Yukiko Takakuwa

Clinical subsets associated with different anti-aminoacyl transfer RNA synthetase antibodies and their association with coexisting anti-Ro52

Abstract Objective: To characterize clinical features of polymyositis/dermatomyositis (PM/DM) patients with different anti-aminoacyl transfer RNA synthetase (ARS) antibodies and their association with anti-Ro52. Methods: Autoantibodies in sera from 97 Japanese patients (36 PM, 56 DM, and 5 clinically amyopathic DM), who satisfied Bohan and Peter or modified Sontheimers criteria, were characterized by immunoprecipitation and enzyme-linked immunosorbent assay. Clinical information was from medical records. Features associated with different anti-ARS and anti-Ro52 antibodies were analyzed. Results: The prevalence of anti-ARS was similar to other studies (Jo-1, 22%; EJ, 4%; OJ, 1%; PL-12, 1%), except for a high prevalence of anti-PL-7 (12%), which allowed us to characterize patients carrying this specificity. Serum creatine kinase >3000 IU/l was less common in anti-PL-7-positive patients (57%) than anti-Jo-1-positive patients (18%) (p = 0.0328) and was not found in anti-EJ-positive individuals. Interstitial lung disease was common in anti-ARS-positive patients (97%) (p < 0.0001 vs. 48% in anti-ARS-negative). Anti-Ro52 antibodies were frequently detected with anti-ARS (59%) (57% in anti-Jo-1, 67% in anti-PL-7) (vs. 21% in anti-ARS-negative, p < 0.0002). Anti-Ro52 was associated with overlap syndrome (26%) (vs. 7% in anti-Ro52-negative, p = 0.0119). Conclusions: Patients with different anti-ARS in combination with anti-Ro52 appear to be associated with distinctive clinical subsets.

Altered posttranslational modification on U1 small nuclear ribonucleoprotein 68k in systemic autoimmune diseases detected by 2D Western blot

Anti‐ribonucleoprotein (anti‐RNP) antibodies are one of the representative autoantibodies detectable in patients with systemic lupus erythematosus (SLE) and mixed connective tissue disease (MCTD). Generally, posttranslational modifications (PTMs) on autoantigens are proposed to be involved in the production of autoantibodies. In this study, we tried to detect the alteration in PTMs on a U1 small nuclear RNP 68k subunit (U1‐68k), a major antigen of anti‐RNP antibodies. Peripheral blood mononuclear cells (PBMCs) were obtained from patients with MCTD, SLE, and rheumatoid arthritis (RA), and from healthy donors. U1‐68ks in the PBMCs were detected by 2D Western blot (WB), where extracted nuclear proteins were separated by 2DE, followed by the detection of U1‐68k using WB. More than 20 PTM isoforms were detected with different molecular weights of 65.0 , 66.5, and 68.0kDa, and different pIs between 6.0 and 8.5. Importantly, the relative intensity of the spot with 66.5 kDa and pI 7.5 was significantly increased in the MCTD and SLE groups compared to the RA and healthy groups. Further, this U1‐68k isoform, in particular, in its RS domain, was found to have significantly decreased phosphorylation compared to the other isoforms. The PTM alternation may be one of the steps to generate the anti‐RNP antibodies.

Peptidomics: Identification of Pathogenic and Marker Peptides

Recent years have seen great advances in mass spectrometry and proteomics, the science dealing with the analysis of proteins, their structure and function. A branch of proteomics dealing with naturally occurring peptides is often referred to as peptidomics. Direct analysis of peptides produced by processing or degradation of proteins might be useful for example for detecting and identifying pathogenic and/or biomarker peptides in body fluids like blood. In this paper, we introduce one of the standard protocols for comprehensive analysis of serum-derived peptides, which consists of methods for purification of serum peptides, detection of peptides, pattern recognition and clustering (bioinformatics), and identification of peptide sequences. Peptide identification should be followed by the investigation of their pathogenic roles using for example synthetic peptides and the establishment of their usefulness as bioclinical markers.

Comparative proteomic analysis of neutrophils from patients with microscopic polyangiitis and granulomatosis with polyangiitis

UNLABELLED Both microscopic polyangiitis (MPA) and granulomatosis with polyangiitis (GPA) belong to ANCA-associated vasculitis (AAV), in which neutrophils play a key role in their pathology. In this study, in order to discriminate between MPA and GPA, protein profiles of peripheral blood polymorphonuclear cells (PMNs) of 11 MPA patients and 9 GPA patients and 10 healthy controls (HC) were analyzed by 2D-DIGE. In all the 864 spots detected, intensity of 55 spots was significantly different (p<0.05) among the three groups by ANOVA. 31 out of the 55 spots were identified by mass spectrometry. Orthogonal partial-least-squares-discriminate analysis revealed that the abundance profile of the protein spots discriminated the AAV group from the HC group, and the MPA group from the GPA group completely. 13 protein spots were considered as biomarker candidates to distinguish between MPA and GPA. In those, spots whose intensity was higher in MPA than in GPA included actin with various pI values, while a considerable part of spots whose intensity was higher in GPA were proteins related with the activity of neutrophils. Among the candidate proteins, ROC analysis showed that a combination of neutrophil gelatinase-associated lipocalin and a-kinase anchor protein 7 isoforms beta had a high diagnostic potential. BIOLOGICAL SIGNIFICANCE In this study, protein profiles of polymorphonuclear cells (PMNs) of microscopic polyangiitis (MPA) and granulomatosis with polyangiitis (GPA) patients and healthy controls (HC) were investigated by 2D-DIGE, and MS analysis. As a result, we found that the protein profiles of PMNs were useful for distinguishing between patients (MPA and GPA) and HC, and between patients with MPA and patients with GPA. Especially, we found that the 13 protein spots that consisted of 10 proteins considerably contributed to the discrimination between MPA and GPA. This is the first to demonstrate that protein profiles of PMNs are different among MPA, GPA and healthy control. The 10 proteins we identified in this study would be new biomarkers for the diagnosis of the diseases, and may be reflect the pathology difference between MPA and GPA.

Comparison of renal response to four different induction therapies in Japanese patients with lupus nephritis class III or IV: A single-centre retrospective study

The recent recommendations for the management of lupus nephritis suggest that racial background should be considered while choosing induction therapy. However, the responses to different induction regimens have been poorly studied in Japanese population. Here, we assessed the renal response to different induction therapies in Japanese patients with lupus nephritis class III or IV. The records of 64 patients with biopsy-proven lupus nephritis class III or IV were retrospectively evaluated according to therapy received: monthly intravenous cyclophosphamide (IVCY), the Euro-lupus nephritis trial (ELNT) protocol-IVCY, tacrolimus (TAC), or mycophenolate mofetil (MMF). We investigated cumulative complete renal response (CR) rates and relapse rates for each group for 3 years. Organ damage was assessed with the Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index (SDI). There were 22 patients on monthly IVCY, 18 on ELNT-IVCY, 13 on TAC, and 11 on MMF. Lower systemic lupus erythematosus disease activity index (SLEDAI) and higher CH50 were found in the TAC group at baseline (p<0.01 and p<0.01, respectively). There were no significant differences of cumulative CR rates and relapse free survival for 3 years among the four different therapeutic regimens (p = 0.2 and p = 0.2, respectively). There was a tendency to have early response and early relapse in TAC group and late response in MMF group. The SDI increase over 3 years was found more frequently in the TAC group than in the monthly-IVCY group (p = 0.04). Multivariate analysis indicated that CR at 3 months was independent prognosticator for low damage accrual. Regarding lower damage accrual, early CR achievement might be essential in induction therapy regardless of immunosuppressant choice.

Cutaneous polyarteritis nodosa associated with HLA-B39-positive undifferentiated spondyloarthritis in a Japanese patient

We present the case of a 43-year-old man diagnosed with HLA-B39-positive spondyloarthritis who developed cutaneous lesions consistent with cutaneous polyarteritis nodosa (CPN). Previous studies indicated an elevated incidence of HLA-B39 in HLA-B27-negative Japanese patients with spondyloarthritis. This case suggested that CPN may also occur in association with forms of HLA-B39-positive spondyloarthritis. The rarity of this association is emphasized. Therapy with corticosteroid and methotrexate improved both the cutaneous lesions and the clinical symptoms of spondyloarthritis.

Renal protective effect of antiplatelet therapy in antiphospholipid antibody-positive lupus nephritis patients without antiphospholipid syndrome

Objective We sought to evaluate the effect of antiplatelet therapy in addition to conventional immunosuppressive therapy for lupus nephritis (LN) patients positive for antiphospholipid antibodies (aPL) without definite antiphospholipid syndrome (APS). Methods Patients with biopsy-proven LN class III or IV were retrospectively evaluated. We selected patients positive for anticardiolipin antibody (aCL) or lupus anticoagulant (LA) who did not meet the criteria for a diagnosis of APS. The patients were divided into two subgroups according to whether antiplatelet therapy was received. The cumulative complete renal response (CR) rate, relapse-free rate, and change in estimated glomerular filtration rate (eGFR) over 3 years after induction therapy were calculated. Results We identified 17 patients who received antiplatelet therapy and 21 who did not. Baseline clinicopathological characteristics and immunosuppressive therapy did not show a significant difference between the two groups except for a higher incidence of LN class IV in the treatment group (p = 0.03). There was no difference in cumulative CR rate, relapse-free rate, or eGFR change between these subgroups. However, when data on LA-positive patients were assessed, an improvement in eGFR was found (p = 0.04) in patients receiving antiplatelet treatment. Conclusion Addition of anti-platelet therapy was associated with an improvement of eGFR in LA-positive patients with LN class III or IV.

Syndrome of inappropriate antidiuretic hormone secretion (SIADH) in a patient with microscopic polyangiitis following low-dose intravenous cyclophosphamide: a possible pathogenic link with disease activity

Abstract Syndrome of inappropriate antidiuretic hormone secretion (SIADH) induced by cyclophosphamide (CY) in rheumatology has been rarely described. Most reported cases developed SIADH at the first administration of high-dose CY. We report the case of a 76-year-old female with microscopic polyangiitis (MPA) who developed SIADH during the seventh course of low-dose intravenous cyclophosphamide (IVCY). Our report suggested that SIADH may be developed at any course of IVCY treatment and a process other than CY toxicity may be involved. An exacerbation of interstitial lung disease (ILD) and overexpression of inflammatory cytokines may have contributed to the development of SIADH in this case. Since the active phase of MPA might be a risk factor for SIADH, MPA patients should be evaluated for subclinical SIADH.

A positive direct Coombs’ test in the absence of hemolytic anemia predicts high disease activity and poor renal response in systemic lupus erythematosus

We determined the clinical utility of the direct Coombs’ test in the absence of hemolytic anemia as an indicator of disease activity and therapeutic response in systemic lupus erythematosus (SLE). SLE patients without hemolytic anemia who visited our hospital from January 2016 to November 2016 were retrospectively evaluated with a direct Coombs’ test. Clinical features, including SLE disease activity index (SLEDAI), treatment and laboratory findings were analyzed. For patients with lupus nephritis, we additionally evaluated the cumulative complete renal response rate over one year after induction therapy. Among 182 patients evaluated, 10 (5.8%) patients had a positive direct Coombs’ test in the absence of hemolytic anemia. They had a higher SLEDAI (p < 0.01), higher circulating immune complex levels (p = 0.01), higher anti-DNA titers (p < 0.01) and a lower complete renal response rate (p = 0.03) compared with those who were negative. Multivariate analysis indicated that SLEDAI was an independent factor correlated with the direct Coombs’ test without hemolytic anemia (odds ratio 2.4, 95% confidence interval 1.66–4.98, p < 0.01). A positive direct Coombs’ test in the absence of hemolytic anemia may therefore represent a useful biomarker for assessing disease activity and therapeutic response.

Rhabdomyolysis in a Patient with Polyarteritis Nodosa

Polyarteritis nodosa (PAN) is a medium vessel vasculitis affecting systemic organs. Muscle involvement of PAN usually lacks elevation of creatinine kinase (CK). We herein report a case of PAN with rhabdomyolysis. A 71-year-old man was hospitalized because of muscle weakness of the lower limbs that persisted for 1 month. On a physical examination, rapidly progressive lower proximal muscle weakness and bilateral drop foot were observed. His blood test showed an elevation in the C-reactive protein (19.5 mg/dL) and CK (13,435 IU/L) levels and negativity for anti-neutrophilic cytoplasmic antibody. Computed tomographic angiography showed stenosis of the left renal artery. Electromyogram indicated mono-neuritis multiplex pattern, and enhanced magnetic resonance imaging demonstrated discretely granular hyperintensities on T2 and slow tau inversion recovery in his femoral muscles. A femoral muscle-biopsy specimen showed fibrinoid necrosis of medium-sized vessels and disruption of the elastic lamina of the vessel wall in fascia. Furthermore, muscle necrosis was localized depending on the arterial distribution, suggesting ischemic changes in the muscles. Given these findings, he was diagnosed with PAN with rhabdomyolysis and treated with methyl-prednisolone pulse therapy followed by oral prednisolone at 50 mg/day. He was additionally treated with monthly intravenous cyclophosphamide at 500 mg. Sustained remission has been obtained for two months since the treatment. Although rhabdomyolysis rarely manifests with PAN, it should be included in a differential diagnosis of febrile patients presenting with acute myalgia and weakness with CK elevation.