Novel Biomarker Of Disease Activity Research Articles

18F-Sodium fluoride PET-CT visualizes disease activity in chronic nonbacterial osteitis in adults.

Chronic nonbacterial osteitis (CNO) is a rare disease spectrum, which lacks biomarkers for disease activity. Sodium fluoride-18 positron emission tomography/computed tomography ([18F]NaF-PET/CT) is a sensitive imaging tool for bone diseases and yields quantitative data on bone turnover. We evaluated the capacities of [18F]NaF-PET/CT to provide structural and functional assessment in adult CNO. A coss-sectional study was performed including 43 adult patients with CNO and 16 controls (patients referred for suspected, but not diagnosed with CNO) who underwent [18F]NaF-PET/CT at our expert clinic. Structural features were compared between patients and controls, and maximal standardized uptake values (SUVmax [g/mL]) were calculated for bone lesions, soft tissue/joint lesions, and reference bone. SUVmax was correlated with clinical disease activity in patients. Structural assessment revealed manubrial and costal sclerosis/hyperostosis and calcification of the costoclavicular ligament as typical features associated with CNO. SUVmax of CNO lesions was higher compared with in-patient reference bone (mean paired difference: 11.4; 95% CI: 9.4-13.5; p < .001) and controls (mean difference: 12.4; 95%CI: 9.1-15.8; p < .001). The highest SUVmax values were found in soft tissue and joint areas such as the costoclavicular ligament and manubriosternal joint, and these correlated with erythrocyte sedimentation rate in patients (correlation coefficient: 0.546; p < .002). Our data suggest that [18F]NaF-PET/CT is a promising imaging tool for adult CNO, allowing for detailed structural evaluation of its typical bone, soft-tissue, and joint features. At the same time, [18F]NaF-PET/CT yields quantitative bone remodeling data that represent the pathologically increased bone turnover and the process of new bone formation. Further studies should investigate the application of quantified [18F]NaF uptake as a novel biomarker for disease activity in CNO, and its utility to steer clinical decision making.

The emerging role of Growth Differentiation Factor 15 as a potential disease biomarker in juvenile dermatomyositis.

We aimed to investigate the potential of Growth Differentiation Factor 15 (GDF-15) as a novel biomarker for disease activity in Juvenile Dermatomyositis (JDM). We recruited children with juvenile myositis including juvenile dermatomyositis (n = 77), polymyositis (n = 6), and healthy controls (n = 22). GDF-15 levels in plasma were measured using ELISA. Statistical analyses were performed using non-parametric tests. Levels of GDF-15 were significantly elevated in JDM compared with healthy controls (p< 0.001). GDF-15 levels exhibited strong positive correlations with disease activity scores, including the Disease Activity Score (DAS) total score, DAS skin score, DAS muscle score, and Childhood Myositis Assessment Scale (CMAS). Additionally, GDF-15 levels could differentiate between active disease and remission based on the Physician Global Assessment of muscle score. Positive correlations were observed between levels of GDF-15 and creatine kinase, neopterin, and nailfold end row loops, indicating the potential involvement of GDF-15 in muscle damage, immune activation, and vascular pathology. ROC curve analysis showed GDF-15 to be more effective in assessing disease activity in JDM than creatine kinase (AUC 0.77, p= 0.001 and AUC 0.6369, p= 0.0738, respectively). GDF-15 may serve as a valuable biomarker for assessing disease activity in JDM. It exhibits better sensitivity and specificity than creatine kinase, and the levels correlate with various disease activity scores and functional measures. GDF-15 may provide valuable information for treatment decision-making and monitoring disease progression in JDM.

Urine L-selectin reflects clinical and histological renal disease activity and treatment response in lupus nephritis across multi-ethnicity.

There is an urgent need for novel biomarkers in lupus nephritis (LN). We report a non-invasive urinary biomarker, L-selectin, in two independent multi-ethnic cohorts. uL-selectin was tested cross-sectionally in a Chinese cohort (n=255) and a US cohort (n=219) of SLE patients and controls using ELISA. A longitudinal cohort includes 20 active Chinese LN patients. uL-selectin was significantly increased in active LN patients compared to active non-renal SLE, inactive LN, inactive non-renal SLE, chronic kidney disease patients, and healthy controls. uL-selectin positively correlated with global and renal disease activities as well as histological activity index and chronicity index (CI). Low uL-selectin was an independent predictor for high CI. During follow-up, uL-selectin levels decreased significantly in the complete renal remission group. uL-selectin is a novel biomarker of disease activity and renal histopathology in LN across multiple ethnicities. It also reflects treatment response in LN patients during follow up.

Clinical Significance of Immature Neutrophils in Relation to Disease Activity in Psoriatic Arthritis Patients

Abstract Background Psoriatic arthritis (PsA) is a complex type of progressive inflammatory arthritis associated with psoriasis. Prevalence of PsA has been reported 5.8–40.9% among patients suffering from psoriasis. A recent meta-analysis indicated that there is no gender predilection in patients with PsA. It shares genetic and clinical features with other forms of spondyloarthritis and affects different structures of the musculoskeletal system in addition to the skin and the nail. Neutrophils are crucial responders of the innate immune response, but the role played by neutrophils in psoriasis has not been well characterized. This thesis discussed the current understanding of the role and potential mechanisms of immature neutrophils in psoriatic arthritis. Objective To study the relation between immature neutrophils in blood and disease activity in psoriatic arthritis patients. Methods This randomized control trial, done on 50 PsA patients from Physical medicine, Rheumatology and Rehabilitation and Dermatology outpatient clinics of Ain Shams University hospital, included by CASPAR criteria, subjected to full medical history taking and thorough clinical examination with calculation of SPARCC, ASDAS and PASI scores.laboratory evaluation include CBC with differential white blood cell count for Immature Neutrophils percentage, ESR, CRP and Rheumatoid factor.CBC was compared with 20 age and sex matched healthy controls. Results The Immature Neutrophils mean ± SD(%) was found to be 5.99±1.57 in patients and was statistically significantly higher in them than in controls 2.25±1.16(P &amp;lt; 0.037) and it was statistically significantly correlated with disease activity according to ASDAS score, SPARCC, PASI score, ESR, CRP, RBCS, WBCS, Platelets, Enthesitis, Dactylitis, skin and nail affection. Conclusion This study highlighted the role of immature neutrophils as a novel biomarker for disease activity in psoriatic arthritis.

P122 Changes in Serum Levels of Cytokines with Methotrexate Monotherapy in Active Rheumatoid Arthritis: Results from a Multicentre, Randomized Controlled Trial on Methotrexate Escalation In Rheumatoid Arthritis

Abstract Background/Aims Identification of novel biomarkers of disease activity and prediction of treatment response in rheumatoid arthritis (RA) is an area of active research. Reliable biomarkers of methotrexate (MTX) response in RA are yet to be identified. We sought to study the change in serum levels of IL-6, MMP-3, and TNF-α with MTX monotherapy in patients with active RA. Methods Patients with active RA (SJC≥2 and TJC≥4) not on DMARDs (except HCQ and low-dose prednisolone) who had been enrolled in the multicentre, parallel-group RCT comparing two different MTX escalation strategies in RA (MEIRA) were included. All these patients received MTX monotherapy started at 15 mg/week and escalated to 25 mg/week by 4-8 weeks. Serum levels of IL-6, MMP-3, and TNF-α were measured by ELISA before starting MTX and after 16 weeks of MTX therapy. Responders were defined as patients who achieved a EULAR good or moderate response (based on DAS28ESR-3v) at 16 weeks. Analyses were intention-to-treat. Results 16 weeks of MTX therapy led to a decline in serum levels of both MMP-3 (p = 0.03, n = 129) and IL-6 (p = 0.048, n = 151). The decline in IL-6 levels was seen only in MTX responders (p = 0.003), but not in non-responders, while the decline in MMP-3 levels was seen in both MTX non-responders (p = 0.015) and responders (p = 0.44) (Table 1). Baseline (pre-treatment) serum levels of both IL-6 and MMP-3 were similar between MTX responders and non-responders. Correlations of serum levels of IL-6 (but not MMP-3) with measures of disease activity (DAS28, tender joint count) were weak but significant at 16 weeks. Serum TNF-α levels (n = 137) were undetectable in a majority of patients (93%). Serum IL-6 was not detectable in 67 (44%) patients at baseline. Conclusion Although serum levels of both MMP-3 and IL-6 reduced significantly with MTX therapy, the decline of IL-6 was seen only in MTX responders but not in non-responders. Pre-treatment serum levels did not predict future MTX response or non-response. IL-6 levels also correlated weakly but significantly with disease activity. Serial measurement of serum IL-6 merits further exploration in larger studies as a potential biomarker of MTX response in RA. Disclosure S. Jain: None. V. Dhir: None. A. Aggarwal: None. R. Gupta: None. S. Maurya: None. B. Leishangthem: None. A. Khullar: None. S. Naidu: None. S.K. Sharma: None. A. Sharma: None. S. Jain: None.

Hsa_circ_0056856 in the serum serves as a potential novel biomarker for disease activity in psoriasis.

To the Editor: Psoriasis is a chronic immune-related inflammatory skin disease. The clinical severity of psoriasis varies greatly, and recurrence occurs frequently.[1] Many attempts have been made to identify biomarkers for psoriasis; however, there are currently no specific markers that can accurately monitor disease severity and treatment efficacy. Circular RNAs (circRNAs) are a novel class of endogenous noncoding RNAs that are highly stable and resistant to RNase R since they can form covalently closed loops without 5′ caps and 3′ tails.[2] It has been reported that circRNAs could exist in body fluids, such as serum and plasma, and could serve as molecular biomarkers in the diagnosis and prognosis of several diseases.[3] In this study, we detected the alteration of circRNAs and investigated the potential value of hsa_circ_0056856 as a biomarker for disease severity in serum samples from psoriasis patients and healthy controls (HCs). A total of 125 patients diagnosed with psoriasis vulgaris were recruited from the Seventh Medical Center of PLA General Hospital. We collected all the patient serum samples and 20 patient peripheral blood mononuclear cell (PBMC) samples from the same cohort at first admission to the hospital. We also obtained 30 patient serum samples after treatment for 7 to 8 weeks. Disease severity was assessed by the psoriasis area and severity index (PASI). None of the patients had other autoimmune diseases or were on systemic treatment at least 1 month prior to study participation. Serum samples from 75 healthy subjects, whose age and sex were matched with the psoriasis group, were collected at the Medical Examination Center of the same hospital. Details of all samples are summarized in Supplementary Table 1, https://links.lww.com/CM9/B135. This study was conducted in adherence with the principles of the Declaration of Helsinki and was approved by the Ethics Committee of the Seventh Medical Center of PLA General Hospital (No. 2020-084). Written informed consent was obtained from all participants. First, we performed a circRNA microarray to identify the expression patterns of circRNAs in serum from five psoriasis patients and five normal controls. A total of 11,582 circRNAs were detected, and a box plot showed a similar distribution of log2 ratios among the ten samples after quantile normalization [Supplementary Figure 1A, https://links.lww.com/CM9/B134]. The genomic locations of their source genes were exonic (9741, 84.10%), intronic (798, 6.89%), sense overlapping (751, 6.48%), antisense overlapping (205, 1.77%), and intergenic (87, 0.75%) [Supplementary Figure 1B, https://links.lww.com/CM9/B134]. Approximately 66.32% (7681/11,582) of the identified circRNAs were >2k bp in length [Supplementary Figure 1C, https://links.lww.com/CM9/B134], and the chromosome distribution of the circRNAs showed that chromosome 1 had the most (1165), while chromosome 21 had the fewest (123) [Supplementary Figure 1D, https://links.lww.com/CM9/B134]. To explore the differentially expressed (DE) profiles of serum circRNAs in psoriasis, hierarchical clustering analysis was employed, and the results showed that the circRNA expression was significantly changed between psoriasis patients and HCs [Supplementary Figure 2A, https://links.lww.com/CM9/B134]. A total of 132 circRNAs were significantly changed (|log2 (fold change)| > 1 and P < 0.05) in serum from psoriasis patients, with 123 upregulated and nine downregulated circRNAs, compared with HCs [Supplementary Figure 2B, https://links.lww.com/CM9/B134 and Supplementary Table 2, https://links.lww.com/CM9/B136]. Among them, the most upregulated circRNA was hsa_circ_0056856, and the circRNA showing the greatest downregulation was hsa_circ_0112550. Subsequently, we screened DE circRNAs based on the raw intensity, fold-change, and statistical value in the RNA-seq data. Ten DE circRNAs were selected for further analysis by real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR) in serum samples from 20 psoriasis patients and 20 HCs. All primers were synthesized by Sangon Biotech (Shanghai, China) and are listed in Supplementary Table 3, https://links.lww.com/CM9/B137. The results showed that six circRNAs, including hsa_circ_0056856 (U = 400, Z = 5.41, P < 0.001) (Mann-Whitney U test, U-value represents the difference of rank-sum and Z score means the difference of median between these two groups), hsa_circ_0023461 (U = 286, Z = 2.326, P = 0.02), hsa_circ_0082326 (U = 291, Z = 2.462, P = 0.013), hsa_circ_0001255 (U = 298, Z = 2.651, P = 0.007), hsa_circ_0055440 (U = 282, Z = 2.218, P = 0.026), and hsa_circ_0112550 (U = 117, Z = −2.245, P = 0.024), were consistent with the RNA-seq data, among which hsa_circ_0056856 exhibited the most significant changes and was finally selected for further validation [Figure 1A].Figure 1: hsa_circ_0056856 as a biomarker for disease severity in psoriasis. (A) The expression profiles of selected circRNAs in serum between 20 psoriasis patients and 20 HCs by RT-qPCR validation. (B) The ROC curve of hsa_circ_0056856 was calculated based on the −ΔCt value in serum from 100 psoriasis patients and 50 HCs. (C) Correlation between serum levels of hsa_circ_0056856 and PASI scores in 100 psoriasis patients. (D) hsa_circ_0056856 expression was measured before and after 7 to 8 weeks of treatment in 30 psoriasis patients by RT-qPCR. ∗ P < 0.05; ∗∗ P < 0.01; ∗∗∗ P < 0.001. circRNAs: Circular RNAs; HCs: Healthy controls; PASI: Psoriasis area and severity index; ROC: Receiver operating characteristic; RT-qPCR: Real-time reverse transcription quantitative polymerase chain reaction.To explore the diagnostic value of hsa_circ_0056856 as a biomarker for psoriasis, we analyzed the expression of hsa_circ_0056856 in a larger cohort of serum samples, including 100 psoriasis patients and 50 HCs. In addition, PBMC samples from 30 patients and 20 controls were also collected from the same cohort [Supplementary Table 1, https://links.lww.com/CM9/B135]. Consistent with the above findings, hsa_circ_0056856 was dramatically increased in both psoriasis patients’ serum (U = 735, Z = 7.037, P < 0.001) and PBMCs (U = 106, Z = 3.842, P < 0.001) compared with HCs [Supplementary Figure 3A, B, https://links.lww.com/CM9/B134]. Moreover, receiver operating characteristic curve analysis showed that the area under curve value of hsa_circ_0056856 was 0.853 (95% CI = 0.792–0.914, P < 0.0001) (Wilson/Brown method) [Figure 1B]. These results suggested that hsa_circ_0056856 could be a potential biomarker for psoriasis diagnosis. Next, we evaluated the potential value of hsa_circ_0056856 as a biomarker for disease severity in psoriasis. We measured the correlation between the expression of hsa_circ_0056856 and the PASI score. Our results identified that the hsa_circ_0056856 levels in serum were positively correlated with PASI scores in psoriasis patients (r = 0.422, P < 0.0001) (Spearman's rank correlation coefficient) [Figure 1C]. In addition, we detected the expression of hsa_circ_0056856 in serum obtained from 30 patients at baseline and after 7 to 8 weeks of treatment. All patients achieved a significant improvement in PASI score [Supplementary Table 1, https://links.lww.com/CM9/B135], and the expression of hsa_circ_0056856 exhibited a significant downregulation after treatment (Z = 2.725, P = 0.006) (Wilcoxon signed-rank test, Z score means the difference of median between these two groups) [Figure 1D]. Previous studies have found that circRNAs can indirectly regulate gene expression by serving as miRNA sponges. Therefore, we predicted the potential miRNAs that could bind to hsa_circ_0056856 using Arraystar's homemade miRNA target prediction software. The top five miRNA response elements were selected as follows: miR-1301-3p, miR-140-3p, miR-93-3p, miR-19b-3p, and miR-187-5p. Next, we analyzed the potential targets of the five miRNAs using three online websites (TargetScan, TarBase, and MicroT-CDS), and 43, 82, 52, 23, and 78 candidate genes were screened out. A hsa_circ_0056856-miRNA- target mRNA interaction network was constructed through a ceRNA mechanism using Cytoscape software [Supplementary Figure 4A, https://links.lww.com/CM9/B134]. To further understand the underlying molecular mechanism of hsa_circ_0056856, we performed GO biological process enrichment analysis to investigate the functions of these target genes. The results showed that the target mRNAs in the ceRNA network were mainly involved in the regulation of protein serine/threonine kinase activity, histone modification, covalent chromatin modification, and peptidyl-lysine modification [Supplementary Figure 4B, https://links.lww.com/CM9/B134]. These results indicate that hsa_circ_0056856 could participate in epigenetic regulation in psoriasis through a ceRNA mechanism. However, further functional verification is required in future studies. Accumulating evidence supports the use of circRNAs as diagnostic and therapy response biomarkers of disease.[4] Because circRNAs are constantly stable and specific in serum and serum is easier to isolate, transport, and preserve in the clinic, these circRNAs could be used as therapeutic response and diagnostic tools for various diseases as well as potential viable therapeutic targets.[3] hsa_circ_0056856 expression has been reported in several cell types including Nhek, Mcf7, A549, and HMEC cells.[5] In the present study, we revealed that hsa_circ_0056856 was significantly increased in serum from psoriasis patients compared with HCs and could serve as an ideal biomarker for psoriasis diagnosis, severity, and therapeutic response evaluation. Moreover, we identified that hsa_circ_0056856 may participate in the pathogenesis of psoriasis through a ceRNA mechanism based on bioinformatic analysis, which could indirectly regulate gene expression and affect the histone modification biological process. Funding This work was supported by grants from the National Natural Science Foundation of China (Nos. 81773316 and 82002120) and the National Science Foundation of Chongqing, China (No. cstc2021jcyj-msxmX0140). Conflicts of interest None.

Serum levels of fetuin-A as a novel biomarker for disease activity in patients with Takayasu arteritis and granulomatous polyangiitis.

The aim of the present study was to investigate serum fetuin-A (Fet-A) levels in patients with Takayasu arteritis (TA) and granulomatous polyangiitis (GPA) and to analyze the relationship between serum Fet-A levels and disease activity scores. Thirty-two TA and 28 GPA patients presented to the rheumatology clinic at Gazi University and met the criteria of American College of Rheumatology 1990 and 2012 International Chapell Hill meeting, respectively, and 20 healthy control subjects were included in the present study. We collected data on serum C-reactive protein (CRP), albumin, calcium, and phosphate levels as well as erythrocyte sedimentation rates. Calcification risk index (CRI) was calculated for each patient. The Birmingham Vasculitis Activity Score (BVAS) and Indian Takayasu Clinical Activity Score (ITAS), were used to assess disease activity in GPA and TA patients respectively. Serum Fet-A levels were significantly lower in the overall vasculitis group compared to control group (p = 0.015). In subgroup analysis, Fet-A levels were significantly lower in those with active disease, compared to control group (p = 0.001, for active TA (n = 18) and GPA (n = 17), respectively). However, there was no significant difference in serum Fet-A levels in inactive cases versus control subjects (p = 0.061, for inactive TA (n = 14) and GPA (n = 11), respectively). Serum Fet-A levels negatively correlated with BVAS (r =  - 0.675) and ITAS scores (r =  - 0.385), as well as with CRP and CRI. Our results suggest that serum Fet-A level could be a novel biomarker for assessment of activity status in patients with GPA or TA. Key Points • Serum Fetuin-A is negative acute phase protein and systemic calcification inhibitor synthesized in hepatocytes and secreted by various inflammation. • Serum Fetuin-A was negatively correlated with CRP, BVAS, and ITAS scores and significantly decreased in vasculitis patients with high disease activity. • Serum Fetuin-A could be a promising and useful biomarker for the assessment of disease activity for vasculitis, also that it might also be a predictor of long-term cardiovascular progression.

Integrative Analysis of the Inflammatory Bowel Disease Serum Metabolome Improves Our Understanding of Genetic Etiology and Points to Novel Putative Therapeutic Targets

Polygenic and environmental factors are underlying causes of inflammatory bowel disease (IBD). We hypothesized that integration of the genetic loci controlling a metabolite's abundance, with known IBD genetic susceptibility loci, may help resolve metabolic drivers of IBD. We measured the levels of 1300 metabolites in the serum of 484 patients with ulcerative colitis (UC) and 464 patients with Crohn's disease (CD) and 365 controls. Differential metabolite abundance was determined for disease status, subtype, clinical and endoscopic disease activity, as well as IBD phenotype including disease behavior, location, and extent. To inform on the genetic basis underlying metabolic diversity, we integrated metabolite and genomic data. Genetic colocalization and Mendelian randomization analyses were performed using known IBD risk loci to explore whether any metabolite was causally associated with IBD. We found 173 genetically controlled metabolites (metabolite quantitative trait loci, 9 novel) within 63 non-overlapping loci (7 novel). Furthermore, several metabolites significantly associated with IBD disease status and activity as defined using clinical and endoscopic indexes. This constitutes a resource for biomarker discovery and IBD biology insights. Using this resource, we show that a novel metabolite quantitative trait locus for serum butyrate levels containing ACADS was not supported as causal for IBD; replicate the association of serum omega-6 containing lipids with the fatty acid desaturase 1/2 locus and identify these metabolites as causal for CD through Mendelian randomization; and validate a novel association of serum plasmalogen and TMEM229B, which was predicted as causal for CD. An exploratory analysis combining genetics and unbiased serum metabolome surveys can reveal novel biomarkers of disease activity and potential mediators of pathology in IBD.

Calprotectin in Chronic Inflammatory Demyelinating Polyneuropathy and Variants-A Potential Novel Biomarker of Disease Activity.

Background: In chronic inflammatory demyelinating polyneuropathy (CIDP), there is an urgent need for biomarkers to monitor ongoing disease activity. Serum calprotectin (CLP) induces signaling pathways involved in inflammatory processes and has been shown to correlate with markers of disease activity in other autoimmune disorders. Thus, we wanted to study the potential value of CLP in comparison to serum neurofilament light chain (sNfl) to monitor disease activity.Materials and Methods: Sera from 63 typical and atypical CIDP and 6 MMN patients with varying degrees of disease activity were analyzed in comparison with 40 healthy controls (HC) in a cross-sectional design. Association of CLP and sNfl levels with socio-demographics, disease duration, CIDP disease activity scale (CDAS), and impairment status [medical research council-sum score (MRC-SS), the inflammatory neuropathy cause and treatment disability score (INCAT-DS), grip strength, and maximum walking distance], patient-reported outcome (PRO) parameters [SF-36 questionnaire, Beck's depression index (BDI), and fatigue severity scale (FSS)], as well as treatment regime were investigated using uni- and multivariate analysis.Results: CLP and sNfl levels were significantly higher in all CIDP patients compared to HC (p = 0.0009). Multivariate analysis adjusted for age and gender revealed that CLP acts as an independent predictor for CIDP and MMN. CLP was significantly associated with active disease course according to CDAS and correlated with MRC-SS, whereas sNfl correlated with parameters of disease impairment. There was no correlation with PRO, except for sNfl and the mental health composite score. Subgroup analysis revealed no differences between typical CIDP and atypical variants.Conclusions: CLP was elevated in CIDP and variants and was associated with active disease course, whereas sNfl shows further potential as biomarker of axonal degeneration. Thus, CLP might be a suitable additive biomarker for measurement of ongoing inflammation, which is greatly needed to guide better patient care in CIDP.

Fc Glycosylation Characterization of Human Immunoglobulins G Using Immunocapture and LC-MS.

Immunoglobulins G (IgG) are proteins produced by the immune system of higher life forms that play a central role in the defense against microbial pathogens. IgG bind pathogens with the hypervariable Fab component and mediate a diversity of effector functions by binding to immune effector cells via their crystallizable (Fc) component. All IgG Fc carry a polymorphic N-glycan that regulates its binding properties and thereby its effector functions. The glycosylation profile of IgG Fc is modulated by physiological and pathological conditions, including infectious diseases and inflammatory disorders. Characterization of IgG Fc glycosylation profiles is a promising approach to understand the pathogenesis of diseases involving the immune system and to develop novel biomarkers of disease activity. Measuring the proportion of the different IgG Fc glycoforms remains an analytical challenge, that requires a sensitive and reproducible analytical approach.This chapter describes an optimized approach for the preparation and the analysis of Fc N-glycans from total serum or plasma IgG using magnetic beads, RapiFluor MS label©, and LC-MS.

CD14 (C-159T) polymorphism is associated with increased susceptibility to SLE, and plasma levels of soluble CD14 is a novel biomarker of disease activity: A hospital-based case-control study.

Cluster of differentiation 14 (CD14) plays a crucial role in the innate immune response of the host in protection against various pathogens. The importance of soluble CD14 in autoimmune disorders has been described in different populations. However, the role of sCD14 in systemic lupus erythematosus (SLE) is poorly understood. Further, the association of functional variants at the promoter region of the CD14 gene (-159 C > T) with susceptibility to SLE or disease severity needs to be defined. Two hundred female SLE patients diagnosed on systemic lupus international collaborating clinics (SLICC) classification criteria and age, sex, matched healthy controls were enrolled in the present study. Polymerase chain reaction- restriction fragment length polymorphism (PCR-RFLP) method was used to genotype CD14 (C-159 T) polymorphism. Plasma levels of IFN-α, TNF-α, and sCD14 were quantified by enzyme-linked immunosorbent assay (ELISA). Prevalence of mutant genotypes (CT and TT) and minor allele (T) of CD14 (C-159T) polymorphism was significantly higher in SLE cases compared to healthy controls (CT: P < 0.0001; OR = 3.26, TT:P < 0.0001; OR = 3.39; T:P = 0.0009, OR = 1.62). Further, lupus nephritis patients had a higher prevalence of homozygous mutants (TT) and mutant allele (T)(TT: P = 0.0002, OR = 8.07; T: P = 0.001, OR = 1.32). SLE patients displayed significantly increased plasma sCD14, TNF-α, and IFN-α levels in comparison to healthy controls. These cytokines were significantly elevated in patients of lupus nephritis compared to those without kidney involvement. Interestingly, sCD14 levels correlated positively with SLE disease activity index-2K (SLEDAI-2K) scores and 24 hours proteinuria. CD14 (C-159T) polymorphism is associated with an increased predisposition to the development of SLE and lupus nephritis: sCD14 is a promising novel biomarker for assessing disease activity and lupus nephritis.

Milk fat globule epidermal growth factor 8 (MFG-E8) on monocytes is a novel biomarker of disease activity in systemic lupus erythematosus.

Milk fat globule epidermal growth factor (MFG-E8) is related secreted protein which links phosphatidylserine on apoptotic cells and integrin αvβ3/5 on phagocytes. To clarify the clinical significance of MFG-E8 in SLE, we analyzed the correlation between expression level of MFG-E8 in circulating phagocytic leukocytes and clinical parameters of patients. The study was conducted under a multi-center, prospective cohort design. Patients with one or both BILAG A or B, or SLEDAI- 2 K ≥ 4 with clinical symptoms were defined as the active SLE group. Expression of MFG-E8 on monocytes and concentration in serum were measured by FACS and ELISA, respectively. 96 subjects were enrolled. The absolute number and proportion of MFG-E8-positive monocytes to total monocytes were significantly higher in the active SLE group (p < 0.01). Importantly, the proportion was also significantly correlated with SLEDAI-2K, clinical SLEDAI, as well as serum levels of anti-ds-DNA antibody and complement and C1q. In addition, the proportion of MFG-E8-positive monocytes to total monocytes was significantly decreased from baseline in active SLE patients after 6 months' treatment and increased concordantly with disease activity in 6 refractory cases. Further, in receiver operating characteristic curve analysis for discrimination between active and inactive SLE, the AUC of the proportion of MFG-E8 was 0.854, which was equivalent to classical activity markers such as anti-ds DNA antibody (0.776), complement (0.897) and C1q (0.815). The proportion of MFG-E8-positive monocytes to total monocytes in peripheral blood was positively associated with disease activity in SLE and may be a novel biomarker of disease activity.

Comment on: Serum sTREM-1 in adult-onset Still's disease: a novel biomarker of disease activity and a potential predictor of the chronic course: reply.

Comment on: Serum sTREM-1 in adult-onset Still's disease: a novel biomarker of disease activity and a potential predictor of the chronic course: reply.

Comment on: Serum Strem-1 in adult-onset Still's disease: a novel biomarker of disease activity and a potential predictor of the chronic course.

Comment on: Serum Strem-1 in adult-onset Still's disease: a novel biomarker of disease activity and a potential predictor of the chronic course.

Serum level of soluble urokinase plasminogen activator receptor (suPAR) as a disease severity marker of myasthenia gravis: a pilot study.

Myasthenia gravis (MG) is an autoantibody-mediated inflammatory disease of the neuromuscular junction. Biomarkers indicating disease activity in MG are warranted. Recently, the soluble urokinase plasminogen activator receptor (suPAR) has been reported to be associated with inflammation, tissue damage, disease activity and prognosis in various diseases, including autoimmune diseases. In this study, serum suPAR levels were measured in 40patients with anti-acetylcholine receptor antibody-positive MG and 30controls, and their correlations with clinical variables and severity scale scores were investigated. We identified that serum suPAR levels significantly correlated with MG activities of daily living scale (Spearman's ρ=0·45; P=0·004) and MG Foundation of America classification (Spearman's ρ=0·37; P=0·02) at serum sampling, but not with anti-acetylcholine receptor antibody titers. In conclusion, serum suPAR levels can be a candidate for a novel biomarker of disease activity in anti-acetylcholine receptor antibody-positive MG.

Neutrophil-derived lipocalin-2 in adult-onset Still's disease: a novel biomarker of disease activity and liver damage.

ObjectiveLiver damage is a common manifestation and can be life-threatening in adult-onset Still’s disease (AOSD), an autoinflammatory disease. The hallmark of AOSD is activation of neutrophils, whose infiltration in liver is suspected to promote tissue injury. Here we aimed to identify a candidate biomarker and to validate its association with liver damage in AOSD.MethodsTranscriptome analysis of neutrophils from treatment-naïve active AOSD patients and healthy donors was performed. Lipocalin-2 (LCN2) expression was assessed in neutrophils, plasma and liver biopsies of AOSD. The correlations of LCN2 with different variables and its ability to identify liver damage from AOSD patients were analysed.ResultsLCN2, a novel biomarker in hepatic inflammation, was found to be upregulated in AOSD neutrophils by RNA sequencing and confirmed at the mRNA and protein levels. Plasma levels of LCN2 were significantly higher in AOSD patients than healthy controls, RA and SLE patients. Plasma LCN2 levels were closely correlated with inflammatory markers, systemic score, HScore and cytokines. Moreover, LCN2 levels were increased in active AOSD with liver involvement and independently associated with liver dysfunction. Enhanced expression of LCN2 was detected in liver biopsies from three patients with ongoing liver injury. Furthermore, the area under the curve value of LCN2 for identifying AOSD with liver injury from other liver diseases was 0.9694.ConclusionOur results reveal that neutrophils-derived LCN2 is higher in plasma and liver tissue in AOSD patients than in healthy controls, and it could serve as a potent biomarker for identifying AOSD with systemic inflammation, especially liver damage caused by hyperinflammation.

Quantitative Proteomic Profile of Psoriatic Epidermis Identifies OAS2 as a Novel Biomarker for Disease Activity.

Psoriasis is a common chronic inflammatory systemic disease. Epidermal proteins are considered to be important in maintaining skin barrier function, innate immunity, and inflammation. To define more possible roles of the epidermis in the pathogenesis of psoriasis, quantified proteomic analysis was used to screen and analyze the differentially expressed epidermal proteins between 16 psoriasis patients and 15 healthy controls. Upregulated differential expression proteins (DEPs) include several significant functional protein clusters, including antimicrobial peptides (AMPs) and antiviral proteins (AVPs). The levels of 2–5-oligoadenylate synthase 2 (OAS2) in both epidermis and serum levels were significantly elevated in psoriasis and were also positively correlated with Psoriasis Area Severity Index (PASI) scores and Body Surface Area (BSA) scores. Moreover, OAS2 expression in psoriatic skin significantly decreased after IL-17R mono-antibody treatment. It has been clarified that inflamed keratinocytes were the main source of abnormally increased OAS2 in psoriasis skin by immunofluorescence and primary cell cultures. Keratinocyte-derived OAS2 can be induced by not only IFNβ, but also psoriasis associated cytokines like IL-17A and IL-6. This study revealed that AMPs and AVPs are two important functional protein clusters altering innate immune in psoriatic epidermis. OAS2 is a novel potential sensitive biomarker, which could predict the severity and activity of psoriasis, and could also be used as an indicator to evaluate or monitor the efficacy of clinical treatment.

Serum sTREM-1 in adult-onset Still's disease: a novel biomarker of disease activity and a potential predictor of the chronic course.

ObjectivesTriggering receptor expressed on myeloid cells-1 (TREM-1) is an amplifier of inflammatory signals. Recently, a soluble form of TREM-1 (sTREM-1) was described. This study aimed to investigate the role of serum sTREM-1 in patients with adult-onset Still’s disease (AOSD).MethodsSerum sTREM-1 levels were detected in 108 AOSD patients, 88 RA patients and 112 healthy controls (HC). The correlations of sTREM-1 with disease activity, clinical characteristics and laboratory parameters in AOSD patients were analysed by the Spearman correlation test. Risk factors for the chronic course of AOSD were evaluated by multivariate logistic regression analysis.ResultsAOSD patients had significantly higher serum sTREM-1 levels than RA patients and HC, and serum sTREM-1 levels were correlated with the systemic score, ferritin, leucocyte count, CRP, IL-1β and IL-6. The elevation in the initial sTREM-1 level by itself could discriminate patients developing the chronic course from patients developing the nonchronic course. Moreover, an elevated sTREM-1 level (> 526.4475 pg/ml) was an independent risk factor for the chronic course in active AOSD patients. Furthermore, interfering with TREM-1 engagement led to reductions in the secretion of pro-inflammatory cytokines, such as IL-1β, IL-6 and TNF-α, in neutrophils and monocytes from active AOSD patients.ConclusionSerum sTREM-1 levels are correlated with disease activity, and an elevation in the initial serum sTREM-1 level is a potential predictor of the chronic course in AOSD patients, which currently provides the best predictive model for identifying patients prone to developing the chronic course of AOSD.

Evaluation of Potential Serum Biomarkers of Disease Activity in Diverse Forms of Vasculitis.

We evaluated potential circulating biomarkers of disease activity in giant cell arteritis (GCA), Takayasu arteritis (TA), polyarteritis nodosa (PAN), and eosinophilic granulomatosis with polyangiitis (EGPA). A panel of 22 serum proteins was tested in patients enrolled in the Vasculitis Clinical Research Consortium Longitudinal Studies of GCA, TA, PAN, or EGPA. Mixed models were used for most analyses. A J48 classification tree method was used to find the most relevant markers to differentiate between active and inactive GCA. Tests were done on 418 samples from 152 patients (60 GCA, 29 TA, 26 PAN, 37 EGPA), during both active vasculitis and remission. In GCA, these showed significant (p < 0.05) differences between disease states: B cell-attracting chemokine 1 (BCA)-1/CXC motif ligand 13 (CXCL13), erythrocyte sedimentation rate (ESR), interferon-γ-induced protein 10/CXC motif chemokine 10, soluble interleukin 2 receptor α (sIL-2Rα), and tissue inhibitor of metalloproteinase-1 (TIMP-1). In EGPA, these showed significant increases during active disease: granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage-CSF, interleukin (IL)-6, IL-15, and sIL-2Rα. BCA-1/CXCL13 also showed such increases, but only after adjustment for treatment. In PAN, ESR and matrix metalloprotease (MMP)-3 showed significant differences between disease states. Differences in biomarker levels between diseases were significant for 11 markers and were more striking (all p < 0.01) than differences related to disease activity. A combination of lower values of TIMP-1, IL-6, interferon-γ, and MMP-3 correctly classified 87% of samples with inactive GCA. We identified novel biomarkers of disease activity in GCA and EGPA. Differences of biomarker levels between diseases, independent of disease activity, were more apparent than differences related to disease activity. Further studies are needed to determine whether these serum proteins have potential for clinical use in distinguishing active disease from remission or in predicting longer-term outcomes.

Circular RNAS: novel biomarkers of disease activity in systemic lupus erythematosus?

Circular RNAs (circRNAs) are a class of non-coding RNAs that regulate gene expression by acting as competitive endogenous RNAs (ceRNAs) and modulating gene transcription. Several studies support the implication of circRNAs in a variety of human diseases, but research on the role of circRNAs in systemic lupus erythematosus (SLE) is lacking. In a study recently published in Clinical Science (2018), Zhang et al. identified hsa_circ_0012919 as a potential biomarker of disease activity in SLE patients. The authors observed different circRNA expression between SLE patients and healthy controls, an association with clinical variables and with the abnormal DNA methylation present in SLE CD4+ T cells. Finally, Zhang et al. demonstrated that hsa_circ_0012919 acts as a miRNA sponge for miR-125a-3p, regulating the gene expression of targets RANTES and KLF13 that are involved in the physiology and pathophysiology of acute and chronic inflammatory processes. These findings support the role of circRNAs in the pathophysiology of SLE.