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Additional file 2: Supplementary Figure 2. Comparisons of pentameric C-reactive protein (pCRP) (A–C), monomeric (m)CRP (D–F), and mCRP/pCRP ratio (G–I) demonstrated between normal vs. subnormal estimated glomerular filtration rate (eGFR) (A, D, G), presence vs. absence of hematuria (B, E, H) and presence vs. absence albuminuria (C, F, I) in patients with ANCA-associated vasculitis. Abnormal eGFR

Additional file 3: Supplementary Figure 3. Standard curves and reactivity of the monoclonal detection antibody 8C10 towards monomeric C-reactive protein (mCRP) and pentameric (p)CRP: (A) standard curve for 8C10 antibody with mCRP; (B) reactivity of 8C10 antibody with pCRP; and (C) standard curve for samples in the present study.

Additional file 1: Supplementary Figure 1. Comparisons of pentameric C-reactive protein (pCRP) (A–D), monomeric (m)CRP (E–H), and mCRP/pCRP ratio (I–L) in 160 patients with systemic lupus erythematosus with lupus nephritis vs. no nephritis (A, E, I), normal vs. subnormal estimated glomerular filtration rate (eGFR) (B, F, J), presence vs. absence of hematuria (C, G, K) and presence vs. absence of albuminuria (D, H, L). Abnormal eGFR

Additional file 3: Supplementary Figure 3. Standard curves and reactivity of the monoclonal detection antibody 8C10 towards monomeric C-reactive protein (mCRP) and pentameric (p)CRP: (A) standard curve for 8C10 antibody with mCRP; (B) reactivity of 8C10 antibody with pCRP; and (C) standard curve for samples in the present study.

BackgroundThere is a need for biomarker to identify patients ‘at risk’ for rheumatoid arthritis (risk-RA) and to better predict the therapeutic response and in this study we tested the hypothesis that novel native and citrullinated heterogeneous nuclear ribonucleoprotein (hnRNP)-DL autoantibodies could be possible biomarkers.MethodsUsing Protein macroarray and ELISA, epitope recognition against hnRNP-DL was analysed in sera from different developed RA disease and diagnosed SLE patients. Toll-like receptor (TLR) 7/9 and myeloid differentiation primary response gene 88 (MyD88)-dependency were studied in sera from murine disease models. HnRNP-DL expression in cultivated cells and synovial tissue was analysed by indirect immunofluorescence, immunoblot and immunohistochemistry. ResultshnRNP-DL was highly expressed in stress granules, citrullinated in the rheumatoid joint and targeted by autoantibodies either as native or citrullinated proteins in patient subsets with different developed RA disease. Structural citrullination dependent epitopes (SCEs) of hnRNP-DL were detected in 58% of the SLE patients although 98% of these sera were α-CCP2-negative. To obtain a specific citrullinated signal value, we subtracted the native antibody value from the citrullinated signal. This CNDL (Citrullinated-Native-hnRNP-DL)-index identified and the bioinformatic value was explored, as a new value for an “individual window of treatment success” in early RA and for the detection of RF-IgM/α-CCP2 seronegative RA patients (24-46%). Negative CNDL-index was found in SLE patients, risk-RA- and early RA-cohorts such as EIRA where the majority of these patients are DAS28-responders to methotrexate (MTX) treatment (87%). High positive CNDL-values were associated with more severe RA, shared epitope and parenchymal changes in the lung. Specifically, native α-hnRNP-DL is TLR7/9-dependent, associated with pain and ROC-analysis revealed an association to initial MTX or etanercept treatment response, especially in seronegative RA patients.ConclusionCNDL-index defines patients is a possible biomarker for develop RA and the “window of treatment success” thereby potentially closing the sensitivity gap.

Supplemental material, sj-pdf-1-lup-10.1177_09612033211014570 for Associations with thrombosis are stronger for antiphosphatidylserine/prothrombin antibodies than for the Sydney criteria antiphospholipid antibody tests in SLE by Sahwa Elbagir, Giorgia Grosso, NasrEldeen A Mohammed, Amir I Elshafie, Elnour M Elagib, Agneta Zickert, Vivek Anand Manivel, Eleftheria Pertsinidou, Musa A. M Nur, Iva Gunnarsson, Johan Rönnelid and Elisabet Svenungsson in Lupus

Supplemental material, sj-pdf-2-lup-10.1177_09612033211014570 for Associations with thrombosis are stronger for antiphosphatidylserine/prothrombin antibodies than for the Sydney criteria antiphospholipid antibody tests in SLE by Sahwa Elbagir, Giorgia Grosso, NasrEldeen A Mohammed, Amir I Elshafie, Elnour M Elagib, Agneta Zickert, Vivek Anand Manivel, Eleftheria Pertsinidou, Musa A. M Nur, Iva Gunnarsson, Johan Rönnelid and Elisabet Svenungsson in Lupus

Seropositive rheumatoid arthritis (RA) is characterized by the presence of rheumatoid factor (RF) and anti-citrullinated protein autoantibodies (ACPA) with different fine-specificities. Yet, other serum anti-modified protein autoantibodies (AMPA), e.g. anti-carbamylated (Carb), -acetylated (KAc), and malondialdehyde acetaldehyde (MAA) modified protein antibodies, have been described. In this comprehensive study, we analyze 30 different IgG and IgA AMPA reactivities to Cit, Carb, KAc, and MAA antigens detected by ELISA and autoantigen arrays in N=1985 newly diagnosed RA patients. Association with patient characteristics such as smoking and disease activity were explored. Carb and KAc reactivities by different assays were primarily seen in patients also positive for anti-citrulline reactivity. Modified vimentin (mod-Vim) peptides were used for direct comparison of different AMPA reactivities, revealing that IgA AMPA recognizing mod-Vim was mainly detected in subsets of patients with high IgG anti-Cit-Vim levels and a history of smoking. IgG reactivity to acetylation was mainly detected in a subset of patients with Cit and Carb reactivity. Anti-acetylated histone reactivity was RA-specific and associated with high anti-CCP2 IgG levels, multiple ACPA fine-specificities, and smoking status. This reactivity was also found to be present in CCP2+ RA-risk individuals without arthritis. Our data further demonstrate that IgG autoreactivity to MAA was increased in RA compared to controls with highest levels in CCP2+ RA, but was not RA-specific, and showed low correlation with other AMPA. Anti-MAA was instead associated with disease activity and was not significantly increased in CCP2+ individuals at risk of RA. Notably, RA patients could be subdivided into four different subsets based on their AMPA IgG and IgA reactivity profiles. Our serology results were complemented by screening of monoclonal antibodies derived from single B cells from RA patients for the same antigens as the RA cohort. Certain CCP2+ clones had Carb or Carb+KAc+ multireactivity, while such reactivities were not found in CCP2- clones. We conclude that autoantibodies exhibiting different patterns of ACPA fine-specificities as well as Carb and KAc reactivity are present in RA and may be derived from multireactive B-cell clones. Carb and KAc could be considered reactivities within the "Cit-umbrella" similar to ACPA fine-specificities, while MAA reactivity is distinctly different.

Supplemental material, sj-pdf-1-lup-10.1177_0961203320945387 for High IgA antiphospholipid autoantibodies in healthy Sudanese explain the increased prevalence among Sudanese compared to Swedish systemic lupus erythematosus patients by Sahwa Elbagir, Amir I Elshafie, Elnour M Elagib, NasrEldeen A Mohammed, Mawahib IE Aledrissy, Vivek Anand Manivel, Eleftheria Pertsinidou, Musa AM Nur, Iva Gunnarsson, Elisabet Svenungsson and Johan Rönnelid in Lupus

Additional file 3: Supplementary Table 3. Baseline characteristics in CCP2-positive and -negative RA. The table show baseline characteristics for the EIRA patients included in the study, based on anti-CCP2 IgG status; p-values indicate differences between subsets with respect to: age, female-to-male ratio, number of smokers, HLA-DRB1 SE-positivity, PTPN22-positivity, DAS28 and CRP.

Additional file 7: Supplementary Table 6. Associations between different ACPA fine-specificities and HLA-DRB1 SE, in anti-CCP2-positive and anti-CCP2-negative RA. Odds ratios with 95% confidence intervals are shown for associations between HLA-DRB1 SE and presence/absence of 19 different ACPA fine-specificities, in anti-CCP2-positive and -negative RA: p-values indicate differences in ORs between ACPA fine-specificity-positive and -negative subsets in the anti-CCP2-positive subset.

Additional file 6: Supplementary Table 5. Associations between different autoantibodies and PTPN22 polymorphism, in anti-CCP2-negative RA. Odds ratios with 95% confidence intervals are shown for associations between PTPN22 polymorphism and presence/absence of ACPA, RF, other autoantibodies, or "any autoantibody" in anti-CCP2-negative RA.

Additional file 6: Supplementary Table 5. Associations between different autoantibodies and PTPN22 polymorphism, in anti-CCP2-negative RA. Odds ratios with 95% confidence intervals are shown for associations between PTPN22 polymorphism and presence/absence of ACPA, RF, other autoantibodies, or "any autoantibody" in anti-CCP2-negative RA.

Additional file 7: Supplementary Table 6. Associations between different ACPA fine-specificities and HLA-DRB1 SE, in anti-CCP2-positive and anti-CCP2-negative RA. Odds ratios with 95% confidence intervals are shown for associations between HLA-DRB1 SE and presence/absence of 19 different ACPA fine-specificities, in anti-CCP2-positive and -negative RA: p-values indicate differences in ORs between ACPA fine-specificity-positive and -negative subsets in the anti-CCP2-positive subset.

Additional file 4: Supplementary Figure 1. ACPA fine-specificity levels in anti-CCP2-negative RA and controls. ACPA levels for 19 individual ACPA fine-specificities are shown in separate graphs with AU values on the Y-axes. Only ACPA levels above cutoff for positivity are shown.

Additional file 1.

Additional file 4: Supplementary Figure 1. ACPA fine-specificity levels in anti-CCP2-negative RA and controls. ACPA levels for 19 individual ACPA fine-specificities are shown in separate graphs with AU values on the Y-axes. Only ACPA levels above cutoff for positivity are shown.

Additional file 8: Supplementary Table 7. Associations between RF isotypes and smoking, in anti-CCP2-positive and anti-CCP2-negative RA. Odds ratios with 95% confidence intervals are shown for associations between smoking and presence/absence of IgM, IgG or IgA RF, in anti-CCP2-positive and -negative RA; p-values indicate differences in ORs between RF isotype-positive and -negative subsets.

Additional file 8: Supplementary Table 7. Associations between RF isotypes and smoking, in anti-CCP2-positive and anti-CCP2-negative RA. Odds ratios with 95% confidence intervals are shown for associations between smoking and presence/absence of IgM, IgG or IgA RF, in anti-CCP2-positive and -negative RA; p-values indicate differences in ORs between RF isotype-positive and -negative subsets.

Illustration of a conditional inference tree for developing RA in pre-symptomatic individuals compared with control subjects. CCP2 anti-CCP2 antibodies, Filaggrin anti-filaggrin antibodies, CEP-1 = anti-α-enolase antibodies, IgM-RF immunoglobulin M rheumatoid factor, HLA-SE human leukocyte antigen shared epitope, n number of individuals, Pr probability of detecting a pre-symptomatic individual, + = positive, − = negative. (DOCX 29 kb)

Prevalence, sensitivity, specificity and OR of numbers of RF isotypes, alone and/or in combination with ten different ACPA specificities, anti-CCP2 antibodies and anti-carbamylated protein antibodies in pre-symptomatic individuals and population controls. (DOCX 17kb)

Illustration of a conditional inference tree for developing RA in pre-symptomatic individuals compared with control subjects. CCP2 anti-CCP2 antibodies, Filaggrin anti-filaggrin antibodies, CEP-1 = anti-α-enolase antibodies, IgM-RF immunoglobulin M rheumatoid factor, HLA-SE human leukocyte antigen shared epitope, n number of individuals, Pr probability of detecting a pre-symptomatic individual, + = positive, − = negative. (DOCX 29 kb)

BackgroundRheumatoid arthritis (RA) can be divided into two major subsets based on the presence or absence of antibodies to citrullinated peptide antigens (ACPA). Until now, data from genome-wide association studies (GWAS) have only been published from ACPA-positive subsets of RA or from studies that have not separated the two subsets. The aim of the current study is to provide and compare GWAS data for both subsets.Methods and resultsGWAS using the Illumina 300K chip was performed for 774 ACPA-negative patients with RA, 1147 ACPA-positive patients with RA and 1079 controls from the Swedish population-based case-control study EIRA. Imputation was performed which allowed comparisons using 1,723,056 single nucleotide polymorphisms (SNPs). No SNP achieved genome-wide significance (2.9 × 10⁻⁸) in the comparison between ACPA-negative RA and controls. A case-case association study was then performed between ACPA-negative and ACPA-positive RA groups. The major difference in this analysis was in the HLA region where 768 HLA SNPs passed the threshold for genome-wide significance whereas additional contrasting SNPs did not reach genome-wide significance. However, one SNP close to the RPS12P4 locus in chromosome 2 reached a p value of 2 × 10⁶ and this locus can thus be considered as a tentative candidate locus for ACPA-negative RA.ConclusionsACPA-positive and ACPA-negative RA display significant risk allele frequency differences which are mainly confined to the HLA region. The data provide further support for distinct genetic aetiologies of RA subsets and emphasise the need to consider them separately in genetic as well as functional studies of this disease.

Figure S1. The correlation between RF-IgM and all proteins analyzed. (PDF 284 kb)

Additional file 1: Supplementary Table 1. Citrullinated peptide antigens on the multiplex microarray. The table lists citrullinated antigens used on the multiplex microarray, including name, protein of origin, and amino acid sequence, as well as references.

A more detailed description of the MS proteomic method is described. (PDF 289 kb)

A more detailed description of the affinity-based proteomic method is described. (PDF 370 kb)

Additional file 5: Supplementary Table 4. Other autoantibodies in anti-CCP2-positive and -negative RA and controls. Frequencies of 17 other autoantibodies in anti-CCP2-positive RA, anti-CCP2-negative RA and controls are shown, as well as median antibody levels in anti-CCP2-positive and -negative RA (p-values indicate differences between anti-CCP2-negative and anti-CCP2-positive RA subsets).

Table S2. Medications for the patients are reported. (PDF 526 kb)

Additional file 9: Supplementary Figure 2. Antibody levels in EIRA RA cases based on smoking status. IgM, IgG and IgA RF levels, as well as anti-CCP2 IgG levels, are shown for never, current and former smokers in all EIRA RA patients where smoking data was available.

Additional file 1: Supplementary Table 1. Citrullinated peptide antigens on the multiplex microarray. The table lists citrullinated antigens used on the multiplex microarray, including name, protein of origin, and amino acid sequence, as well as references.

Additional file 5: Supplementary Table 4. Other autoantibodies in anti-CCP2-positive and -negative RA and controls. Frequencies of 17 other autoantibodies in anti-CCP2-positive RA, anti-CCP2-negative RA and controls are shown, as well as median antibody levels in anti-CCP2-positive and -negative RA (p-values indicate differences between anti-CCP2-negative and anti-CCP2-positive RA subsets).

Additional file 2: Supplementary Table 2. Other (non-citrullinated) autoantigens on the multiplex microarray. The table lists the other antigens used on the multiplex microarray, including name, protein of origin, and major associated disease(s).

Figure S1. The correlation between RF-IgM and all proteins analyzed. (PDF 284 kb)

Additional file 9: Supplementary Figure 2. Antibody levels in EIRA RA cases based on smoking status. IgM, IgG and IgA RF levels, as well as anti-CCP2 IgG levels, are shown for never, current and former smokers in all EIRA RA patients where smoking data was available.

Additional file 1: Figure 1. Sequence, structure and major immunogenic region (mir) of hnRNP-D and hnRNP-DL. A, Schematic representation of hnRNP-D (isoform p45), hnRNP-DL and the different recombinant hnRNP-DL variants studied. The main structural features are highlighted. Mir-region is the major immunogenic region, RBD1 and RBD2 are RNA-binding domains 1 and 2, Gly-rich is the C-terminal glycine-rich region of the proteins. B, Global amino acid sequence alignment of hnRNP-D and hnRNP-DL1 (isoform 1). HnRNP-D and -DL share 89.1% similarity by sequenc e[1]. Regions “mir”, “RBD1”, “RBD2” and “Gly-rich” are highlighted. Figure 2. Characterisation of autoantibodies against, A, citrullinated α-hnRNP-DLmir (cit-DL), B, α-hnRNP-DLmir (DL) and C, ∆OD between cit-DL and DL (ΔDL) determed by ELISA in sera of other diseases (n=127; MS n=20, reA n=7, Sclero n=20, Sjö n=20, PsA n=20, MB n=20, OA n=20). The dotted lines markes the cutoff vs. other diseases (except systemic lupus erythematosus) or healthy controls with 98% specificity each. OD, optical density; nm, nano meter; vs., versus; MS, multiple sclerosis; reA, reactive arthritis; Sclero, scleroderma; Sjö, Sjögren´s syndrome; PsA, psoriasis arthritis; MB, ankylosing spondylitis; OA. Osteoarthritis. Table 1. Mann Whitney U-test of (cit) α-hnRNP-DLmir-OD signals of seropositive and seronegative data sets of RA-cohorts. Table 2. Mann Whitney U-test of cit α-hnRNP-DLmir-OD signals of seronegative data sets of RA-cohorts and data sets of other inflammatory diseases. Figure 3. XY-Plot and Spearman Correlation of citrullinated or native α-hnRNP-DLmir versus ΔhnRNP-DLmir for the early RA cohort EIRA (A/D; n=404), the seropositive EIRA sera (B/E; n=202) and the seronegative EIRA sera (C/F; n=202). Table 3. Spearman correlation of the early RA sera of the EIRA cohort (n=404). The results are given as R value (left of slash) with the corresponding p-value (right of slash). Table 4. Spearman correlation of the 242 EIRA sera treated with MTX (α-CCP2 positive n=133, α-CCP2 negative n=109). The results are given as R value (left of slash) with the corresponding p-value (right of slash). Table 5. Spearman correlation of the established RA sera of the Predict cohort (n=94; RF IgM and/or α-CCP2 positive n=64, RF IgM and α-CCP2 negative n=30). The results are given as R value (left of slash) with the corresponding p-value (right of slash). Table 6. ROC analysis of native hnRNP-DLmir of MTX-treated EIRA patients (n=192; seropositive n=93, seronegative n=99). Table 7. Negative CNDL-index of MTX-treated EIRA patients n=192 (Resp. n=161, non-Resp. n=31). Table 8. ROC analysis of native hnRNP-DLmir of Enbrel®-treated Predict patients (n=94; seropositive n=63, seronegative n=31). Figure 4. High baseline titer against α-hnRNP-DLmir (DL) is rather present in 6-month EULAR Responder RA patients who had received MTX or α-TNF inhibitor therapy (Enbrel®). A-C, Citrullinated α-hnRNP-DLmir (citDL) (A), α-hnRNP-DLmir (DL) (B) and ∆ OD between citDL and DL (ΔDL) (C) were measured by ELISA in patient sera from the EIRA cohort treated with MTX (n=192) with 161 EULAR Responder and 31 EULAR non-Responder among 6 months. The evaluation was done according to the cutoff versus other diseases. D, α-DL were measured by ELISA in patient sera from the Predict cohort treated with α-TNF inhibitor therapy with 6-month EULAR response data (n=94, responder n=63, non-Responder n=31). Based on the signals, a response-cutoff (dotted line, OD 0.174) was determined, from which only responders are recognized as positive. OD, optical density; nm, nano meter; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus; MTX, Methotrexate; Resp., 6-month EULAR Responder. Figure 5. A, Influence of cytokines on hnRNP-DL expression determined by immunoblotting. Cellular extracts from unstimulated, IL1α- or TNFα-stimulated HeLa cells and from unstimulated and IL6-stimulated HepG2 cells were probed with α-hnRNP-DL1/2-peptide specific rabbit serum. B, Citrullination of hnRNP-DL in synovial tissue from a patient with rheumatoid arthritis was investigated with an α-deiminated arginine antibody and an α-hnRNP-DL antibody. Both positive bands were labled with hnRNP-DL, which isoforms were not analysed.

Table S1. The number of missing data points for each variable is shown. (PDF 668 kb)

Additional file 1: Figure 1. Sequence, structure and major immunogenic region (mir) of hnRNP-D and hnRNP-DL. A, Schematic representation of hnRNP-D (isoform p45), hnRNP-DL and the different recombinant hnRNP-DL variants studied. The main structural features are highlighted. Mir-region is the major immunogenic region, RBD1 and RBD2 are RNA-binding domains 1 and 2, Gly-rich is the C-terminal glycine-rich region of the proteins. B, Global amino acid sequence alignment of hnRNP-D and hnRNP-DL1 (isoform 1). HnRNP-D and -DL share 89.1% similarity by sequenc e[1]. Regions “mir”, “RBD1”, “RBD2” and “Gly-rich” are highlighted. Figure 2. Characterisation of autoantibodies against, A, citrullinated α-hnRNP-DLmir (cit-DL), B, α-hnRNP-DLmir (DL) and C, ∆OD between cit-DL and DL (ΔDL) determed by ELISA in sera of other diseases (n=127; MS n=20, reA n=7, Sclero n=20, Sjö n=20, PsA n=20, MB n=20, OA n=20). The dotted lines markes the cutoff vs. other diseases (except systemic lupus erythematosus) or healthy controls with 98% specificity each. OD, optical density; nm, nano meter; vs., versus; MS, multiple sclerosis; reA, reactive arthritis; Sclero, scleroderma; Sjö, Sjögren´s syndrome; PsA, psoriasis arthritis; MB, ankylosing spondylitis; OA. Osteoarthritis. Table 1. Mann Whitney U-test of (cit) α-hnRNP-DLmir-OD signals of seropositive and seronegative data sets of RA-cohorts. Table 2. Mann Whitney U-test of cit α-hnRNP-DLmir-OD signals of seronegative data sets of RA-cohorts and data sets of other inflammatory diseases. Figure 3. XY-Plot and Spearman Correlation of citrullinated or native α-hnRNP-DLmir versus ΔhnRNP-DLmir for the early RA cohort EIRA (A/D; n=404), the seropositive EIRA sera (B/E; n=202) and the seronegative EIRA sera (C/F; n=202). Table 3. Spearman correlation of the early RA sera of the EIRA cohort (n=404). The results are given as R value (left of slash) with the corresponding p-value (right of slash). Table 4. Spearman correlation of the 242 EIRA sera treated with MTX (α-CCP2 positive n=133, α-CCP2 negative n=109). The results are given as R value (left of slash) with the corresponding p-value (right of slash). Table 5. Spearman correlation of the established RA sera of the Predict cohort (n=94; RF IgM and/or α-CCP2 positive n=64, RF IgM and α-CCP2 negative n=30). The results are given as R value (left of slash) with the corresponding p-value (right of slash). Table 6. ROC analysis of native hnRNP-DLmir of MTX-treated EIRA patients (n=192; seropositive n=93, seronegative n=99). Table 7. Negative CNDL-index of MTX-treated EIRA patients n=192 (Resp. n=161, non-Resp. n=31). Table 8. ROC analysis of native hnRNP-DLmir of Enbrel®-treated Predict patients (n=94; seropositive n=63, seronegative n=31). Figure 4. High baseline titer against α-hnRNP-DLmir (DL) is rather present in 6-month EULAR Responder RA patients who had received MTX or α-TNF inhibitor therapy (Enbrel®). A-C, Citrullinated α-hnRNP-DLmir (citDL) (A), α-hnRNP-DLmir (DL) (B) and ∆ OD between citDL and DL (ΔDL) (C) were measured by ELISA in patient sera from the EIRA cohort treated with MTX (n=192) with 161 EULAR Responder and 31 EULAR non-Responder among 6 months. The evaluation was done according to the cutoff versus other diseases. D, α-DL were measured by ELISA in patient sera from the Predict cohort treated with α-TNF inhibitor therapy with 6-month EULAR response data (n=94, responder n=63, non-Responder n=31). Based on the signals, a response-cutoff (dotted line, OD 0.174) was determined, from which only responders are recognized as positive. OD, optical density; nm, nano meter; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus; MTX, Methotrexate; Resp., 6-month EULAR Responder. Figure 5. A, Influence of cytokines on hnRNP-DL expression determined by immunoblotting. Cellular extracts from unstimulated, IL1α- or TNFα-stimulated HeLa cells and from unstimulated and IL6-stimulated HepG2 cells were probed with α-hnRNP-DL1/2-peptide specific rabbit serum. B, Citrullination of hnRNP-DL in synovial tissue from a patient with rheumatoid arthritis was investigated with an α-deiminated arginine antibody and an α-hnRNP-DL antibody. Both positive bands were labled with hnRNP-DL, which isoforms were not analysed.

Table S1. The number of missing data points for each variable is shown. (PDF 668 kb)

A more detailed description of the affinity-based proteomic method is described. (PDF 370 kb)

Table S3. The correlation between RF-IgM and the 25 proteins in Table 2 are shown. (PDF 256 kb)

A more detailed description of the MS proteomic method is described. (PDF 289 kb)

Table S3. The correlation between RF-IgM and the 25 proteins in Table 2 are shown. (PDF 256 kb)

Table S2. Medications for the patients are reported. (PDF 526 kb)

usc Autoimmune polyendocrine syndrome type 1 (APS-1) is a multiorgan autoimmune disease caused by mutations in the autoimmune regulator (AIRE) gene. Chronic mucocutaneous candidiasis, hypoparathyroidism and adrenal failure are hallmarks of the disease. The critical mechanisms causing chronic mucocutaneous candidiasis in APS-1 patients have not been identified although autoantibodies to cytokines are implicated in the pathogenesis. To investigate whether the Th reactivity to Candida albicans (C. albicans) and other stimuli was altered, we isolated PBMC from APS-1 patients and matched healthy controls. The Th17 pathway was upregulated in response to C. albicans in APS-1 patients, whereas the IL-22 secretion was reduced. Autoantibodies against IL-22, IL-17A and IL-17F were detected in sera from APS-1 patients by immunoprecipitation. In addition, Aire-deficient (Aire 0/0 ) mice were much more susceptible than Aire +/+ mice to mucosal candidiasis and C. albicans-induced Th17- and Th1-cell responses were increased in Aire 0/0 mice. Thus an excessive IL-17A reactivity towards C. albicans was observed in APS-1 patients and Aire 0/0 mice. Refereed/Peer-reviewed