (smoking and obesity), in the triggering of the disease, so that patients with different genotypes would have the same pathogenic phenotype.

Rheumatology

Ankylosing spondylitis

non-radiographic axial spondyloarthritis

SNP

HLA-B27 allele

pathogenesis

Spondyloarthritis (SpA) are a group of rheumatic, inflammatory diseases that share a series of characteristics, such as affected axial skeleton [spine and sacroiliac joints (SIJs)], and peripheral and extra-articular manifestations [1]. SpA traditionally include ankylosing spondylitis (AS), psoriatic arthritis (PsA), reactive arthritis, undifferentiated spondyloarthritis, juvenile spondyloarthritis and inflammatory bowel disease-associated spondyloarthritis (IBD) (Crohn´s disease (CD) and ulcerative colitis) [2].

The classification criteria for the diagnosis of SpA developed by the Assessment of Spondyloarthritis International Society (ASAS) (Table 1), were created and published in 2009 [3,4], due to the diagnostic limitations of the previous criteria [modified New York criteria [5] and European Spondyloarthropathy Study Group criteria [6]]. Based on the ASAS criteria, SpA can be classified as axial (axSpA) or peripheral [3,4]. Within axSpA, there are patients with radiographic changes in SIJs (AS) and other patients without radiographic manifestations (nr-axSpA).

The ASAS classification criteria (image branch) use magnetic resonance imaging (MRI) to diagnose nr-axSpA by detecting the active inflammation (bone marrow edema). However, in the case of AS, conventional x-ray imaging shows advanced and irreversible changes of the sacroiliitis [7], although several years are necessary for these changes to take place, which makes the diagnosis delay years after the beginning of the symptoms [7], thus decreasing the quality of life of the patient.

Sacroiliitis on Imaging + ≥1 SpA Feature OR HLA-B27 + ≥2 Other SpA Features

SpA Features

Sacroiliitis on Imaging

Inflammatory back pain

Arthritis

Enthesitis (heel)

Uveitis

Dactylitis

Psoriasis

Crohn´s disease/ulcerative colitis

Good response to NSAIDs

Family history for SpA

HLA-B27(+)

Elevated CRP

Active (acute) inflammation on MRI highly suggestive of sacroiliitis associated with SpA

Definitive radiographic sacroiliitis according to modified New York criteria

Table 1. The ASAS classification criteria for axial spondyloarthritis in patients with back pain 3 months or more and age at onset younger than 45 years [3,4]. ASAS, Assessment of Spondyloarthritis International Society; SpA, spondyloarthritis; NSAIDs, non-steroidal anti-inflammatory drug; HLA-B27, human leukocyte antigen-B27; CRP, C-reactive protein, MRI, magnetic resonance imaging.

There is current debate on whether nr-axSpA is a different form of AS [8,9], an early form of AS [10,11], or whether both are two expressions of the same disease [12,13]. Some patients with nr-axSpA will develop AS after years of disease [14-17], which occurs in 10% of patients with over 2 years of follow-up, and in over 20% of patients after 2 years with high levels of C-reactive protein (CRP) or an active inflammation detected by MRI [15]. However, other patients with nr-axSpA will suffer the disease for decades, and probably for life, without any evidence of radiographic damage [18], whereas remission will take place in others, either spontaneously or through the administration of drugs.

Patients with AS and nr-axSpA share some clinical characteristics, such as peripheral manifestations (arthritis, enthesitis, dactylitis) and extra-articular manifestations [uveitis, IBD, psoriasis (Ps)] [10-12], with uveitis being slightly more prevalent in AS [19]. Different researchers believe that intestinal dysbiosis plays a fundamental role in the genesis of SpA [20]; in fact, SpA have been associated with IBD, since approximately 5-10% of patients with SpA develop IBD, and approximately 70% may have a subclinical intestinal disease [21]. Furthermore, common genes and loci have been identified for these two diseases [22], as well as the influence of specific alterations in the composition of the gut microbiota with several immune-mediated disorders [23]. Moreover, 10% of patients with AS suffer from Ps, whereas in the case of Ps, 7-42% of patients develop SpA-type axial affectation [24].

Other common characteristics among patients with AS and nr-axSpA are related to the clinical activity, damage, functional deterioration, quality of life [10,12,15,25], similar response to anti-TNF treatment and similar average age in the appearance of symptoms, with small differences in the duration of the disease.

Regarding the differences between AS and nr-axSpA, we can highlight the presence of structural changes in the spine and SIJs, which limit the mobility of patients with AS, whose functionality is influenced by inflammation and the formation of new bone material [26]. Furthermore, several authors have reported a greater number of inflammatory lesions and high levels of CRP in patients with AS [27]. This could be due to the fact that patients with axSpA and high levels of CRP or positive MRI progress more rapidly to radiographic sacroiliitis and, therefore, there may be a larger percentage of patients with radiographic progression if there are signs of inflammation (CRP and MRI) [28]. It has been described that smoking increases the pace and severity of the progression to the radiographic form of SpA [29]. With respect to gender, there is a larger proportion of women among the population of patients with nr-axSpA and a larger percentage of men among patients with AS [10,16,30]. In studies with Asian cohorts, the prevalence of nr-axSpA was twice as high in women with respect to men [31,32], whereas in European and North American cohorts the prevalence of nr-axSpA was similar in both gender [8,11,12]. Therefore, basal radiographic damage, high levels of CRP, smoking and being male are factors that could predict the future radiographic damage in patients with early SpA [33].

The etiopathogenesis of SpA results from the complex interaction between genetic and environmental factors [34]. Although some studies report that genetic factors have a great influence on the susceptibility to develop AS (up to 80-90% in studies with relatives and twins [34]), this type of disease is very complex from the genetic perspective. It has been suggested that allele HLA-B27 contributes to 20-25% of the genetic inheritance of this disease [35], and the single nucleotide polymorphisms (SNPs) found in genome-wide association studies (GWASs) seem to contribute to 3-7% [36]. These data suggest that there may still be many unknown genetic variants that are part of the background of the disease.

Among the SpA, AS has the strongest association with allele HLA-B27 [37,38], since this allele has been found in almost 90% of patients with AS [38]. However, in the general population, only 5% of HLA-B27(+) individuals will eventually develop this disease, with the peculiarity that homozygous for this allele have a greater risk of suffering from AS compared to heterozygous [39], although not all subtypes of HLA-B27 have a predisposition for the disease [40]. Despite the reported association between HLA-B27 and susceptibility to AS, it is currently unknown how HLA-B27 contributes to the development of AS [41]. Different studies have stated that HLA-B27 does not play a direct role in the formation of new bone and cartilage [42,43], and that it is indirectly involved in the inflammatory process [43]. It has been reported that HLA-B27(+) patients have a younger disease onset age and a shorter delay in the diagnosis than HLA-B27(-) patients [30]. Furthermore, it has been suggested that other alleles of the HLA-B system increase or decrease the susceptibility to develop SpA, such as alleles HLA-B07, HLA-B13, HLA-B40, HLA-B47, HLA-B51 or HLA-B57 among others [44,45]. In recent years, thanks to GWAS, alleles of different genes, both inside and outside of the Major Histocompatibility Complex (MHC), have been associated with the susceptibility to develop AS [34,36,46-49].

Some studies indicate that the prevalence of HLA-B27 is similar in patients with nr-axSpA and in those with AS [8,10,11,27,50-52]. However, others indicate that patients with AS have a greater frequency of HLA-B27 [34,36,46-49].

Considering the hypothesis that AS and nr-axSpA are subsets of a single disease with common clinical characteristics and a similar disease burden, the genetic background could be expected to be similar for the two entities. The aim of the present study was to evaluate this hypothesis, through the analysis of a set of risk SNPs localised in genes involved in the pathogenesis of AS, Ps, PsA and IBD in a sample of patients with axSpA.

Patients

DNA was obtained from blood samples of 62 patients of the University Hospital of Basurto (Bilbao, Basque Country, Spain). These samples were supplied through the Basque Biobank for Research (Biobanko, Bioef), along with the informed consent of the patients, following the protocols of the Drug Research Ethics Committee of Euskadi (CEImE, PI2017141).

This study included patients over 18 years of age who suffered from inflammatory back pain and met the New York modified criteria for AS or the ASAS classification criteria for axSpA. Moreover, clinical, demographic and laboratory data were also included (CRP, ESR, HLA-B27). The functional state, the activity of the disease and the spinal and pelvic mobility were evaluated through Bath Ankylosing Spondylitis Functional Index (BASFI), Bath Ankylosing Spondylitis Disease Activity Index (BASDAI) and Bath Ankylosing Spondylitis Metrology Index (BASMI), respectively.

Pelvic plain radiography was evaluated by three rheumatologists and the existence of sacroiliitis, defined according to the New York modified criteria [5], was decided by consensus with the positive opinion of at least 2 of the 3 rheumatologists. The patients fulfilling criteria with radiographic sacroiliitis were diagnosed with AS, whereas those without radiographic criteria, were diagnosed with nr-AxSpA by MRI, according to ASAS image branch criteria [3,4].

Selection of SNPs associated with rheumatic diseases

For the selection of SNPs, a search was carried out in different bibliographic databases: 1) PubMed (http://www.ncbi.nlm.nih.gov/pubmed/), 2) dbSNP (http://www.ncbi.nlm.nih.gov/snp/) of NCBI (National Center for Biotechnology Information, USA), and 3) databases that gather the results obtained in different GWAS analyses (GWAS Catalog, GWAS Central, Ensembl Genome Browser), which identify those allele variations of SNPs associated with the susceptibility to developing a disease.

In this study, we selected those SNPs that had been previously associated with the diagnosis, prognosis and treatment of AS, Ps, PsA and IBD (Supplementary table). These SNPs are located in genes that are involved in different routes and mechanisms that could partially explain the pathogenesis of the disease.

For the selection of the SNPs, the following criteria were applied: (1) GWAS described in Caucasian populations, and (2) the p_value of the association between the SNP and the disease had to be lower than 5*10^-6. Moreover, the SNPs that were in linkage disequilibrium (r²> 0.8) were excluded from the study.

Genotyping

The DNA samples were genotyped through SNP Type Assay, using the BioMark HD platform of Fluidigm (96.96 Dynamic Array^TM IFC, Fluidigm, South San Francisco, CA, USA). The genotyping was carried out at the Sequencing and Genotyping Department of the SGIker General Genomic Services of the University of the Basque Country (UPV/EHU, Bizkaia, Spain). All the SNPs that did not obtain a call rate greater than 0.90 were discarded from the analysis. The genotyping of HLA-B27 was conducted using tagSNP rs11688202 in the mentioned platform. The sensitivity and specificity of this SNP is >98% for the typing of HLA-B27 [49]. The rest of the alleles of the HLA-B gene were analysed through sequencing, following the methodology proposed by Laza et al., 2016 [56].

Statistical analysis

The continuous variables, expressed as mean and standard deviation, were compared using the Student’s T-test, and the results were shown as mean and standard deviation. Regarding the categorical variables, a statistically significant p_value<0.05 was considered for the clinical and laboratory variables, as well as for the allele and genotype frequencies, which were compared using the Fisher’s exact test (applying the Bonferroni correction for multiple comparisons). These analyses were carried out using SPSS Statistics software v25 [57]. The Hardy-Weinberg equilibrium test for the frequencies of the SNPs was conducted using the HardyWeinberg package of RStudio [58,59], considering that there was no equilibrium in those with p_value<0.05 (applying the Bonferroni correction for multiple comparisons). To carry out the Principal Component Analysis (PCA) and Heatmap, we used the FactoMineR [60] and Pheatmap [61] packages of RStudio [59], respectively.

In the present study, 62 patients diagnosed with axSpA were analysed, all of whom had a history of inflammatory back pain. Of these, 49 patients (79%) met the radiographic criteria being diagnosed with AS, and 13 (21%) were diagnosed with nr-axSpA using MRI. Table 2 shows the demographic and clinical characteristics and laboratory parameters of the patients of both groups. The patients of the AS group showed a higher inclusion age and a significantly longer duration of the disease with respect to those of the nr-axSpA group (p_value<0.05), although the symptoms onset age was lower, the diagnostic delay was shorter and the diagnosis age was significantly lower (p_value<0.05). In the group of patients with AS there was a predominant proportion of males (77.55% males vs 22.45% females), whereas in the nr-axSpA group the proportions of males and females were similar (53.85% vs 46.15%). The prevalence of smokers (including ex-smokers) was larger in the AS group compared to the nr-axSpA group (Table 2). The presence of syndesmophytes in the spine was statistically higher in the AS group (p_value<0.05), confirmed by the BASMI index (p_value<0.05), which measures the mobility of the spine and SIJs (Table 2). In the present study, no statistically significant differences were found for peripheral and extra-articular manifestations, although dactylitis, peripheral arthritis and uveitis were mostly diagnosed in the AS group (Table 2).

Regarding the laboratory variables analysed in this study, the levels of CRP were higher among the patients with AS. It is worth highlighting the significantly higher prevalence of allele HLA-B27 in individuals with AS compared to those with nr-axSpA (87.8% vs 38.5%) (Table 2). In the patients who did not have allele HLA-B27 (N=14), the sequencing revealed the presence of alleles HLA-B40 (N=5), HLA-B55 (N=4), HLA-B56 (N=2), HLA-B07 (N=1), HLA-B39 (N=1) and HLA-B47 (N=1), and no statistically significant differences were found between AS and nr-axSpA patients, since 4 of these alleles were detected in patients with AS [HLA-B40 (N=3), HLA-B55 (N=2), HLA-B56 (N=1), HLA-B07 (N=1)), and 5 in those with nr-axSpA (HLA-B40 (N=2), HLA-B55 (N=2), HLA-B56 (N=1), HLA-B39 (N=1), HLA-B47 (N=1)].

*Characteristic*	AS (N = 49)	Nr-axSpA (N=13)	p_value
Current age, mean ± S.D. (years)	51.3 ± 13.5	47.5 ± 12.2	0.360
Male gender (N, %)	38 (77.6%)	7 (53.9%)	0.176
Female gender (N, %)	11 (22.5%)	6 (46.2%)	0.176
Family history (N, %)	15 (30.6%)	5 (38.5%)	0.838
Inclusion age mean ± S.D. (years)	49.7 ± 13.4	45.9 ± 12.0	0.355
Age at diagnosis mean ± S.D. (years)	33.3 ± 13.0	42.2 ± 13.4	0.039*
Age at symptom onset mean ± S.D. (years)	28.6 ± 12.5	32.7 ± 9.9	0.276
Disease duration mean ± S.D. (years)	17.9 ± 13.3	4.4 ± 4.3	2E-07*
Diagnostic delay mean ± S.D. (years)	5.00 ± 5.3	9.7 ± 11.1	0.180
HLA-B27(+) (N, %)	43 (87.8%)	5 (38.5)	0.0007*
CRP mean ± S.D. (mg/L)	21.7 ± 43.1	9.7 ± 10.1	0.367
ESR mean ± S.D. (mm/h)	22.2 ± 21.7	24.1 ± 19.2	0.795
BMI mean ± S.D. (kg/m²)	26.7 ± 4.4	27.0 ± 5.4	0.395
Enthesitis (N, %)	2 (4.1%)	2 (15.4%)	0.401
Dactylitis (N, %)	6 (12.2%)	0 (0%)	0.424
Peripheral arthritis (N, %)	12 (24.5%)	1 (7.7%)	0.348
Uveitis (N, %)	14 (28.6%)	2 (15.4%)	0.542
Psoriasis (N, %)	3 (6.1%)	3 (23.1%)	0.190
IBD (N, %)	2 (4.1%)	3 (23.1%)	0.096
Obesity (N, %)	11 (22.5%)	4 (30.8%)	0.796
Smoker (N, %)	34 (69.4%)	7 (53.9%)	0.470
Presence of syndesmophytes (N, %)	28 (57.1%)	1 (7.7%)	0.004*
Cardiovascular disease (N, %)	12 (24.5%)	3 (23.1%)	0.796
BASDAI mean ± S.D. (score)	2.5 ± 2.1	3.53 ± 2.5	0.138
BASFI mean ± S.D. (score)	2.6 ± 2.4	3.16 ± 2.7	0.448
BASMI mean ± S.D. (score)	1.6 ± 0.5	1.25 ± 0.3	0.0005*

Table 2. Clinical and laboratory parameters of patients diagnosed with AS and nr-axSpA included in the present study, as well as the p_value of the comparison between the two patient groups (*p_value<0.05). AS, ankylosing spondylitis; nr-axSpA, non-radiographic axial spondyloarthritis; HLA-B27, human leukocyte antigen-B27; CRP, C-reactive protein; ESR, erythrocyte sedimentation rate; BMI, body mass index; IBD, inflammatory bowel disease; BASDAI, bath ankylosing spondylitis disease activity index; BASFI, bath ankylosing spondylitis functional index; BASMI, bath ankylosing spondylitis metrology index.

The analysis of the 28 risk SNPs located in genes involved in the pathogenesis to develop AS according to the existing GWAS studies, showed that over 75% of the patients of this study shared 60% of the risk alleles (Figure 1). However, no statistically significant differences were found in the frequencies of the risk alleles of these 28 SNPs between the individuals with AS and those with nr-axSpA, except for allele HLA-B27, whose frequency was significantly higher in the AS group (p_value<0.05) (Table 2).

Considering that some SNPs showed a dominance relationship, thus the existence of a single copy of that allele increases or decreases the risk of developing AS, we analysed the different genotypes of the 28 SNPs associated with AS.

Regarding the genotypic frequencies of these SNPs, no statistically significant differences were found in this study between the patients with AS and those with nr-axSpA. However, Figure 2 shows that some risk alleles in homozygous or heterozygous patients (in the case of showing a dominance relationship) relative to genes involved in important pathogenic routes, such as ERAP1, ERAP2, GPR25, 2p15 or IL-23R, appear at high frequencies in all the individuals of this study. Nevertheless, there is no defined pattern in any of the two pathological entities in terms of risk genotypes, which shows the great genetic heterogeneity underlying these two entities (Figure 2).

A PCA was carried out considering the genotypic frequencies of such markers in the individuals with AS and those with nr-axSpA, obtaining two principal components that explain 19.42% of the variance (Figure 3A). In the first component (10.20% of the variance), the genetic markers with greater correlation were: HLA-B27 (HLA-B27, 0.582), rs7282490 (rs7282490-ICOSLG, -0.316) and rs1801274 (rs1801274-FCGR2A, 0.3). In the second component (9.22% of the variance), the variables with greater correlation were: rs30187 (rs30187-ERAP1, 0.531), rs1128905 (rs1128905-CARD9, -0.383), HLA-B27 (HLA-B27, 0.314) and rs6600247 (rs6600247, RUNX3, -0.3).

When the 62 individuals were represented on the first two components of the PCA (Figure 3B), there was no significant spatial distribution of the individuals of the two entities regarding the genotypic frequencies. On the other hand, there were some common genotypes of SNPs among the analysed individuals. Likewise, no common genetic pattern was found in the patients of each group, which indicates the existence of great intragroup variability.

Given that the presence of SNPs associated with the susceptibility to develop Ps and IBD has been described in patients with AS, which suggests the existence of a common pathogenetic mechanism, we analysed in the patients of this study, 6 SNPs associated with the susceptibility to develop Ps and PsA and 9 SNPs associated with IBD. The analysis of the genotypic frequencies of these SNPs did not reveal any statistically significant difference between the two patient groups analysed in the present study.

A second PCA was conducted considering the genotypes of the SNPs associated with the susceptibility to develop AS, Ps, PsA and IBD, as well as the alleles found for the HLA-B gene, obtaining two principal components that explain 15.61% of the variance (Figure 5A). In the first component of this PCA (8.13% of the variance), the genetic markers with greater correlation were: rs27222427 (rs27222427-NOD2, 0.648), rs30187 (rs30187-ERAP1, -0.474), HLA-B27 (HLA-B27, -0.366) and rs2076756 (rs2076756-NOD2, 0.335). In the second component (7.48% of the variance), the variables with greater correlation were: rs1801274 (rs1801274-FCGR2A, -0.409), rs10865331 (rs10865331-2p15, 0.384), rs6600247 (rs6600247, RUNX3, -0.355) and rs1128905 (rs1128905-CARD9, 0.320).

When the 62 individuals were represented on the first 2 components (Figure 5B), there were no patterns based on the genotypic frequencies of the SNPs associated with AS, Ps, PsA an IBD. Figure 5 shows what was already observed in the first PCA, regarding the genetic heterogeneity underlying these two entities. However, it is worth highlighting two variables that showed a higher correlation values with respect to axis 1; these are two SNPs of the NOD2 gene (rs27221427 and rs2076756), which is a gene that shows a stronger association with CD and whose association with AS has not been described to date (Figure 5A). Moreover, the combination of the risk genotypes of these two SNPs of the NOD2 gene is present in 16 individuals, almost exclusively in the AS group (N=15, 30.61%), appearing only in one nr-axSpA individual (7.69%). These results suggest an association of these alleles with a greater radiographic progression of the disease, although no statistically significant differences were detected between the two groups (p_value>0.0001) due probably to the sample size.

There is current debate on whether nr-axSpA is a different disease from AS [8,9], an early form of AS [10,11] or whether both are two expressions of the same disease [12,13]. In the present study, we analysed the clinical and demographic characteristics, laboratory markers and SNPs associated with the susceptibility to develop AS, Ps, PsA and IBD in 62 patients of the University Hospital of Basurto (Bilbao, Spain), with the aim of determining whether these two entities, apart from sharing clinical characteristics, had a common genetic background.

It was observed that the patients with AS had a lower symptom onset age, shorter diagnostic time and longer duration of the disease compared to the patients with nr-axSpA (Table 2). This could be due to the fact that AS can be more aggressive than nr-axSpA, with a faster radiographic progression, which would imply an earlier symptom onset and diagnosis compared to nr-axSpA. Regarding peripheral manifestations (arthritis, enthesitis, dactylitis), our data confirm those obtained in other studies related to the prevalence of enthesitis in the two patient groups [10-12]. However, the prevalence of peripheral arthritis and dactylitis was greater in AS patients [10-12] (Table 2). With respect to extra-articular manifestations, we found a greater prevalence of uveitis among the patients with AS, as has been reported in other studies [19], which could be due to the fact that uveitis is associated with the duration of the disease, which is longer in AS [62] (Table 2). Nevertheless, the prevalence of Ps and IBD is greater among individuals diagnosed with nr-axSpA, which is in contrast to what has been reported in the literature to date [10-12] (Table 2).

The BASMI index is statistically higher among individuals diagnosed with AS [11,12,25,32] (Table 2), due to the decreased mobility of the axial skeleton as a consequence of the formation of syndesmophytes and even bone bonds and changes in the SIJs [26]. The BASFI and BASDAI indices, which measure the clinical activity, damage and functional deterioration, would be expected to be higher in individuals with AS, given their greater severity and progression [10,11,32]; however, in the present study, higher values of these indices were found in those individuals with nr-axSpA (Table 2). In this study, high levels of CRP were more frequent in patients with AS (Table 2), which could be due to the fact that axSpA patients with high CRP values tend to progress more rapidly to radiographic sacroiliitis [28].

Regarding the gender of the patients, our results confirmed the predominance of males among the individuals diagnosed with AS (77.55%) (Table 2), which could be due to the fact that these patients have a faster progression than females, as well as more structural changes, which in turn, may cause a more severe disability [28]. Another possible influencing factor is physical activity, which, in the case of men usually involves greater mechanic stress, which would increase the inflammatory activity [63]. However, in the group of nr-axSpA patients, the proportion of women (46.15%) was very similar to that of men (53.85%) (Table 2), as has been described in other studies conducted in European and North American cohorts [8,11,12]. The explanation suggested for the differential prevalence between the two entities in terms of gender is that women have a lower radiographic damage and a slower progression to a radiographic state, remaining in the non-radiographic form for longer periods of time [64], suggesting the possible differential role of hormones in the formation of new bone material in patients with SpA.

Some studies carried out in relatives and twins indicate that the genetic component has a fundamental role in the immunopathogenesis of SpA, although the primary trigger is still unknown [34]. Allele HLA-B27, which has been traditionally granted considerable relevance, could contribute to up to 25% of the total inheritance of AS [35]. In the present study, HLA-B27 showed a significantly greater frequency among the patients with AS compared to those with nr-axSpA (87.8% vs 38.5%), which is in line with the results obtained in other studies [12,28,53-55] (Table 2). Furthermore, this greater frequency of HLA-B27 in the AS group confirms the hypothesis that associates the presence of HLA-B27 with a lower onset age that characterize the AS and an earlier diagnosis [30] (Table 2).

Apart from allele HLA-B27, 6 additional alleles of the HLA-B gene were found in the patients of this study (HLA-B39, HLA-B40, HLA-B47, HLA-B55, HLA-B56 and HLA-B07). Alleles HLA-B39, HLA-B40 and HLA-B47 have been previously associated with AS [65-67] and, in the case of HLA-B39, with PsA [68]. However, allele HLA-B07 is associated with a decrease in the risk of developing AS [67] and alleles HLA-B55 and HLA-B56 are not associated with AS. Although the proportion of individuals with other alleles of the HLA-B gene different from HLA-B27 is greater in the group with nr-axSpA with respect to the AS group (61.5% vs 12.2%), both entities share some alleles (HLA-B40, HLA-B55 and HLA-B56). According to these data, it can be stated that there is a predominance of allele HLA-B27 in patients with AS (87.8%); however, the patients with nr-axSpA showed a greater heterogeneity regarding the alleles of the HLA-B gene.

GWAS studies have revealed a considerable number of genes or gene regions that contribute to the susceptibility to develop AS. In our study, we analysed the allele frequencies of 28 risk SNPs, and found no statistically significant differences between the AS and nr-axSpA patients. It was observed that 75% of the individuals of both groups shared a large number of risk alleles (60%) (Figure 1). These results indicate that both pathological entities have a common genetic background, at least at the level of risk SNPs.

At the genotype level, no statistically significant differences were found between the two groups of patients for risk SNPs located in genes involved in the pathogenesis of AS. Some SNPs showed similar genotypes in most of the individuals of the two groups, e.g., ERAP1 (rs30187), ERAP2 (rs10045403), IL-23R (rs11209026), GPR25 (rs41299637), and in intergenic region 2p15 (rs10865331) (Figure 2). Outside of the MHC, it is worth highlighting the ERAP1 gene for its strong association with the susceptibility to develop AS. ERAP1 is an aminopeptidase involved in the cleavage of peptides to a length of 8-9 amino acids, in order for the new peptide segments to be presented by MHC class I molecules, such as HLA-B27 [47]. The SNPs located in the ERAP1 gene only appear to be associated with AS in HLA-B27(+) and HLA-B40(+) patients [65,67]. The ERAP2 is another aminopeptidase, whose association with AS has been described in HLA-B27(+) and HLA-B27(-) patients [49]. The high frequency of risk SNPs of the ERAP-1 and ERAP-2 genes found in the analysed patients could suggest the alteration of the correct functioning of both genes [69], thus influencing the supply of optimum peptides for MHC class I molecules, such as HLA-B27, which could explain their relationship with the development of SpA [48,70].

The IL-23R gene encodes for IL-23 receptor. IL-23 is a key cytokine involved in the differentiation of naive CD4⁺ T-lymphocytes into Th17 lymphocytes, which produce IL-17, IL-6, IL-22, TNF-α and other similar proinflammatory cytokines. Furthermore, other genes of the IL-23 route have been associated with AS, which shows that this route is an important pathway in the pathogenesis of AS [46,48,49]. There are polymorphisms of IL-23R, such as rs11209026, associated with AS [46], IBD [71] and Ps [72]. This SNP has been found with a very high frequency among the patients of this study (Figure 2), which confirms the evidence that IL-23 and its entire pathogenic route are involved in the susceptibility to develop axSpA, granting inflammation a main role in the triggering of the disease.

GWAS studies have shown the association of SNPs of intergenic region 2p15 (rs10865331) with AS, although their role in the pathogenesis of AS is unknown. It has been hypothesised that this region has non-coding RNA species or protein-coding genes unknown to date, which could be involved in the susceptibility to develop AS [47]. G-protein coupled receptor 25 (GPR25), which is associated with AS [49] is strongly expressed in memory T-cells and NK-cells and involved in the positive regulation of B-cell proliferation [73], suggesting a potential mechanism in different autoimmune diseases. Although it is still unknown how the SNPs located in 2p15 (rs10865331) and GPR25 (rs41299637) may influence the pathogenesis of AS, their high prevalence in our study could suggest a key role in the onset and development of axSpA (Figure 2).

The results obtained for SNPs located in genes (ERAP1, ERAP2, IL-23R, GPR25) and gene regions (2p15), along with the different HLA-B gene subtypes, demonstrate their importance in the pathogenesis of axSpA, probably with an essential role in the onset of both AS and nr-axSpA. Despite the fact that the individuals who suffer from different axSpA have common genotypes for some SNPs of genes that are important in the pathogenesis of AS, we observed great genetic heterogeneity within each group and between the two pathological entities, which demonstrates the complex nature of this type of diseases (Figure 3).

Moreover, given that AS, IBD and Ps could have a common pathogenic mechanism, we analysed 15 SNPs associated with the susceptibility to develop IBD, Ps and PsA, finding no statistically significant differences for any of these SNPs between the two groups of patients (AS and nr-axSpA). However, the combination of the risk genotypes of 2 SNPs located in the NOD2 gene associated with CD was found mainly in individuals with AS (AS: 30.7% vs nr-axSpA: 7.7%), which could indicate that these two SNPs are associated with a greater progression of this disease (Figures 4 and 5). The NOD2 gene was identified as the most relevant risk factor of CD [74], although its association with AS had not been described to date. NOD2 is an intracellular pathogen recognition receptor [75] and plays a role in the immune response to bacterial lipopolysaccharides, since it regulates the response of Th17 cells for the elimination of bacteria by inducing the secretion of cytokines IL-23 and IL-1B, which appear in individuals with CD who have a mutation in NOD2 [76]. The results suggest that individuals with these two SNPs of the NOD2 gene will be at greater risk of developing intestinal lesions characteristic of IBD throughout their lives (Figure 4). Furthermore, the presence of these and other SNPs located in genes associated with AS and IBD would confirm the hypothesis about the relevance of intestinal dysbiosis in the genesis of axSpA [20].

Between 20-25% of the known inheritance of AS is attributed to allele HLA-B27, and 3-7% to SNPs identified in GWAS studies [35,36], thus approximately 70% of the inheritance of this pathology could be related to genetic variants that have not been described to date. Missing heritability is a common issue in complex genetic diseases, and it may be caused by multiple factors. It has also been suggested that general heritability is overestimated, with the probability that epigenetic factors, especially environmental factors (smoking and obesity), may be more relevant in the susceptibility to develop AS.

Smoking is an important environmental factor in the inflammatory process of rheumatic diseases, including SpA, and it is a risk factor of cardiovascular disease. In the present study, 66.1% of the patients were smokers or ex-smokers, with a higher proportion of smokers in the individuals diagnosed with AS (69.4%) with respect to those diagnosed with nr-axSpA (53.9%) (Table 2). This suggests that smoking could contribute to increasing the progression to the radiographic form of SpA [29], promoting spine damage and an earlier onset of the inflammatory back pain, which characterises AS [29].

Obesity and overweight are an increasing issue worldwide due to their influence on the development of metabolic, cardiovascular and rheumatic diseases, which increases the mortality and morbidity of patients [77,78]. In our study, it was observed that 61.3% of the patients were obese (BMI>29.99 kg/m²) or overweight (BMI>24.99 kg/m²), with BMI and the prevalence of obesity being similar between patients with AS and those with nr-axSpA (Table 2). Excess adipose tissue in overweight and obese individuals may have immunomodulating properties that affect the course of the disease [79], which could be associated with an increase in the production of proinflammatory cytokynes [78]. Previous studies have reported a greater prevalence of obesity and overweight [80] and a significantly higher BMI in AS patients compared to the healthy population [81]. It has been described that overweight and obese patients with AS have greater functional limitations, greater activity of the disease and a lower response to anti-TNF-α therapies [79,82], which would contribute to increasing physical inactivity and, thus, gaining weight [80,82]. The high prevalence of smoking and obesity or overweight among the individuals analysed in the present study would suggest the relationship between these life habits and the development of these diseases, with inflammation playing a key role in their triggering.

In the SNPs analysed in the present study, we observed that AS and nr-axSpA have a common genetic background associated with the pathogenic development of these diseases; therefore, from the genetic perspective, it could be suggested that the two entities constitute two different expressions of the same disease. However, the pathogenesis of autoimmune diseases, such as SpA, is very complex, and it is the result of intricate mechanisms interaction where the genetic and environmental factors are involved. Among the genetic factors, this study shows the importance of genes and intergenic regions involved in the pathogenesis of AS, such as HLA-B27, ERAP1, ERAP2, IL-23R, GPR25 and 2p15, whose role may influence the onset, development and severity of the disease. Nevertheless, we also observed great genetic heterogeneity among individuals who had the same clinical diagnosis, which could indicate the involvement of exogenous factors in the triggering of the disease, thus patients with different genotypes would have the same pathogenic phenotype. The most relevant of these external factors are: infections by pathogens that alter the intestinal microbiota, endocrine alterations such as overweight and obesity, and unhealthy life habits (e.g., smoking). Numerous studies have granted great relevance to the genetic component in the development of AS; however, this relevance may be overestimated, since environmental factors could be more important than reported to date in the triggering of the disease. Our results should be confirmed in larger observational studies with a larger nr-axSpA population.

Sieper J, Poddubnyy D. Axial spondyloarthritis. Lancet 2017;390(10089):73–84.
Dougados M, Beaten D. Spondylo-arthritis. Lancet 2011;377(9783):2127-37.
Rudwaleit M, Landewé R, van der Heidge D, Listing J, Akkoc N, Brandt J, et al. The development of Assessment of SpondyloArthritis international Society classification criteria for axial spondyloarthritis (part I): classification of paper patients by expert opinion including uncertainty appraisal. Ann Rheum Dis 2009;68(6):770-6.
Rudwaleit M, van der Heijde D, Landewé R, Listing J, Akkoc N, Brandt J, et al. The development of Assessment of SpondyloArthritis international Society classification criteria for axial spondyloarthritis (part II): validation and final selection. Ann Rheum Dis 2009;68(6):777-83.
van der Linden S, Valkenburg HA, Cats A. Evaluation of diagnostic criteria for ankylosing spondylitis. A proposal for modification of the New York criteria. Arthritis Rheum 1984;27(4):361-8.
Dougados M, van der Linden S, Juhlin R, Huiffeldt B, Amor B, Calin A, et al. The European Spondyloarthropathy Study Group preliminary criteria for the classification of spondyloarthropathy. Arthritis Rheum 1991;34(10):1218-27.
Poddubnyy D, Rudwaleit M. Early spondyloarthritis. Rheum Dis Clin N Am 2012;38(2):387-403.
Wallis D, Haroon N, Ayearst R, Carty A, Inman RD. Ankylosing spondylitis and nonradiographic axial spondyloarthritis: part of a common spectrum or distinct diseases? J Rheumatol 2013;40(12):2038–41.
Robinson PC, Wordsworth BP, Reveille JD, Brown M. Axial spondyloarthritis: a new disease entity, not necessarily early ankylosing spondylitis. Ann Rheum Dis 2013;72(2):162–4.
Kiltz U, Baraliakos X, Karakostas P, Igelmann M, Kalthoff L, Klink C, et al. Do patients with non-radiographic axial spondylarthritis differ from patients with ankylosing spondylitis? Arthritis Care Res (Hoboken) 2012;64(9):1415–22.
Ciurea A, Scherer A, Exer P, Bernhard JJ, Dudler J, Beyeler B, et al. Tumor necrosis factor alpha inhibition in radiographic and nonradiographic axial spondyloarthritis: results from a large observational cohort. Arthritis Rheum 2013;65(12):3096–106.
Rudwaleit M, Haibel H, Baraliakos X, Listing J, Märker-Hermann E, Zeidler H, et al. The early disease stage in axial spondylarthritis: results from the GErman SPondyloarthritis Inception Cohort. Arthritis Rheum 2009;60(3):717–27.
Baeten D, Breban M, Lories R, Schett G, Sieper J. Are spondylarthritides related but distinct conditions or a single disease with a heterogeneous phenotype? Arthritis Rheum 2013;65(1):12–20.
Sampaio-Barros PD, Bertolo MB, Kraemer MH, Marques-Neto JF, Samara AM. Undifferentiated spondyloarthropathies: a 2-year follow-up study. Clin Rheumatol 2001;20(3):201-6.
Sampaio-Barros PD, Bortoluzzo AB, Conde RA, Costallat LT, Samara AM, Bertolo MB. Undifferentiated spondyloarthritis: a longterm followup. J Rheumatol 2010;37(6):1195-9.
Poddubnyy D, Rudwaleit M, Haibel H, Listing J, Marker-Hermann E, Zeidler H, et al. Rates and predictors of radiographic sacroiliiitis progression over 2 years in patients with axial spondyloarthritis. Ann Rheum Dis 2011;70(8):1369-74.
Poddubnyy D, Brandt H, Vahldiek J, Spiller I, Song IH, Rudwaleit M, et al. The frequency of non-radiographic axial spondyloarthritis in relation to symptom duration in patients referred because of chronic back pain: results from the Berlin early spondyloartrhitis clinic. Ann Rheum Dis 2012;71(12):1998-2001.
Said-Nahal R, Miceli-Richard C, Berthelot JM, Duché A, Dernis-Labous E, Le Blévec G, et al. The familial form of spondyloarthrotpathy: a clinical study of 115 multiplex families. Groupe Français d´Etude Génétique des Spondylarthropathies. Arthritis Rheum 2000;43(6):1356-65.
de Winter JJ, van Mens LJ, van der Heijde D, Landewé R, Baeten D. Prevalence of peripheral and extra-articular disease in ankylosing spondylitis versus non-radiographic axial spondyloarthritis: a meta-analysis. Arthritis Res Ther 2016;18:196.
Costello ME, Ciccia F, Willner D, Warrington N, Robinson PC, Gardiner , et al. Intestinal dysbiosis in ankylosing spondylitis. Arthritis Rheumatol 2015;67(3):686-91.
Mielants H, De Vos M, Goemaere S, Schelstraete K, Cuvelier C, Goethals K, et al. Intestinal mucosal permeability in inflammatory rheumatic disease. II. Role of disease. J Rheumatol 1991;18(3):394-400.
Jostins L, Ripke S, Weersma RK, Duerr RH, McGovern DP, Hui KY, et al. . Host-microbe interactions have shaped the genetic architecture of inflammatory bowel disease. Nature 2012;491(7422):119–24.
Scher JU, Littman DR, Abramson SB. Microbiome in inflammatory arthritis and human rheumatic diseases. Arth Rheumatol 2016;68(1):35–45.
Gladman DD. Natural history of psoriatic arthritis. Baillieres Clin Rheumatol 1994;8(2):379–94.
Poddubnyy D, Haibel H, Braun J, Rudwaleit M, Sieper J. Brief report: clinical course over two years in patients with early nonradiographical axial spondyloarthritis and patients with ankylosing spondylitis not treated with tumor necrosis factor blockers: results from the German Spondyloarthritis Inception Cohort. Arhtritis Rheumatol 2015;67(9):2369-75.
Machado P, Landewé R, Braun J, Hermann KG, Baker D, van der Heijde D. Both structural damage and inflammation of the spine contribute to impairment of spinal mobility in patients with ankylosing spondylitis. Ann Rheum Dis 2010;69(8):1465–70.
Reveille JD, Witter JP, Weisman MH. Prevalence of axial spondyloarthritis in the United States: estimates from a cross-sectional survey. Arthritis Care Res (Hoboken) 2012;64(6):905–10.
Sieper J, van der Heijde D, Dougados M, Mease PJ, Maksymowych WP, Brown MA, et al. Efficacy and safety of adalimumab in patients with non-radiographic axial spondyloarthritis: results of a randomized placebo-controlled trial (ABILITY-1). Ann Rheum Dis 2013;72(6):815–22.
Poddubnyy D, Haibel H, Listing J, Märker-Hermann E, Zeidler H, Braun J, et al. Cigarette smoking has a dose-dependent impact on progression of structural damage in the spine in patients with axial spondyloarthritis: results from the GErman SPondyloarthritis Inception Cohort (GESPIC). Ann Rheum Dis 2013;72(8):1430–2.
Feldtkeller E, Khan MA, van der Heijde D, van der Linden S, Braun J. Age at disease onset and diagnosis delay in HLA-B27 negative vs. positive patients with ankylosing spondylitis. Rheumatol Int 2003;23(2):61-6.
Jeong H, Yoon JY, Park EJ, Hwang J, Kim H, Ahn JK, et al. Clinical characteristics of nonradiographic axial spondyloarthritis in Korea: a comparison with ankylosing spondylitis. Int J Rheum Dis 2015;18(6):661-8.
Malaviya AN, Kalyani A, Rawat R, Gogia SB. Comparion of patients with ankylosing spondylitis (AS) and non-radiographic axial spondyloarthritis (nr-axSpA) from a single rheumatology clinic in New Delhi. Int J Rheum Dis 2015;18(7):736-41.
Poddubnyy D, Haibel H, Listing J, Märker-Hermann E, Zeidler H, Braun J, et al. Baseline radiographic damage, elevated acute-phase reactant levels, and cigarette smoking status predict spinal radiographic progression in early axial spondylarthritis. Arthritis Rheum 2012;64(5):1388-98.
Brown MA, Kennedy LG, MacGregor AJ, Darke C, Duncan E, Shatford JL, et al. Susceptibility to ankylosing spondylitis in twins: the role of genes, HLA and the environment. Arthritis Rheum 1997;40(10):1823-8.
Reveille JD. Genetics of spondyloarthritis-beyond the MHC. Nat Rev Rheumatol 2012;8(5):296-304.
Ellinghaus D, Jostins L, Spain SL, Cortes A, Bethune J, Han B, et al. Analysis of five chronic inflammatory diseases identifies 27 new associations and highlights disease-specific patterns at shared loci. Nat Genet 2016;48(5):510-8.
Caffrey MF, James DC. Human lymphocyte antigen association in ankylosing spondylitis. Nature 1973;242(5393):121.
Chatzikyriakidou A, Voulgari PV, Drosos AA. What is the new role of HLA-B27 in spondiloarthropathies? Autoimmun Rev 2011;10(8):464-8.
Jaakkola E, Herzberg I, Laiho K, Barnardo MC, Pointon JJ, Kauppi M, et al. Finnish HLA studies confirm the increased risk conferred by HLA-B27 homozygosity in ankylosing spondylitis. Ann Rheum Dis 2006;65(6):775-80.
Khan MA. An update on the genetic polymorphism of HLA-B*27 with 213 alleles encompassing 160 subtypes (and still counting). Curr Rheumatol Rep 2017;19(2):9.
Bowness P. HLA-B27. Annu Rev Immunol 2015;33:29–48.
Lories RJ, Haroon N. Bone formation in axial spondyloarthritis. Best Pract Res Clin Rheumatol 2014;28(5):765–77.
Neerinkx B, Kollnberger S, Shaw J, Lories R. No evidence for a direct role of HLA-B27 in pathological bone formation in axial SpA. RMD Open 2017;3:e000451.
Laza IM, Ventades NG, Hervella M, de-la-Rúa C. Contribution of ancient human remains analysis to the understanding of the variability in HLA-B gene variants in relation to the diagnosis of spondyloarthropathies. J Autoimmun 2018;94:70-82.
Reveille JD, Zhou X, Lee M, Weisman MH, Yi L, Gensler LS, et al. HLA class I and II alleles in susceptibility to ankylosing spondylitis. Ann Rheum Dis 2019;78(1):66-73.
Wellcome Trust Case Control Consortium, Australo-Anglo-American Spondylitis Consortium (TASC), Burton PR, Clayton DG, Cardon LR, Craddock N, et al. Association scan of 14,500 nonsynonymous SNPs in four diseases identifies autoimmunity variants. Nat Genet 2007;39(11):1329–37.
Australo-Anglo-American Spondyloarthritis Consortium (TASC), Reveille JD, Sims A-M, Danoy P, Evans DM, Leo P, et al. Genome-wide association study of ankylosing spondylitis identifies non-MHC susceptibility loci. Nat Genet 2010;42(2):123–7.
Evans DM, Spencer CCA, Pointon JJ, Su Z, Harvey D, Kochan G, et al. Interaction between ERAP1 and HLA-B27 in ankylosing spondylitis implicates peptide handling in the mechanism for HLA-B27 in disease susceptibility. Nat Genet 2011;43(8):761–7.
International Genetics of Ankylosing Spondylitis Consortium (IGAS), Cortes A, Hadler J, Pointon JP, Robinson PC, Karaderi T, et al. Identification of multiple risk variants for ankylosing spondylitis through high-density genotyping of immune-related loci. Nat Genet 2013;45(7):730–8.
van den Berg R, de Hooge M, van Gaalen F, Reijnierse M, Huizinga T, van der Heijde D. Percentage of patients with spondyloarthritis in patient referred because of chronic back pain and performance of classification criteria: experience from the SPondyloArthritis Caught Early (SPACE) cohort. Rheumatology (Oxford) 2013;52(8):1492–9.
Bakland G, Alsing R, Singh K, Nossent JC. Assessment of SpondyloArthritis International Society criteria for axial spondyloarthritis in chronic back pain patients with a high prevalence of HLA-B27. Arthritis Care Res (Hoboken) 2013;65(3):448e53.
Burgos-Vargas R, Wei JCC, Rahman MU, Akkoc N, Haq SA, Hammoudeh M, et al. The prevalence and clinical characteristics of non-radiographic axial spondyloarthritis among patients with inflammatory back pain in rheumatology practices: a multinational multicenter study. Arthritis Res Ther 2016;18(1):132.
Haibel H, Rudwaleit M, Listing J, Heldmann F, Wong RL, Kupper H, et al. Efficacy of adalimumab in the treatment of axial spondylarthritis without radiographically defined sacroiliitis: results of a twelve-week randomized, double-blind, placebo-controlled trial followed by an open-label extension up to week fifty-two. Arthritis Rheum 2008;58(7):1981–91.
Althoff CE, Sieper J, Song IH, Haibel H, Weiss A, Diekhoff T, et al. Active inflammation and structural change in early active axial spondyloarthritis as detected by whole-body MRI. Ann Rheum Dis 2013;72(6):967-73.
Kilic E, Kilic G, Akgul O, Ozgocmen S. Discriminant validity of the Ankylosing Spondylitis Disease Activity Score (ASDAS) in patients with non-radiographic axial spondyloarthritis and ankylosing spondylitis: a cohort study. Rheumatol Int 2015;35(6):981e9.
Laza IM, Hervella M, de-la-Rúa C. Genetic markers in a medieval case of ankylosing spondylitis. J Rheumatol 2016;43(3):679-81.
IBM Corp. Released 2017. IBM SPSS Statistics for Windows, Version 25.0. Armonk, NY: IBM Corp.
Graffelman J. Exploring Diallelic Genetic Markers: The HardyWeinberg Package. Journal of Statistical Software 2015;64(3):1-23.
RStudio Team. RStudio: Integrated Development for R. RStudio, Inc., Boston, MA 2018; URL http://www.rstudio.com/.
Lê, S, Josse, J, Husson, F. FactoMineR: An R Package for Multivariate Analysis. Journal of Statistical Software 2008;25(1):1-18.
Kolde R. pheatmap: Pretty Heatmaps [Internet]. 2015 Available: https://cran.r-project.org/package=pheatmap.
Stolwijk C, Essers I, van Tubergen A, Boonen A, Bazelier MT, De Bruin ML, et al. The epidemiology of extra-articular manifestations in ankylosing spondylitis: a population-based matched cohort study. Ann Rheum Dis 2015;74(7):1373-8.
Ward MM, Reveille JD, Learch TJ, Davis JC Jr, Weisman MH. Occupational physical activities and long-term functional and radiographic outcomes in patients with ankylosing spondylitis. Arthritis Rheum 2008;59(6):822-32.
Poddubnyy D, Sieper J. Similarities and differences between nonradiographic and radiographic axial spondyloarthritis: a clinical, epidemiological and therapeutic assessment. Curr Opin Rheumatol 2014;26(4):377-83.
Robinson WP, van der Linden SM, Khan MA, Rentsch HU, Cats A, Russell A, et al. HLA-Bw60 increases susceptibility to ankylosing spondylitis in HLA-B27+ patients. Arthritis Rheum 1989;32(9):1135–41.
Yamaguchi A, Tsuchiya N, Mitsui H, Shiota M, Ogawa A, Tokunaga K, et al. Association of HLA-B39 with HLA-B27-negative ankylosing spondylitis and pauciarticular juvenile rheumatoid arthritis in Japanese patients. Evidence for a role of the peptide-anchoring B pocket. Arthritis Rheum 1995;38(11):1672-7.
Cortes A, Pulit SL, Leo PJ, Pointon JJ, Robinson PC, Weisman MH, et al. Major histocompatibility complex associations of ankylosing spondylitis are complex and involve further epistasis with ERAP1. Nat Commun 2015;6:7146.
Gladman DD, Anhorn KA, Schachter RK, Mervat H. HLA antigens in psoriatic arthritis. J Rheumatol 1986;13(3):586-92.
Reeves E, Edwards CJ, Elliott T, James E. Naturally occurring ERAP1 haplotypes encode functionally distinct alleles with fine substrate specificity. J Immunol 2013;191(1):35–43.
Dangoria NS, DeLay ML, Kingsbury DJ, Mear JP, Uchnaska-Ziegler B, Ziegler A, et al. HLA-B27 misfolding is associated with aberrant intermolecular disulfide bond formation (dimerization) in the endoplasmic reticulum. J Biol Chem 2002;277(26):23459–68.
Duerr RH, Taylor KD, Brant SR, Rioux JD, Silverberg MS, Daly MJ, et al. A genome-wide association study identifies IL23R as an inflammatory bowel disease gene. Science 2006;314(5804):1461–3.
Cargill M, Schrodi SJ, Chang M, Garcia VE, Brandon R, Callis KP, et al. A large-scale genetic association study confirms IL12B and leads to the identification of IL23R as psoriasis-risk genes. Am J Hum Genet 2007;80(2):273–90.
Ricaño-Ponce I, Zhernakova DV, Deelen P, Luo O, Li X, Isaacs A, et al. Refined mapping of autoimmune disease associated genetic variants with gene expression suggests an important role for non-coding RNAs. J Autoimmun 2016; 68: 62-74.
Hugot JP, Chamaillard M, Zouali H, Lesage S, Cézard JP, Belaiche J, et al. Association of NOD2 leucine-rich repeat variants with susceptibility to Crohn’s disease. Nature 2001;411(6837):599-603.
Strober W, Murray PJ, Kitani A, Watanabe T. Signalling pathways and molecular interactions of NOD1 and NOD2. Nat Rev Immunol 2006;6(1):9–20.
Philpott DJ, Sorbara MT, Robertson SJ, Croitoru K, Girardin SE. NOD proteins: regulators of inflammation in health and disease. Nat Rev Immunol 2014;14(1):9–23.
Adams KF, Schatzkin A, Harris TB, Kipnis V, Mouw T, Ballard-Barbash R, et al. Overweight, obesity, and mortality in a large prospective cohort of persons 50 to 71 years old. N Engl J Med 2006;355(8):763-78.
Versini M, Jeandel PY, Rosenthal E, Shoenfeld Y. Obesity in autoimmune diseases: not a passive bystander. Autoimmun Rev 2014;13(9):981-1000.
Gremese E, Bernardi S, Bonazza S, Nowik M, Peluso G, Massara A, et al. Body weight, gender and response to TNF-α blockers in axial spondyloarthritis. Rheumatology (Oxford) 2014;53(5):875-81.
Maas F, Arends S, van der Veer E, Wink F, Efde M, Bootsma H, et al. Obesity is common in axial spondyloarthritis and is associated with poor clinical outcome. J Rheumatol 2016;43(2):383-7.
Zhang X, Ding W. Association of genetic variants in pentraxin 3 gene with ankylosing spondylitis. Med Sci Monit 2016;22:2911–6.
Durcan L, Wilson F, Conway R, Cunnane G, O´Shea FD. Increased body mass index in ankylosing spondylitis is associated with greater burden of symptoms and poor perceptions of the benefits of exercise. J Rheumatol 2012;39(12):2310-4.
Genetic Analysis of Psoriasis Consortium & the Wellcome Trust Case Control Consortium, Strange A, Capon F, Spencer CC, Knight J, Weale ME, et al. A genome-wide association study identifies new psoriasis susceptibility loci an interaction between HLA-C and ERAP1. Nat Genet 2010;42(11):985-90.
Kenny EE, Pe´er I, Karban A, Ozelius L, Mitchell AA, Ng SM, et al. A genome-wide scan of Ashkenazi Jewish Crohn´s disease suggests novel susceptibility loci. PLoS Genet 2012;8(3):e1002559.
Anderson CA, Boucher G, Lees CW, Franke A, D´Amato M, Taylor KD, et al. Meta-analysis identifies 29 additional ulcerative colitis risk loci, increasing the number of confirmed associations to 47. Nat Genet 2011;43(3):246-52.
Wellcome Trust Case Control Consortium, Burton PR, Clayton DG, Cardon LR, Craddock N, Deloukas P, et al. Genome-wide association study of 14,000 cases of seven common diseases and 3,000 shared controls. Nature 2007;447(7145):661-78.
Tsoi LC, Spain SL, Knight J, Ellinghaus E, Stuart PE, Capon F, et al. Identification of 15 new psoriasis susceptibility loci highlights the role of innate immunity. Nat Genet 2012;44(12):1341-8.
Ellinghaus E, Stuart PE, Ellinghaus D, Nair RP, Debrus S, Raelson JV, et al. Genome-wide meta-analysis of psoriatic arthritis identifies susceptibility locus at REL. J Invest Dermatol 2012;132(4):1133-40.
Liu JZ, van Sommeren S, Huang H, Ng SC, Alberts R, Takahashi A, et al. Association analyses identify 38 susceptibility loci for inflammatory bowel disease and highlight shared genetic risk across populations. Nat Genet 2015;47(9):979-86.
Hüffmeier U, Uebe S, Ekici AB, Bowes J, Giardina E, Korendowych E, et al. Common variants at TRAF3IP2 are associated with susceptibility to psoriatic arthritis and psoriasis. Nat Genet 2010;42(11):996-9.
Stuart PE, Nair RP, Tsoi LC, Tejasvi T, Das S, Kang HM, et al. Genome-wide association analysis of psoriatic arthritis and cutaneous psoriasis reveals differences in their genetic architecture. Am J Hum Genet 2015;97(6):816-36.
Franke A, McGovern DP, Barrett JC, Wang K, Radford-Smith GL, Ahmad T. Genome-wide meta-analysis increases to 71 the number of confirmed Crohn´s diasease susceptibility loci. Nat Genet 2010;42(12):1118-25.

AS: Ankylosing spondylitis

ASAS: Assessment of Spondyloarthritis International Society

axSpA: Axial spondyloarthritis

BASDAI: Bath Ankylosing Spondylitis Disease Activity Index

BASFI: Bath Ankylosing Spondylitis Functional Index

BASMI: Bath Ankylosing Spondylitis Metrology Index

BMI: Body mass index

CD: Crohn disease

CRP: C-reactive protein

ERAP1: Endoplasmic reticulum aminopeptidase 1

ERAP2: Endoplasmic reticulum aminopeptidase 2

ESR: Erythrocyte sedimentation rate

GPR25:

GWAS: Genome-wide association study

HLA-B27: Human leukocyte antigen B27

IBD: Inflammatory bowel disease

IL-23R: G protein-coupled receptor 25

MHC: Major histocompatibility complex

MRI: Magnetic resonance imaging

NOD2: Nucleotide-binding oligomerization domain 2

nr-axSpA: non-radiographic axial spondyloarthritis

NSAID: Non-steroidal anti-inflammatory drug

PCA: Principal component analysis

Ps: Psoriasis

PsA: Psoriatic arthritis

SIJ: Sacroiliac joint

SNP: Single nucleotide polymorphism

SpA: Spondyloarthritis

TNF: Tumor necrosis factor

UC: Ulcerative colitis

Acknowledgments

We thank the investigators of University Hospital of Basurto (Bilbao, Basque Country) who recruited and followed up the patients, the investigators of the Basque Biobank for Research (Biobanko, Bioef) who supplied the DNA samples and the investigators of Sequencing and Genotyping Department of the SGIker General Genomic Services of the University of the Basque Country (UPV/EHU, Bizkaia, Spain) who carried out the sequencing process.

Funding

This work was funded by the Spanish Ministry of Economy, Industry and Competitiveness (GCL2016-79093/P), and grants from the Basque Government to Research Groups of the Basque University System (IT1138-16) and to Imanol Martín Laza (2014_1_326).

Author information

Affiliations

Department of Genetics, Physical Anthropology and Animal Physiology, Faculty of Science and Technology, University of the Basque Country (UPV-EHU); Leioa; Biscay; Basque Country; 48940; Spain. Laza IM, Hervella M, de-la-Rúa C

Department of Rheumatology, Basurto University Hospital, Montevideo Avenue 13-18, 48013 Bilbao, Spain. Blanco Madrigal JM, Galíndez E, Rivera NA, Garcia Vivar ML

Corresponding autor

*Correspondence to IM Laza, [email protected]

Contributions

All authors were involved in drafting the manuscript or revising it critically for important intellectual content, and all authors approved the final version to be published. IML, MH, JMBM, EG, NAR, MLGV and CdR had full access to all of the data in the study and take responsibility for the integrity of the data. IML, MH and CdR carried out the statistical analysis and the accuracy of the data analysis. IML, MH, MLGV and CdR contributed to the study conception and design. IML, MH, JMBM, EG, NAR, MLGV and CdR contributed to the analysis and interpretation of data.

Ethics declarations

Ethics approval and consent to participate

This study was conducted with the approval of the Drug Research Ethics Committee of Euskadi (CEImE, PI2017141). All patients gave their signed informed consent to participate in the study.

Consent for publication

No individual person´s data are present in this manuscript. All data are completely anonymized. All patients gave their signed informed consent to participate in the study.

Competing interests

The author declares that they have no competing interests.

SupplementaryTable.docx

Download PDF

Version 1

posted

You are reading this latest preprint version

Is Ankylosing Spondylitis a single disease? A clinical and genetic perspective

Status:

Version 1

Abstract

Figures

Background

Methods

Results

Discussion

Conclusions

References

Abbreviations

Declarations

Supplementary Files

Status:

Version 1