Anti-integrin αvß6 antibody in Takayasu arteritis patients with or without ulcerative colitis.

Background: It has been well documented that Takayasu arteritis (TAK) and ulcerative colitis (UC) coexist in the same patients. HLA-B*52 characterizes the co-occurrence, which is one of the common genetic features between these two diseases, indicating shared underlying pathologic mechanisms. Anti-integrin αvß6 antibody (Ab) is present in sera of UC patients in a highly specific manner. We investigated if there were any associations between anti-integrin αvß6 Ab and TAK, considering the risk HLA alleles. Methods: A total of 227 Japanese TAK patients were recruited in the current study and their serum samples were subjected to measurement of anti-integrin αvß6 Ab by ELISA. The clinical information, including the co-occurrence of UC, was collected. The HLA allele carrier status was determined by Luminex or genotype imputation. Results: The information about the presence of UC was available for 165 patients, among which eight (4.84%) patients had UC. Anti-integrin αvß6 antibody was identified in 7 out of 8 TAK subjects with UC (87.5%) while only 5 out of 157 (3.18%) TAK subjects without UC had the antibody (OR 121, p=7.46×10-8). A total of 99 out of 218 (45.4%) patients were HLA-B*52 carriers. There was no significant association between the presence of anti-integrin αvß6 Ab and HLA-B*52 carrier status in those without UC (OR 2.01, 95% CI 0.33-12.4, p = 0.189). Conclusions: The prevalence of anti-integrin αvß6 Ab was high in TAK patients with UC, but not in the absence of concomitant UC. The effect of HLA-B*52 on anti-integrin αvß6 Ab production would be minimal.

Attenuation of HOIL-1L ligase activity promotes systemic autoimmune disorders by augmenting linear ubiquitin signaling.

Linear ubiquitin chains, which are generated specifically by the linear ubiquitin assembly complex (LUBAC) ubiquitin ligase, play crucial roles in immune signaling, including NF-κB activation. LUBAC comprises catalytic large isoform of heme-oxidized iron regulatory protein 2 ubiquitin ligase 1 (HOIL-1L) interacting protein (HOIP), accessory HOIL-1L, and SHANK-associated RH domain-interacting protein (SHARPIN). Deletion of the ubiquitin ligase activity of HOIL-1L, an accessory ligase of LUBAC, augments LUBAC functions by enhancing LUBAC-mediated linear ubiquitination, which is catalyzed by HOIP. Here, we show that HOIL-1L ΔRING1 mice, which exhibit augmented LUBAC functions upon loss of the HOIL-1L ligase, developed systemic lupus erythematosus (SLE) and Sjögren's syndrome in a female-dominant fashion. Augmented LUBAC activity led to hyperactivation of both lymphoid and myeloid cells. In line with the findings in mice, we sought to identify missense single nucleotide polymorphisms/variations of the RBCK1/HOIL-1L gene in humans that attenuate HOIL-1L ligase activity. We found that the R464H variant, which is encoded by rs774507518 within the RBCK1/HOIL-1L gene, attenuated HOIL-1L ligase activity and augmented LUBAC-mediated immune signaling, including that mediated by Toll-like receptors. We also found that rs774507518 was enriched significantly in patients with SLE, strongly suggesting that RBCK1/HOIL-1L is an SLE susceptibility gene and that augmented linear ubiquitin signaling generated specifically by LUBAC underlies the pathogenesis of this prototype systemic autoimmune disease.

Clinical Characteristics of Cryopyrin-Associated Periodic Syndrome and Long-Term Real-World Efficacy and Tolerability of Canakinumab in Japan: Results of a Nationwide Survey.

OBJECTIVE: We assess the clinical characteristics of patients with cryopyrin-associated periodic syndrome (CAPS) in Japan and evaluate the real-world efficacy and safety of interleukin-1 (IL-1) inhibitors, primarily canakinumab. METHODS: Clinical information was collected retrospectively, and serum concentrations of canakinumab and cytokines were analyzed. RESULTS: A total of 101 patients were included, with 86 and 15 carrying heterozygous germline and somatic mosaic mutations, respectively. We identified 39 mutation types, and the common CAPS-associated symptoms corresponded with those in previous reports. Six patients (5.9% of all patients) died, with four of the deaths caused by CAPS-associated symptoms. Notably, 73.7% of patients (100%, 79.6%, and 44.4% of familial cold autoinflammatory syndrome, Muckle-Wells syndrome, and chronic infantile neurological cutaneous articular syndrome/neonatal onset multisystem inflammatory disease, respectively) achieved complete remission with canakinumab, and early therapeutic intervention was associated with better auditory outcomes. In some patients, canakinumab treatment stabilized the progression of epiphysial overgrowth and improved height gain, visual acuity, and renal function. However, 23.7% of patients did not achieve inflammatory remission with crucial deterioration of organ damage, with two dying while receiving high-dose canakinumab treatment. Serological analysis of canakinumab and cytokine concentrations revealed that the poor response was not related to canakinumab shortage. Four inflammatory nonremitters developed inflammatory bowel disease (IBD)-unclassified during canakinumab treatment. Dual biologic therapy with canakinumab and anti-tumor necrosis factor-α agents was effective for IBD- and CAPS-associated symptoms not resolved by canakinumab monotherapy. CONCLUSION: This study provides one of the largest epidemiologic data sets for CAPS. Although early initiation of anti-IL-1 treatment with canakinumab is beneficial for improving disease prognosis, some patients do not achieve remission despite a high serum concentration of canakinumab. Moreover, IBD may develop in CAPS after canakinumab treatment.

Monocyte-derived transcriptomes explain the ineffectiveness of abatacept in rheumatoid arthritis.

BACKGROUND: The biological mechanisms underlying the differential response to abatacept in patients with rheumatoid arthritis (RA) are unknown. Here, we aimed to identify cellular, transcriptomic, and proteomic features that predict resistance to abatacept in patients with RA. METHODS: Blood samples were collected from 22 RA patients treated with abatacept at baseline and after 3 months of treatment. Response to treatment was defined by the European League Against Rheumatism (EULAR) response criteria at 3 months, and seven patients were classified as responders and the others as non-responders. We quantified gene expression levels by RNA sequencing, 67 plasma protein levels, and the expression of surface molecules (CD3, 19, and 56) by flow cytometry. In addition, three gene expression data sets, comprising a total of 27 responders and 50 non-responders, were used to replicate the results. RESULTS: Among the clinical characteristics, the number of monocytes was significantly higher in the non-responders before treatment. Cell type enrichment analysis showed that differentially expressed genes (DEGs) between responders and non-responders were enriched in monocytes. Gene set enrichment analysis, together with single-cell analysis and deconvolution analysis, identified that Toll-like receptor 5 (TLR5) and interleukin-17 receptor A (IL17RA) pathway in monocytes was upregulated in non-responders. Hepatocyte growth factor (HGF) correlated with this signature showed higher concentrations in non-responders before treatment. The DEGs in the replication set were also enriched for the genes expressed in monocytes, not for the TLR5 and IL17RA pathway but for the oxidative phosphorylation (OXPHOS) pathway. CONCLUSIONS: Monocyte-derived transcriptomic features before treatment underlie the differences in abatacept efficacy in patients with RA. The pathway activated in monocytes was the TLR5 and IL17RA-HGF signature in the current study, while it was the OXPHOS pathway in the replication set. Elevated levels of HGF before treatment may serve as a potential biomarker for predicting poor responses to abatacept. These findings provide insights into the biological mechanisms of abatacept resistance, contributing valuable evidence for stratifying patients with RA.

A 52-Year-Old With Painful Fingertips.

A patient had cold, cyanotic fingertips with small ulcerations. Laboratory testing showed leukocytosis, eosinophilia, and elevated D-dimer level; results of tests for antinuclear antibodies, antiphospholipid antibodies, antineutrophil cytoplasmic antibodies against proteinase 3 and myeloperoxidase, and HIV were negative. What is the diagnosis and what would you do next?

Positive and negative regulation of the Fcγ receptor-stimulating activity of RNA-containing immune complexes by RNase.

The U1RNP complex, Ro/SSA, and La/SSB are major RNA-containing autoantigens. Immune complexes (ICs) composed of RNA-containing autoantigens and autoantibodies are suspected to be involved in the pathogenesis of some systemic autoimmune diseases. Therefore, RNase treatment, which degrades RNA in ICs, has been tested in clinical trials as a potential therapeutic agent. However, no studies to our knowledge have specifically evaluated the effect of RNase treatment on the Fcγ receptor-stimulating (FcγR-stimulating) activity of RNA-containing ICs. In this study, using a reporter system that specifically detects FcγR-stimulating capacity, we investigated the effect of RNase treatment on the FcγR-stimulating activity of RNA-containing ICs composed of autoantigens and autoantibodies from patients with systemic autoimmune diseases such as systemic lupus erythematosus. We found that RNase enhanced the FcγR-stimulating activity of Ro/SSA- and La/SSB-containing ICs, but attenuated that of the U1RNP complex-containing ICs. RNase decreased autoantibody binding to the U1RNP complex, but increased autoantibody binding to Ro/SSA and La/SSB. Our results suggest that RNase enhances FcγR activation by promoting the formation of ICs containing Ro/SSA or La/SSB. Our study provides insights into the pathophysiology of autoimmune diseases involving anti-Ro/SSA and anti-La/SSB autoantibodies, and into the therapeutic application of RNase treatment for systemic autoimmune diseases.

Synovial Tissue Heterogeneity in Japanese Patients With Rheumatoid Arthritis Elucidated Using a Cell-Type Deconvolution Approach.

OBJECTIVE: Recent advances in single-cell RNA sequencing technology have improved our understanding of the immunological landscape of rheumatoid arthritis (RA). We aimed to stratify the synovium from East Asian patients with RA by immune cell compositions and gain insight into the inflammatory drivers of each synovial phenotype. METHODS: Synovial tissues were obtained from East Asian patients in Japan with RA (n = 41) undergoing articular surgery. The cellular composition was quantified by a deconvolution approach using a public single-cell-based reference. Inflammatory pathway activity was calculated by gene set variation analysis, and chromatin accessibility was evaluated using assay of transposase accessible chromatin-sequencing. RESULTS: We stratified RA synovium into three distinct subtypes based on the hierarchical clustering of cellular composition data. One subtype was characterized by abundant HLA-DRAhigh synovial fibroblasts, autoimmune-associated B cells, GZMK+ GZMB+ CD8+ T cells, interleukin (IL)1-ß+ monocytes, and plasmablasts. In addition, tumor necrosis factor (TNF)-α, interferons (IFNs), and IL-6 signaling were highly activated in this subtype, and the expression of various chemokines was significantly enhanced. Moreover, we found an open chromatin region overlapping with RA risk locus rs9405192 near the IRF4 gene, suggesting the genetic background influences the development of this inflammatory synovial state. The other two subtypes were characterized by increased IFNs and IL-6 signaling, and expression of molecules associated with degeneration, respectively. CONCLUSION: This study adds insights into the synovial heterogeneity in East Asian patients and shows a promising link with predominant inflammatory signals. Evaluating the site of inflammation has the potential to lead to appropriate drug selection that matches the individual pathology.

Expression of S100A8 protein on B cells is associated with disease activity in patients with systemic lupus erythematosus.

BACKGROUND: Systemic lupus erythematosus (SLE) is an intractable disease characterized by autoantibody production and autoreactive B and T cell proliferation. Although several studies have revealed multiple genetic and environmental associations, the underlying mechanisms remain unknown. METHODS: We performed proteomics and transcriptomics using liquid chromatography-mass spectrometry and DNA microarray, using peripheral blood B cells from patients with SLE, and healthy controls (HC). We explored molecules associated with the pathophysiology of SLE by flow cytometry and B cell stimulation assay. RESULTS: We identified for the first time that expression of both S100A8 protein and mRNA were markedly upregulated in SLE B cells. The results obtained using flow cytometry showed that S100A8 was highly expressed on the surface of B cells of patients with active SLE (MFI; HC 102.5 ± 5.97, stable SLE 111.4 ± 12.87, active SLE 586.9 ± 142.9), and S100A8 on the cell surface was decreased after treatment (MFI; pre-treat 1094.5 ± 355.38, post-treat 492.25 ± 247.39); therefore, it is suggested that S100A8 may be a marker for disease activity. The mRNA expression of S100A8 was particularly upregulated in memory B cells of SLE (56.68 fold higher than HC), suggesting that S100A8 may be mainly secreted by memory B cells in the pathogenesis of SLE. CONCLUSIONS: Our results imply that the S100A8 proteins secreted from memory B cells may stimulate granulocytes and monocytes through pattern recognition receptors, activate the innate immune system, and are involved in the pathogenesis of SLE.

Anti-synthetase Syndrome That Relapsed with Pulmonary Arterial Hypertension and Malignancy.

A 69-year-old man with a history of anti-synthetase antibody-positive polymyositis and interstitial lung disease (ILD) stable for more than 20 years suddenly developed pulmonary artery hypertension (PAH) with a mean PA pressure of 46 mmHg. At this stage, ILD was mild, but it became acutely exacerbated later, and high-dose corticosteroid and intravenous cyclophosphamide ameliorated both PAH and ILD. The tricuspid regurgitation pressure gradient decreased from 80 to 49 mmHg and ILD recovered almost completely. During a systemic examination, bone metastatic cancer of unknown origin was found. We herein report the relationship between anti-synthetase syndrome (ASS) and PAH as well as ASS and malignancy.

Rheumatoid factor recognizes specific domains of the IgG heavy chain complexed with HLA class II molecules.

OBJECTIVE: We previously reported that RF recognized the IgG heavy chain (IgGH)/RA-susceptible HLA class II molecule complex. In the present study, we investigated the molecular mechanisms underlying HLA binding to and the RF recognition of IgGH. METHODS: We synthesized various types of IgGH segments, including VH, CH1, CH2 and CH3, and transfected them with or without HLA class II molecules into the Human Embryonic Kidney 293T cell line. IgGH single domains linked with the HLA-Cw3 peptide, which binds to the binding groove of the HLA class II molecule, were also synthesized. The expression of IgGH domains on the cell surface and their recognition by RF were examined using flow cytometry. RESULTS: Flag-tagged IgGH segments containing CH1 (CH1, VH-CH1, CH1-CH2, VH-CH1-CH2, CH1-CH2-CH3 and VH-CH1-CH2-CH3) were clearly presented on the cell surface by HLA-DR4, while segments without the CH1 domain were expressed at a low level, and the CH3 single domain was only weakly detected on the cell surface, even with HLA-DR4. We then transfected IgGH single domains linked to the Cw3 peptide together with HLA-DR4 and showed that RF-containing sera from RA patients only recognized the CH3 domain and none of the other single domains. When various segments without the Cw3 peptide were transfected with HLA-DR4, only the CH1-CH2-CH3 segment and full-length IgGH were detected by the sera of RA patients. CONCLUSION: The CH1 domain of IgGH binds to the RA-susceptible HLA-DR molecule and is expressed on the cell surface. RF specifically recognizes the CH3 domain of the IgGH/HLA-DR4 complex.

Factors associated with osteoporosis and fractures in patients with systemic lupus erythematosus: Kyoto Lupus Cohort.

OBJECTIVES: Osteoporosis and compression fractures of the lumbar spine are some of the major adverse effects of glucocorticoid therapy in patients with systemic lupus erythematosus (SLE). This study examined the association between bone mineral density, bone turnover markers, presence of vertebral fractures, and Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index in SLE patients. METHODS: This was a cross-sectional study of 246 outpatients with SLE at the Kyoto University Hospital. Lumbar and femoral bone mineral density was measured with dual-energy X-ray absorptiometry, and the presence of vertebral fractures was determined using X-ray, computed tomography, or magnetic resonance imaging. RESULTS: On multiple regression analysis, both high lumbar and femoral T-scores were associated with the concomitant use of hydroxychloroquine (P = .018 and P = .037, respectively), no use of bisphosphonate or denosumab (P = .004 and P = .038, respectively), high body mass index (P < .001), and low bone-specific alkaline phosphatase level (P = .014 and P = .002, respectively). Vertebral fractures showed a significant association with Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index score (P < .001) and femoral T-score (P < .001). CONCLUSION: Vertebral fracture was associated with SLE-associated organ damage, and serum bone-specific alkaline phosphatase level is a potentially useful marker for osteoporosis monitoring in SLE patients.

Genetic influences on human blood metabolites in the Japanese population.

An increase in ethnic diversity in genetic studies has the potential to provide unprecedented insights into how genetic variations influence human phenotypes. In this study, we conducted a quantitative trait locus (QTL) analysis of 121 metabolites measured using gas chromatography-mass spectrometry with plasma samples from 4,888 Japanese individuals. We found 60 metabolite-gene associations, of which 13 have not been previously reported. Meta-analyses with another Japanese and a European study identified six and two additional unreported loci, respectively. Genetic variants influencing metabolite levels were more enriched in protein-coding regions than in the regulatory regions while being associated with the risk of various diseases. Finally, we identified a signature of strong negative selection for uric acid ( S ˆ  = -1.53, p = 6.2 × 10-18). Our study expanded the knowledge of genetic influences on human blood metabolites, providing valuable insights into their physiological, pathological, and selective properties.

Genetic architecture underlying IgG-RF production is distinct from that of IgM-RF.

OBJECTIVE: HLA-DRB1 alleles, particularly the shared epitope (SE) alleles, are strongly associated with RA. Different genetic structures underlie the production of the various autoantibodies in RA. While extensive genetic analyses have been conducted to generate a detailed profile of ACPA, a representative autoantibody in RA, the genetic architecture underlying subfractions of RF other than IgM-RF, namely IgG-RF, known to be associated with rheumatoid vasculitis, is not well understood. METHODS: We enrolled a total of 743 RA subjects whose detailed autoantibody (IgG-RF, IgM-RF, and ACPA) data were available. We evaluated co-presence and correlations of the levels of these autoantibodies. We analysed associations between the presence or levels of the autoantibodies and HLA-DRB1 alleles for the 743 RA patients and 2008 healthy controls. RESULTS: We found both IgG-RF(+) and IgG-RF(-) RA subjects showed comparable associations with SE alleles, which was not observed for the other autoantibodies. Furthermore, there was a clear difference in SE allele associations between IgG-RF(+) and (-) subsets: the association with the IgG-RF(+) subsets was solely driven by HLA-DRB1*04:05, the most frequent SE allele in the Japanese population, while not only HLA-DRB1*04:05 but also HLA-DRB1*04:01, less frequent in the Japanese population but the most frequent SE allele in Europeans, were main drivers of the association in the IgG-RF(-) subset. We confirmed that these associations were irrespective of ACPA presence. CONCLUSION: We found a unique genetic architecture for IgG-RF(-) RA, which showed a strong association with a SE allele not frequent in the Japanese population but the most frequent SE allele in Europeans. The findings could shed light on uncovered RA pathology.

Four cases of dermatomyositis with abnormally high anti-MDA-5 antibody titres and not high levels of serum ferritin.

Anti-melanoma differentiation-associated gene 5 (MDA-5) antibody-positive dermatomyositis is a fatal disease presenting with rapidly progressive interstitial lung disease. High ferritin levels are a well-known poor prognostic factor. A high anti-MDA-5 antibody titre was also recently identified as a poor prognostic factor. We encountered four cases that had extremely high anti-MDA-5 antibody titres without high levels of ferritin in the initial examination. All cases were female with ages ranging between 29 and 54 years (mean age, 44 years). In the initial examination, anti-MDA-5 antibody titres were 2060-3040 (normal range, <32 index), ferritin levels were 87-480 ng/ml (normal range, 2.6-129.4 ng/ml), KL-6 level was 186-1806 U/ml (normal range, <500 U/ml), and creatine kinase level was normal in all patients. One patient had respiratory distress on exertion. Computed Tomography (CT) images showed mild ground-glass attenuation/reticular shadows near the pleura in all patients. Three patients were treated with a combination of high-dose glucocorticoids, intermittent intravenous cyclophosphamide, and calcineurin inhibitors, and two required plasma exchange due to the worsening of lung lesion. In these patients, ferritin and KL-6 levels tended to elevate after the beginning of treatment. Very mild pulmonary lesions disappeared in one patient treated with moderate doses of a glucocorticoid and calcineurin inhibitor. All patients survived, and one required oxygen on exertion at discharge. The condition of patients with abnormally high anti-MDA-5 antibody titres may deteriorate even though ferritin levels were not high and lung shadows are minimal at presentation. Therefore, intensive treatment needs to be considered early in the course of the disease regardless of the serum ferritin level.

A case of aortoduodenal fistula caused by IgG4-related periaortitis.

An 86-year-old man who underwent endovascular aortic repair for impending rupture of an abdominal aortic aneurysm a year ago presented to our hospital because of fatigue and black stools. Multiple bacterial specimens were detected in blood cultures, and computed tomography following oral administration of gastrografin demonstrated gastrografin in the abdominal aorta. The diagnosis of aortic duodenal fistula was confirmed and emergency abdominal aortic replacement was performed. The pathological findings of the aorta included a large number of immunoglobulin G4 (IgG4)-positive plasma cells infiltrating all layers of the aortic wall, with particularly marked thickening of the adventitia. The serum IgG4 level was 241 mg/dl and IgG4-related periaortitis was diagnosed. Aortoduodenal fistula is a rare but fatal complication of IgG4-related periaortitis. Patients should be followed carefully after endovascular aortic repair for inflammatory abdominal aortic aneurysms.

Rituximab in the real-world treatment of lupus nephritis: A retrospective cohort study in Japan.

OBJECTIVES: Although recommended in established international guidelines for lupus nephritis (LN), rituximab is not officially approved for LN treatment, making all such use off-label. The Japan College of Rheumatology (JCR) conducted a retrospective observational study on real-world efficacy and safety of rituximab treatment for LN in Japan. METHODS: Clinical data were collected from 47 hospitals for LN patients treated with rituximab to retrospectively investigate dosing schedule, efficacy, and safety. RESULTS: This retrospective analysis included 115 patients: 84 (73%) received 375 mg/m2 weekly up to four doses, and 31 (27%) received 1000 mg/body in one or two doses 2 weeks apart. Rituximab significantly improved findings for urinalysis, systemic lupus erythematosus serology, and systemic lupus erythematosus disease activity and was assessed as 'extremely effective' in 24.8% of patients and 'effective' in 60.2%. The renal response by the JCR-I criteria was 52.5% for overall response rate (ORR) (complete renal response rate 20.8% and partial renal response rate 31.7%) and that by the JCR-II criteria was 49.5% (21.8% and 27.7%, respectively). Corticosteroid dose was significantly reduced. Rituximab was well tolerated, with frequent but manageable adverse events of infusion reaction and infection. CONCLUSIONS: Rituximab is effective for the treatment of Japanese patients with LN refractory to conventional therapy.

SARS-CoV-2 ORF8 is a viral cytokine regulating immune responses.

Many patients with severe COVID-19 suffer from pneumonia and the elucidation of the mechanisms underlying the development of this severe condition is important. The in vivo function of the ORF8 protein secreted by SARS-CoV-2 is not well understood. Here, we analyzed the function of ORF8 protein by generating ORF8-knockout SARS-CoV-2 and found that the lung inflammation observed in wild-type SARS-CoV-2-infected hamsters was decreased in ORF8-knockout SARS-CoV-2-infected hamsters. Administration of recombinant ORF8 protein to hamsters also induced lymphocyte infiltration into the lungs. Similar pro-inflammatory cytokine production was observed in primary human monocytes treated with recombinant ORF8 protein. Furthermore, we demonstrated that the serum ORF8 protein levels are well-correlated with clinical markers of inflammation. These results demonstrated that the ORF8 protein is a SARS-CoV-2 viral cytokine involved in the immune dysregulation observed in COVID-19 patients, and that the ORF8 protein could be a novel therapeutic target in severe COVID-19 patients.