Correlation of serum citrullinated histone H3 levels with disease activity in patients with rheumatoid arthritis.

OBJECTIVES: Serum citrullinated histone H3 (CitH3) levels in humans with rheumatoid arthritis (RA) were examined and the associations between CitH3 levels and disease variables were investigated. METHODS: Serum CitH3 levels were measured using an enzyme-linked immunosorbent assay in 151 RA patients (69 with highly, 32 with moderately, and 20 with mildly active RA and 30 with RA in remission) and 56 healthy controls. Receiver operating characteristic curve analysis was performed to evaluate the discriminant capacity of CitH3 between highly/moderately active RA and RA in remission/mild activity. Furthermore, machine-learning methods were applied to construct a predictive model. RESULTS: CitH3 concentration was more upregulated in patients with highly and moderately active RA than in those with mild activity and remission. The area under the curve for CitH3 was 0.825 for discriminating between highly and mildly active RA, 0.840 for discriminating between highly active RA and RA in remission, 0.789 for discriminating between moderately and mildly active RA, and 0.829 for discriminating between moderately active RA and RA in remission. The correlation analysis revealed that serum CitH3 levels were positively associated with Disease Activity Score 28-Erythrocyte Sedimentation Rate (DAS28-ESR), ESR, C-reactive protein, rheumatoid factor, and some routine blood parameters (WBC, RDW, PLT, N, and N%), while negatively associated with haemoglobin, lymphocyte percentage, and thyroid-stimulating hormone. Through machine learning, the optimal predictive model was selected and had high performance. CONCLUSIONS: CitH3 is significantly associated with disease activity and could serve as a useful candidate biomarker to assess disease activity in patients with RA.

Low serum level of citrullinated histone H3 in patients with dermatomyositis.

OBJECTIVES: We aimed at analyzing the serum levels of citrullinated histone H3 (CitH3) in patients with dermatomyositis (DM) and their association with disease activity. METHODS: Serum CitH3 levels were measured using enzyme-linked immunosorbent assays in serum samples obtained from 93 DM patients and 56 healthy controls (HCs). Receiver operating characteristic (ROC) curve analysis was performed to evaluate the discriminant capacity of CitH3 and other disease variables. The association between CitH3 and disease variables was analyzed using Pearson's rank correlation. RESULTS: Serum CitH3 level was significantly lower in DM patients than in HCs (p < 0.001). The ROC curve analysis revealed that CitH3 strongly discriminated DM patients from HCs (area under the curve [AUC], 0.86), and a combination of CitH3 and the ratio of neutrophil to lymphocyte counts (NLR) showed a greater diagnostic value (AUC, 0.92). Serum CitH3 levels were markedly lower in DM patients with normal muscle enzyme levels than in HCs (all p < 0.001), and when compared to an elevated group, the CitH3 levels were comparable (all p > 0.05). The CitH3 levels showed no difference between DM in active and remission groups. However, in a paired test with 18 hospitalized DM patients, the CitH3 levels were higher in remission state than in active state. Moreover, the CitH3 levels showed no correlation with disease variables that were associated with the disease activity of DM. CONCLUSIONS: Serum CitH3 level may serve as a useful biochemical marker for screening patients with DM from HCs, while its role in monitoring DM disease activity requires further research.

Anti-citrullinated protein antibodies are associated with neutrophil extracellular trap formation in rheumatoid arthritis.

OBJECTIVES: We evaluated the associations of anti-citrullinated protein antibodies (ACPAs) and serological and cytological levels of neutrophil extracellular trap (NET) formation in rheumatoid arthritis (RA) patients. METHODS: Serum levels of myeloperoxidase-DNA and elastase-DNA complexes (NET remnants) were examined in 51 patients with RA and 40 healthy controls using a modified enzyme-linked immunosorbent assay. Neutrophils were isolated by density gradient centrifugation. IgG antibodies were purified by affinity chromatography. NET formation in RA and control neutrophils was assessed by microscopy in vitro. NETs were purified and co-incubated with fibroblast-like synoviocyte (FLS) cells. Interleukin (IL)-6 and IL-8 mRNA expression and protein levels in FLS cells were determined by real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. RESULTS: In RA patients, NET remnants in the peripheral circulation were higher in extremely high ACPA titers when compared to in moderate ACPA titers. And IgG antibodies containing ACPA can stimulate neutrophils to form NETs in a concentration-dependent manner. Furthermore, significantly higher expression of the pro-inflammatory cytokines IL-6 and IL-8 is detected after FLS cells interacted with NETs which derived from neutrophils stimulated with ACPA-containing IgG antibodies. CONCLUSIONS: Anti-citrullinated protein antibodies may enhance NET formation and contribute to inflammation development in RA by stimulating NET formation, such as by subsequent activation of FLS cells by NETs.

Correlation of PMN elastase and PMN elastase-to-neutrophil ratio with disease activity in patients with myositis.

BACKGROUND: Polymorphonuclear (PMN) elastase plays an important role in a variety of inflammatory disorders. Our aim was to analyse PMN elastase in idiopathic inflammatory myopathies (IIMs) and its association with disease activity. METHODS: PMN elastase levels were measured using enzyme-linked immunosorbent assay in serum samples obtained from 74 patients with myositis (58 with dermatomyositis [DM] and 16 with polymyositis [PM]) and 22 healthy controls. Receiver operating characteristic (ROC) curve analysis was performed to evaluate the discriminant capacity of PMN elastase level and PMN elastase-to-neutrophil ratio (ENR) in patients with active and remission myositis. The association of serum PMN elastase level and ENR with disease variables was evaluated in patients with IIMs. The disease specificity of PMN elastase level and ENR was further examined in 60 patients with other systemic autoimmune diseases. RESULTS: PMN elastase level and ENR were significantly higher in patients with active IIMs, DM, and PM than in patients with remission. ROC curve analysis revealed that PMN elastase level and ENR both outperformed creatine kinase (CK), the currently used laboratory marker, and strongly discriminated patients with active disease and those with remission of IIMs, DM, and PM (area under the ROC curve [AUC] 0.9, 0.9, and 0.88 for PMN elastase; AUC 0.96, 0.96, and 1.0 for ENR; AUC 0.72, 0.70, and 0.80 for CK, respectively). PMN elastase level and ENR were positively correlated with myositis disease activity assessment, CK, lactate dehydrogenase, aspartate aminotransferase, alanine aminotransferase, C-reactive protein, and erythrocyte sedimentation rate. PMN elastase level and ENR were higher in the anti-PM-Scl positive myositis group than those in the anti-PM-Scl negative myositis group. Nevertheless, PMN elastase was not a specific disease marker for IIMs when compared with other autoimmune diseases. CONCLUSIONS: PMN elastase, particularly ENR, were significantly correlated with disease activity and could serve as useful biochemical markers for evaluating the disease activity of patients with IIMs. Thus, they are potentially helpful in monitoring disease progression and guiding treatment.

Increased levels of serum histone H4 and activated protein C in patients with active rheumatoid arthritis.

OBJECTIVES: We aimed to examine the levels of serum H4 and activated protein C (APC) in rheumatoid arthritis (RA) and other autoimmune conditions, and investigate the associations between H4 or APC levels and disease activity indicators in RA. METHODS: Serum H4 and APC distribution was examined in samples from patients with RA, systemic lupus erythematosus (SLE), polymyositis (PM), and ankylosing spondylitis (AS), as well as in samples from healthy controls, using commercial ELISA kits. Associations of serum H4 or APC levels with disease variables in patients with RA were evaluated. Receiver operating characteristic (ROC) curve analysis was performed to assess the discriminant capacity of APC against RA and non-RA. RESULTS: The patients with RA, PM, and AS showed higher serum levels of H4 and APC than those from the healthy control individuals, while the SLE patients showed higher serum levels of APC only. Moderate positive correlations between H4 levels and absolute neutrophil count (ANC), platelet count (PLT), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), brinogen (FIB), D-dimer (DD), and complement fraction 3 (C3) were observed. Positive correlations between APC levels and PLT, RF, DD, or DAS28 were additionally found. ROC curve analysis revealed that APC discriminated well between RA and non-RA. CONCLUSIONS: H4 and APC concentrations are elevated in patients with chronic in ammatory autoimmune diseases. The observed associations between H4 and APC and disease variables in patients with RA support a role for H4 and APC in the in ammatory process of the disease.

Increased rates of indeterminate results in QuantiFERON-TB gold in-tube during the COVID-19 epidemic in Hunan province, China.

Increased rates of indeterminate QuantiFERON-TB Gold In-Tube (QFT-GIT) results have been reported since the COVID-19 epidemic in Hunan Province, China. The indeterminate result (ITR) rate of QFT increased from an average of 5.2% to 12.4%, paralleling the first COVID-19 pandemic wave in the region. QFT-GIT results of 243 hospitalized patients with COVID-19 from January 2022 to April 2023 at Xiangya Hospital of Central South University were analyzed. Of the 243 patients, 71 (29.2%) had ITRs due to reduced interferon-gamma production in the positive control. Multiple factors are associated with ITRs, such as disease severity, respiratory failure incidence, immunosuppressant use, and prognosis. Additionally, interferon-gamma (Mitogen-Nil) levels differed significantly depending upon disease severity, prognosis, immunosuppressant use, sepsis symptoms, respiratory failure, or hyperlipidemia. An abnormal increase in the ITR rate in the QFT was observed after the COVID-19 pandemic, and an optimal machine learning predictive model for indeterminate QFT results was established.

Indeterminate results of QuantiFERON-TB Gold in-tube due to high nil concentration and methodological exploration and improvement.

BACKGROUND: The features of indeterminate results (ITRs) due to high interferon gamma (IFN-γ > 8.0 IU/mL) concentrations in Nil tubes of QuantiFERON-TB Gold in-Tube (QFT-GIT) have not been well studied. Therefore, this study aimed at investigating the features of ITRs and optimization alternatives for this method. METHODS: We used the plasma exchange method to reduce the ITR rate due to high Nil concentrations (Nil > 8.0 IU/mL) between March 2020 and September 2021 at Xiangya Hospital of Central South University in China. One hundred and eleven out of 28,193 patient samples were considered ITRs due to Nil > 8.0 IU/mL. Of these 111, blood was re-sampled from 82 patients (pretreated group) who received the plasma exchange. Moreover, 40 out of 82 (blood volume ≥ 5 mL) received pretreated QFT-GIT and standard QFT-GIT (untreated group) simultaneously, while the remaining 42 received pretreated QFT-GIT only. The statistical difference between groups was evaluated. Cohen's kappa coefficient was used to assess the level of agreement between the pretreated and untreated group. RESULTS: Of the 28,193 subjects, 1,083 (3.8%) had ITRs, and 111 (0.4%) had ITRs due to Nil > 8.0 IU/mL. The population studied had a mean age of 52.9 years, and 70.3% were men, which is a larger proportion than that in patients with ITR and the overall population. Significantly decreased IFN-γ levels in Nil tubes were detected using the pretreated QFT-GIT compared with standard QFT-GIT (p < 0.01). The determinate results rate in the pretreated group was significantly higher than that of the untreated group (80% (32/40) vs 57.5%, (23/40), p = 0.03). Further comparison revealed that the pretreated group was consistent with the untreated group in 17/20 (85%) positive tests, 3/3 (100%) negative tests, and 6/17 (35.3%) ITRs. The overall agreement rate was 26/40 (65%) among all 40 subjects, and the κ value was 0.39 (minimal agreement). The majority of results obtained after pretreatment were positive (71.2%, 59/82) and the agreement rate between clinical diagnosis and pretreated QFT-GIT was at least 61.0% (39/59). CONCLUSION: Plasma exchange pretreated QFT-GIT yields more reliable results than untreated QFT-GIT when processing high Nil concentrations.

Increased levels of neutrophil extracellular trap remnants in the serum of patients with rheumatoid arthritis.

AIMS: Neutrophil extracellular traps (NETs) comprise a unique form of non-apoptotic cell death exhibited by neutrophils, which occurs in a stepwise process termed NETosis. It has been postulated that NETosis plays an important role in the pathogenesis of autoimmune disorders. The aim of this study was to evaluate serum levels of NET remnants in patients with rheumatoid arthritis (RA), as well as potential associations between NET remnants and indicators of RA. METHODS: Serum levels of myeloperoxidase (MPO)-DNA complexes (NET remnants) were examined in 74 RA patients and 50 healthy controls using a modified enzyme-linked immunosorbent assay. Associations between the levels of these complexes and indicators of RA were then statistically evaluated. RESULTS: RA patients exhibited significantly higher levels of MPO-DNA complexes than the healthy controls, and these levels were associated with increased neutrophil counts and positivity for rheumatoid factor (RF) and anti-citrullinated protein/peptide antibodies (ACPA). Among the 63 ACPA-positive RA patients examined, those with ACPA titers > 1600 U/mL showed significantly increased MPO-DNA levels. Receiver operating characteristic analysis determined that the area under the curve for all 74 RA patients was 0.798, with a sensitivity of 91.9% and a specificity of 56.0%, while that for the ACPA-negative patients was 0.891, with a sensitivity and specificity of 81.8% and 84.0%, respectively. CONCLUSIONS: The results of this study suggest that the disease status of RA is associated with increased NETosis. In particular, evaluation of serum MPO-DNA levels may comprise a useful complementary tool for discriminating RA patients from healthy individuals.

Evaluation of our self-designed nanometer silicon membrane sandwich cup system for diagnosing tuberculosis.

BACKGROUND AND AIMS: A nanometer silicon membrane sandwich cup system was self-designed. It could concentrate the bacilli via 0.45-µm microporous filter membrane and semi-automate the acid-fast bacilli (AFB) by a bacteria-staining machine. The aim of the study was to assess the clinical value of our self-designed system for diagnosing tuberculosis (TB). METHODS: A total of 1993 sputum specimens obtained from patients with confirmed or suspected TB were subjected to direct or concentrated specimens smear at XiangYa Hospital, Central South University between May 2012 and February 2013. In addition, all the specimens were also inoculated into Lowenstein-Jensen (L-J) media, and culture results were considered as the gold standard for calculating sensitivity and specificity. RESULTS: Compared with direct smear examination, an increased density of red stained bacilli was observed in the self-designed nanometer silicon membrane sandwich cup analysis under the microscope. The positive rate of the self-designed analysis was significantly higher than that of direct AFB smear [10.9% (217/1993) vs 6.2% (123/1993), P < 0.05]. The sensitivity of the self-designed system increased (97.3% vs 55.2%, P < 0.05) without a loss of specificity (100% vs 100%) for identifying positive TB cases compared with the direct smear method. CONCLUSION: The self-designed nanometer silicon membrane sandwich cup and semi-automatic bacteria-staining machine could more efficiently and rapidly detect the AFB in respiratory specimens than direct microscopy. This is a novel and safe examination, and it may replace direct smear examination for the diagnosis of patients with TB.

Detection of the novel IMP-38 among carbapenemase-producing Enterobacteriaceae in a university hospital, China.

INTRODUCTION: This study set out to investigate the molecular epidemiology of carbapenemase-producing Enterobacteriaceae isolates collected from Xiang Ya Hospital, Hunan, China. METHODOLOGY: The carbapenemase genes from Enterobacteriaceae isolates were determined by PCR and sequencing. Relatedness of Klebsiella pneumoniae isolates was evaluated by pulsed-field gel electrophoresis. RESULTS: Twenty-four out of 738 non-repetitive Enterobacteriaceae isolates harbored carbapenemase genes including IMP-38, a novel IMP-type metallo-enzyme. Nine IMP-38-producing isolates were shown to originate from the same clone and caused a small outbreak in the neonatal ward. CONCLUSIONS: IMP-38, a novel IMP-type metallo-enzyme, was one of the predominant types of carbapenemase in the clinical carbapenem-resistant Enterobacteriaceae isolates in our hospital.