Search

Objective: To discover antibody biomarkers that can predict a lack of response to first-line therapy in rheumatoid arthritis (RA) patients., Methods: Two RA cDNA phage display libraries were screened for novel antibodies in baseline RA sera from the Care in early RA (CareRA) trial, differentiating between patients who did or did not reach remission after first-line therapy (n=20 each). Antibody reactivity to identified University Hasselt (UH)-RA antigens was validated in baseline samples from 136 additional CareRA participants. The novel antibodies' potential to predict failure to reach remission or low disease activity (LDA), according to the Disease Activity Score 28-joint C-reactive protein/erythrocyte sedimentation rate (DAS28CRP/ESR) and Clinical/Simplified Disease Activity Index (CDAI/SDAI), was studied by multivariate analyses. The presence of the antibody targets in RA synovial tissue and the fibroblast-like synoviocyte (FLS) cell line SW982 was determined by immunofluorescence., Results: We identified antibodies to 41 novel antigens. Antibodies against any of three antigens, UH-RA.305/318/329, discriminated between RA patients not reaching week (w)8 DAS28CRP remission and those that did (36% vs 13%,p=0.0031). In all patients, anti-UH-RA.305/318/329 antibody reactivity was associated with failure to reach week 8 DAS28CRP and DAS28ESR remission (OR 3.63,p=0.0031; OR 2.92,p=0.016; respectively), SDAI/CDAI sustained remission (OR 5.59,p=0.039 for both) and DAS28CRP and DAS28ESR sustained LDA (OR 3.7,p=0.009; OR 2.76,p=0.042; respectively). In rheumatoid factor/anti-citrullinated protein antibody (RF/ACPA) seronegative patients, these antibodies were strongly associated with failure to achieve week 8 DAS28CRP remission (OR 17.3,p=0.0029). Anti-UH-RA.305/329 antibodies were shown to target FLS in RA synovial tissue and SW982 cells., Conclusion: We identified three antibody biomarkers that are associated with failure to achieve remission/LDA after first-line RA therapy., Competing Interests: Competing interests: PVandormael, VS and PVerschueren have a patent application filed on the biomarkers described in this report., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

The water channel aquaporin 4 (AQP4) contributes to water flow and waste removal across the blood-brain barrier and its levels, organization and localization are perturbed in various neurological diseases, including Alzheimer's Disease. This renders AQP4 a potentially valuable therapeutic target. However, most functional assays aimed at identifying modulators of AQP4 function are performed with primary rodent cells and do not consider inter-cellular variations in AQP4 abundance and presentation. To address this, we have established and applied a robust live cell microscopy assay that captures the contribution of AQP4 in the osmotically driven (de-)quenching of the vital dye Calcein-AM with single-cell resolution. Using human astrocytoma cells, we found that performing measurements on cellular regions instead of whole fields of view yielded a more sensitive readout of the osmotic response, which correlated with AQP4 abundance. Stable co-expression of the two major AQP4 isoforms, but not of the individual isoforms, provoked a faster adaptation to osmotic changes, while siRNA-mediated knockdown of AQP4 had the opposite effect. Post-hoc correlation with the canonical membrane marker CD44 revealed that the speed of the osmotic response scaled with AQP4 membrane enrichment. Coating the substrate with laminin promoted AQP4 membrane enrichment, while cell confinement with fixed-size micropatterns further increased the speed of osmoregulation, underscoring the influence of extracellular factors. The osmotic response of primary fetal astrocytes and human iPSC-derived astrocyte models was comparable to AQP4-deficient astrocytoma cells, in line with their low AQP4 levels and indicative of their immature state. In conclusion, a correlative single-cell approach based on the quantification of Calcein-AM quenching capacity, AQP4 abundance and AQP4 membrane enrichment, allows resolving osmoregulation in a more sensitive manner and reveals heterogeneity between and within human astrocyte (-like) cultures, which could prove crucial for future screens aimed at identifying AQP4 modulators., (© 2024 International Society for Advancement of Cytometry.)

Traumatic spinal cord injury (SCI) is a severe condition leading to long-term impairment of motor, sensory, and autonomic functions. Following the initial injury, a series of additional events is initiated further damaging the spinal cord. During this secondary injury phase, both an inflammatory and immune modulatory response are triggered that have damaging and anti-inflammatory properties, respectively. The proinflammatory cytokine macrophage migration inhibitory factor (MIF) and its receptor CD74 have been extensively studied in traumatic SCI. MIF expression is increased in spinal cord tissue after experimental SCI, mainly in astrocytes and microglia, as well as in the plasma of SCI patients. Functionally, MIF and CD74 were shown to regulate astrocyte viability, proliferation and cholesterol metabolism, microglia migration, and neuronal viability. Moreover, inhibition of the MIF/CD74 axis improved the functional recovery of SCI animals. We provide a detailed overview of studies analyzing the role of MIF and CD74 in traumatic SCI. We describe results from animal studies, using rat and mouse models for SCI, and human studies. Furthermore, we propose a new path for investigation, focused on B cells, that might lead to a better understanding of how MIF and CD74 contribute to the secondary injury cascade following traumatic SCI., (© 2024 Wiley‐VCH GmbH.)

Purpose: Cardiovascular health disparities are present in African Americans, but it remains unknown whether this phenomenon affect Somali Americans. Study of Somali Americans is warranted due to distinct genetic and cultural differences from African Americans of western African ancestry. Orthostatic hemodynamic responses have implications for cardiovascular risk, especially among African American females. We sought to examine race and sex differences in systolic (SAP) and diastolic (DAP) arterial pressure and heart rate (HR) responsiveness to standing. We hypothesized that SAP, DAP, and HR change from supine to standing position would be higher in Somali Americans., Methods: We studied blood pressure and HR responsiveness in 139 (70 Somali; age: 29±10 years, 69 White; age: 31±9 years) participants. Supine SAP, DAP, and HR were measured after at least five minutes of supine rest, and again after one minute of standing. SAP, DAP, and HR change was compared between groups., Results: ΔSAP and ΔDAP were similar between groups (race × sex: p>0.05). However, HR responsiveness to orthostasis varied between race and sex comparisons (race×sex: p=0.011). Somali females exhibited an augmented HR response to orthostasis compared to White females (Δ19±13 vs. 11±9 beats/min, p=0.005) and Somali males (Δ19±13 vs. 12±9 beats/min, p=0.020)., Conclusion: ΔHR to standing is augmented in young female Somali Americans. These findings highlight an early potential impairment in hemodynamic regulation that may heighten future cardiovascular risk. Further work is warranted to identify the potential autonomic nervous system underpinnings that may contribute to potentiated orthostatic responses and cardiovascular risk in Somali American females., Clinical Trial Registration: www.clinicaltrials.gov; unique identifier, NCT04124848; NCT05411029; NCT03308578., Competing Interests: Declarations VKS has consulted for Lilly, Zoll, Jazz Pharma, Axsome, and Know Labs and is on the Sleep Number Scientific Advisory Board. All other authors have no relevant conflicts to disclose.

In autoimmunity, FOXP3+ Tregs skew toward a proinflammatory, nonsuppressive phenotype and are, therefore, unable to control the exaggerated autoimmune response. This largely affects the success of autologous Treg therapy, which is currently under investigation for autoimmune diseases, including multiple sclerosis (MS). There is a need to ensure in vivo Treg stability before successful application of Treg therapy. Using genetic fate-mapping mice, we demonstrate that inflammatory, cytokine-expressing exFOXP3 T cells accumulate in the CNS during experimental autoimmune encephalomyelitis. In a human in vitro model, we discovered that interaction with inflamed blood-brain barrier endothelial cells (BBB-ECs) induces loss of function by Tregs. Transcriptome and cytokine analysis revealed that in vitro migrated Tregs have disrupted regenerative potential and a proinflammatory Th1/17 signature, and they upregulate the mTORC1 signaling pathway. In vitro treatment of migrated human Tregs with the clinically approved mTORC1 inhibitor rapamycin restored suppression. Finally, flow cytometric analysis indicated an enrichment of inflammatory, less-suppressive CD49d+ Tregs in the cerebrospinal fluid of people with MS. In summary, interaction with BBB-ECs is sufficient to affect Treg function, and transmigration triggers an additive proinflammatory phenotype switch. These insights help improve the efficacy of autologous Treg therapy of MS.

One of the major challenges in multiple sclerosis (MS) is to accurately monitor and quantify disability over time. Thus, there is a pressing need to identify new biomarkers for disease progression. Peripheral blood DNA methylation has been demonstrated to be an easily accessible and quantifiable marker in many neurodegenerative diseases. In this study, we aimed to investigate whether methylation patterns that were previously determined in chronic inactive white matter lesions of patients with progressive MS are also reflected in the blood, and whether the latter can serve as a biomarker for disease progression in MS. While our initial analysis revealed differences in the blood methylation state of important myelin-related genes between patients with progressive MS and controls, these findings could not be validated in other independent patient cohorts. Subsequent investigation suggests that sample storage can selectively influence DNA methylation patterns, potentially hindering accurate epigenetic analysis. Therefore, sample storage time should be taken into consideration during the initial sample selection stage in biomarker studies.

B cell depletion by the anti-CD20 antibody ocrelizumab is effective in relapsing-remitting (RR) and primary progressive (PP) multiple sclerosis (MS). We investigated immunological changes in peripheral blood of a real-world MS cohort after 6 and 12 months of ocrelizumab. All RRMS and most PPMS patients (15/20) showed treatment response. Ocrelizumab not only reduced CD20 + B cells, but also numbers of CD20 + T cells. Absolute numbers of monocytes, dendritic cells and CD8 + T cells were increased, while CD56 hi natural killer cells were reduced after ocrelizumab. The residual B cell population shifted towards transitional and activated, IgA + switched memory B cells, double negative B cells, and antibody-secreting cells. Delaying the treatment interval by 2-3 months increased mean B cell frequencies and enhanced naive B cell repopulation. Ocrelizumab reduced plasma levels of interleukin(IL)-12p70 and interferon(IFN)-α2. These findings will contribute to understanding ineffective treatment responses, dealing with life-threatening infections and further unravelling MS pathogenesis., Competing Interests: Declaration of competing interest All authors have no conflict of interest to declare relevant to this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The immune response in patients with Coronavirus Disease 2019 (COVID-19) is highly variable and is linked to disease severity and mortality. However, antibody and cytokine responses in the early disease stage and their association with disease course and outcome are still not completely understood. In this large, multi-centre cohort study, blood samples of 434 Belgian COVID-19 hospitalized patients with different disease severities (ranging from asymptomatic/mild to critically ill) from the first wave of the COVID-19 pandemic were obtained. Baseline antibody and cytokine responses were characterized and associations with several clinical outcome parameters were determined. Anti-spike immunoglobulin (Ig)G and IgM levels were elevated in patients with a more severe disease course. This increased baseline antibody response however was associated with decreased odds for hospital mortality. Levels of the pro-inflammatory cytokines IL-6, IP-10 and IL-8, the anti-inflammatory cytokine IL-10 and the antiviral cytokines IFN-α, IFN-β and IFN-λ1 were increased with disease severity. Remarkably, we found significantly lower levels of IFN-λ2,3 in critically ill patients compared to patients of the moderate and severe disease category. Finally, levels of IL-8, IL-6, IP-10, IL-10, IFN-α, IFN-β, IFN-γ and IFN-λ1 at baseline were positively associated with mortality, whereas higher IFN-λ2,3 levels were negatively associated with mortality., (© 2023. The Author(s).)

Acute decompensated aortic stenosis (ADAS) is common. The cumulative burden of ADAS from a clinical, health care resource, and financial perspective is unknown. This study sought to assess the national impact of ADAS compared with electively treated, stable patients with aortic stenosis (non-ADAS). Using the National Readmissions Database between 2016 and 2019, patients with ADAS and non-ADAS were identified using International Classification of Diseases, Tenth Revision codes. Patients with ADAS were propensity-matched to non-ADAS patients (1:2) using age, gender, and Charlson co-morbidity index. We compared in-hospital mortality, length of stay (LOS), health care-associated costs, and 90-day readmission data between the 2 cohorts. A total of 51,498 propensity-matched patients were included in this study: median age 75 years, 64% men. The in-hospital mortality for ADAS was higher than non-ADAS (2.8% vs 1.5%, p <0.0001). The LOS during the index admission was longer for ADAS (9 [5 to 13] vs 4 [2 to 6] days, p <0.0001). The health care-associated costs per patient was greater for ADAS ($55,450.0 [41,860.4 to 74,500.7] vs $43,405.7 [34,218.5 to 56,034.8], p <0.0001). Readmission to hospital within 90 days was more frequent in ADAS (21.1 vs 16.8%, p <0.001). The in-hospital mortality during readmission was higher with ADAS (3.9% vs 2.8%, p = 0.004). The readmission LOS was longer with ADAS (4 [2 to 7] vs 3 [2 to 6] days, p <0.0001). In conclusion, ADAS imposes a significant burden clinically and financially and on health care resources compared with non-ADAS during the index admission and 90-day follow-up. There is an urgent need to predict ADAS and optimize the timing of aortic valve replacement to reduce the incidence and the burden associated with ADAS., Competing Interests: Declaration of Competing Interest Drs. Patel and Mullen have an unrestricted grant from Edwards Lifesciences. The remaining authors have no competing interests to declare., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Fatty liver disease in the absence of excessive alcohol consumption is an increasingly common condition with a global prevalence of ~ 25-30% and is also associated with cardiovascular disease (CVD). Since systemic metabolic dysfunction underlies its pathogenesis, the term metabolic (dysfunction)-associated fatty liver disease (MAFLD) has been proposed for this condition. MAFLD is closely intertwined with obesity, type 2 diabetes mellitus and atherogenic dyslipidemia, which are established cardiovascular risk factors. Unlike CVD, which has received attention in the literature on fatty liver disease, the CVD risk associated with MAFLD is often underestimated, especially among Cardiologists., Methods and Results: A multidisciplinary panel of fifty-two international experts comprising Hepatologists, Endocrinologists, Diabetologists, Cardiologists and Family Physicians from six continents (Asia, Europe, North America, South America, Africa and Oceania) participated in a formal Delphi survey and developed consensus statements on the association between MAFLD and the risk of CVD. Statements were developed on different aspects of CVD risk, ranging from epidemiology to mechanisms, screening, and management., Conculsions: The expert panel identified important clinical associations between MAFLD and the risk of CVD that could serve to increase awareness of the adverse metabolic and cardiovascular outcomes of MAFLD. Finally, the expert panel also suggests potential areas for future research., (© 2023. Asian Pacific Association for the Study of the Liver.)

Human B cells can be divided into four main subsets based on differential expression of immunoglobulin (Ig)D and CD27. IgD - CD27 - double negative (DN) B cells make up a heterogeneous group of B cells that have first been described in relation to aging and systemic lupus erythematosus but have been mostly disregarded in B cell research. Over the last few years, DN B cells have gained a lot of interest because of their involvement in autoimmune and infectious diseases. DN B cells can be divided into different subsets that originate via different developmental processes and have different functional properties. Further research into the origin and function of different DN subsets is needed to better understand the role of these B cells in normal immune responses and how they could be targeted in specific pathologies. In this review, we give an overview of both phenotypic and functional properties of DN B cells and provide insight into the currently proposed origins of DN B cells. Moreover, their involvement in normal aging and different pathologies is discussed., Competing Interests: Declaration of Competing Interest The authors have no conflict of interest to declare relevant to this article., (Copyright © 2023 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.)

Introduction: There is an unmet need for biomarkers to identify patients with axial spondyloarthritis (axSpA). Increasing evidence suggest the presence of autoantibodies in a subset of axSpA patients. The aim of this study was to identify novel IgA antibodies in early axSpA patients and to determine their diagnostic potential in combination with previously determined IgG antibodies against UH (Hasselt University)-axSpA-IgG antigens., Methods: An axSpA cDNA phage display library constructed from axSpA hip synovium, was used to screen for novel IgA antibodies in plasma from early axSpA patients. The presence of these antibodies against novel UH-axSpA-IgA antigens was determined in two independent axSpA cohorts, in healthy controls and in patients with chronic low back pain., Results: We identified antibodies to 7 novel UH-axSpA-IgA antigens, of which 6 correspond to non-physiological peptides and 1 to the human histone deacetylase 3 (HDAC3) protein. IgA antibodies against 2 of these 7 novel UH-axSpA-IgA antigens and IgG antibodies against 2 of the previously identified antigens were significantly more present in early axSpA patients from the UH cohort (18/70, 25.7%) and the (Bio)SPAR cohort (26/164, 15.9%), compared to controls with chronic low back pain (2/66, 3%). Antibodies to this panel of 4 antigens were present in 21.1% (30/142) of patients with early axSpA from the UH and (Bio)SPAR cohorts. The positive likelihood ratio for confirming early axSpA using antibodies to these 4 UH-axSpA antigens was 7.0. So far, no clinical correlation between the novel identified IgA antibodies and inflammatory bowel disease could be identified., Discussion: In conclusion, screening an axSpA cDNA phage display library for IgA reactivity resulted in the identification of 7 novel UH-axSpA-IgA antigens, of which 2 show promising biomarker potential for the diagnosis of a subset of axSpA patients, in combination with previously identified UH-axSpA-IgG antigens., Competing Interests: VS, PV, KV, and DQ have a patent application pending on the markers described in this report. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Ruytinx, Vandormael, Quaden, Luyten, Geusens, Vanhoof, Agten, Vandenabeele, de Vlam and Somers.)

Introduction: Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder for which early recognition is a major challenge. Autoantibodies against fetal brain antigens have been found in the blood of mothers of children with ASD (m-ASD) and can be transferred to the fetus where they can impact neurodevelopment by binding to fetal brain proteins. This study aims to identify novel maternal autoantibodies reactive against human fetal brain antigens, and explore their use as biomarkers for ASD screening and diagnosis., Methods: A custom-made human fetal brain cDNA phage display library was constructed, and screened for antibody reactivity in m-ASD samples from the Simons Simplex Collection (SSC) of the Simons Foundation Autism Research Initiative (SFARI). Antibody reactivity against 6 identified antigens was determined in plasma samples of 238 m-ASD and 90 mothers with typically developing children (m-TD)., Results: We identified antibodies to 6 novel University Hasselt (UH)-ASD antigens, including three novel m-ASD autoantigens, i.e., ribosomal protein L23 (RPL23), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and calmodulin-regulated spectrin-associated protein 3 (CAMSAP3). Antibody reactivity against a panel of four of these targets was found in 16% of m-ASD samples, compared to 4% in m-TD samples ( p = 0.0049)., Discussion: Maternal antibodies against 4 UH-ASD antigens could therefore provide a novel tool to support the diagnosis of ASD in a subset of individuals., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Mazón-Cabrera, Liesenborgs, Brône, Vandormael and Somers.)

Excessive daytime sleepiness (EDS) is classically viewed as a consequence of insufficient sleep or a symptom of sleep disorders. Epidemiological and clinical evidence have shown that patients reporting EDS in tandem with sleep disorders (e.g., obstructive sleep apnoea) are at greater cardiovascular risk than non-sleepy patients. While this may simply be attributable to EDS being present in patients with a more severe condition, treatment of sleep disorders does not consistently alleviate EDS, indicating potential aetiological differences. Moreover, not all patients with sleep disorders report EDS, and daytime sleepiness may be present even in the absence of any identifiable sleep disorder; thus, EDS could represent an independent pathophysiology. The purpose of this review is twofold: first, to highlight evidence that EDS increases cardiovascular risk in the presence of sleep disorders such as obstructive sleep apnoea, narcolepsy and idiopathic hypersomnia and second, to propose the notion that EDS may also increase cardiovascular risk in the absence of known sleep disorders, as supported by some epidemiological and observational data. We further highlight preliminary evidence suggesting systemic inflammation, which could be attributable to dysfunction of the gut microbiome and adipose tissue, as well as deleterious epigenetic changes, may promote EDS while also increasing cardiovascular risk; however, these pathways may be reciprocal and/or circumstantial. Additionally, gaps within the literature are noted followed by directions for future research., Competing Interests: Competing interests: VS has consulted for Baker Tilly, Respicardia, Bayer and Jazz Pharmaceuticals, and is on the Scientific Advisory Board for Sleep Number Corporation., (© Author(s) (or their employer(s)) 2022. No commercial re-use. See rights and permissions. Published by BMJ.)

The WHO Health Emergencies Programme combined online tools with adult learning techniques to develop an innovative online leadership course. A combination of self-paced learning and online classes were used to deliver skills-based leadership training tailored to public health staff working in emergency response roles. Although using an online approach was considered a temporary solution to counter travel restrictions during the pandemic, the advantages have challenged preconceptions that effective learning, networking and peer exchange for leadership can only be achieved through face-to-face learning.

Following a spinal cord injury (SCI), an inflammatory immune reaction is triggered which results in advanced secondary tissue damage. The systemic post-SCI immune response is poorly understood. This study aimed to extensively analyse the circulating immune cell composition in traumatic SCI patients in relation to clinical parameters. High-dimensional flow cytometry was performed on peripheral blood mononuclear cells of 18 traumatic SCI patients and 18 healthy controls to determine immune cell subsets. SCI blood samples were collected at multiple time points in the (sub)acute (0 days to 3 weeks post-SCI, (s)aSCI) and chronic (6 to >18 weeks post-SCI, cSCI) disease phase. Total and CD4 + T cell frequencies were increased in cSCI patients. Both CD4 + T cells and B cells were shifted towards memory phenotypes in (s)aSCI patients and cSCI patients, respectively. Most profound changes were observed in the B cell compartment. Decreased immunoglobulin (Ig)G + and increased IgM + B cell frequencies reflected disease severity, as these correlated with American Spinal Injury Association (ASIA) impairment scale (AIS) scores. Post-SCI B cell responses consisted of an increased frequency of CD74 + cells and CD74 expression level within total B cells and B cell subsets. Findings from this study suggest that post-SCI inflammation is driven by memory immune cell subsets. The increased CD74 expression on post-SCI B cells could suggest the involvement of CD74-related pathways in neuroinflammation following SCI. In addition, the clinical and prognostic value of monitoring circulating IgM + and IgG + B cell levels in SCI patients should be further evaluated., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Fraussen, Beckers, van Laake-Geelen, Depreitere, Deckers, Cornips, Peuskens and Somers.)

Background: The contribution of native or modified oligodendroglia-derived extracellular vesicles (OL-EVs) in controlling chronic inflammation is poorly understood. In activated microglia, OL-EVs contribute to the removal of cytotoxic proteins following a proteotoxic stress. Intracellular small heat shock protein B8 (HSPB8) sustain this function by facilitating autophagy and protecting cells against oxidative stress mediated cell death. Therefore, secretion of HSPB8 in OL-EVs could be beneficial for neurons during chronic inflammation. However, how secreted HSPB8 contribute to cellular proteostasis remains to be elucidated., Methods: We produced oligodendroglia-derived EVs, either native (OL-EVs) or HSPB8 modified (OL-HSPB8-EVs), to investigate their effects in controlling chronic inflammation and cellular homeostasis. We analyzed the impact of both EV subsets on either a resting or activated microglial cell line and on primary mixed neural cell culture cells. Cells were activated by stimulating with either tumor necrosis factor-alpha and interleukin 1-beta or with phorbol-12-myristate-13-acetate., Results: We show that OL-EVs and modified OL-HSPB8-EVs are internalized by C20 microglia and by primary mixed neural cells. The cellular uptake of OL-HSPB8-EVs increases the endogenous HSPB8 mRNA expression. Consistently, our results revealed that both EV subsets maintained cellular homeostasis during chronic inflammation with an increase in the formation of autophagic vesicles. Both EV subsets conveyed LC3B-II and BAG3 autophagy markers with an enhanced effect observed for OL-HSPB8-EVs. Moreover, stimulation with either native or modified OL-HSPB8-EVs showed a significant reduction in ubiquitinated protein, reactive oxygen species and mitochondrial depolarization, with OL-HSPB8-EVs exhibiting a more protective effect. Both EV subsets did not induce cell death in the C20 microglia cell line or the primary mixed neural cultures., Conclusion: We demonstrate that the functions of oligodendroglia secreted EVs enriched with HSPB8 have a supportive role, comparable to the native OL-EVs. Further development of engineered oligodendroglia derived EVs could be a novel therapeutic strategy in countering chronic inflammation. Video Abstract., (© 2022. The Author(s).)

Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

The WHO defines different COVID-19 disease stages in which the pathophysiological mechanisms differ. We evaluated the characteristics of these COVID-19 disease stages. Forty-four PCR-confirmed COVID-19 patients were included in a prospective minimal invasive autopsy cohort. Patients were classified into mild-moderate (n = 4), severe-critical (n = 32) and post-acute disease (n = 8) and clinical, radiological, histological, microbiological and immunological data were compared. Classified according to Thoracic Society of America, patients with mild-moderate disease had no typical COVID-19 images on CT-Thorax versus 71.9% with typical images in severe-critical disease and 87.5% in post-acute disease (P < 0.001). Diffuse alveolar damage was absent in mild-moderate disease but present in 93.8% and 87.5% of patients with severe-critical and post-acute COVID-19 respectively (P = 0.002). Other organs with COVID-19 related histopathological changes were liver and heart. Interferon-γ levels were significantly higher in patients with severe-critical COVID-19 (P = 0.046). Anti-SARS CoV-2 IgG was positive in 66%, 40.6% and 87.5% of patients with mild-moderate, severe-critical and post-acute COVID-19 respectively (n.s.). Significant differences in histopathological and immunological characteristics between patients with mild-moderate disease compared to patients with severe-critical disease were found, whereas differences between patients with severe-critical disease and post-acute disease were limited. This emphasizes the need for tailored treatment of COVID-19 patients., (© 2022. The Author(s).)

The differentiation of oligodendrocyte precursor cells (OPCs) into myelinating oligodendrocytes is the prerequisite for remyelination in demyelinated disorders such as multiple sclerosis (MS). Epigenetic mechanisms, such as DNA methylation, have been suggested to control the intricate network of transcription factors involved in OPC differentiation. Yet, the exact mechanism remains undisclosed. Here, we are the first to identify the DNA-binding protein inhibitors, Id2 and Id4, as targets of DNA methylation during OPC differentiation. Using state-of-the-art epigenetic editing via CRISPR/dCas9-DNMT3a, we confirm that targeted methylation of Id2/Id4 drives OPC differentiation. Moreover, we show that in the pathological context of MS, methylation and gene expression levels of both ID2 and ID4 are altered compared to control human brain samples. We conclude that DNA methylation is crucial to suppress ID2 and ID4 during OPC differentiation, a process that appears to be dysregulated during MS. Our data do not only reveal new insights into oligodendrocyte biology, but could also lead to a better understanding of CNS myelin disorders., (© 2021. The Author(s).)

The study of cell proliferation is of great importance for medical and biological research, as well as for industrial applications. To render the proliferation process accurately over time, real-time cell proliferation assay methods are required. This work presents a novel real-time and label-free approach for monitoring cell proliferation by continuously measuring changes in thermal properties that occur at the sensor interface during the process. The sensor consists of a single planar resistive structure deposited on a thin foil substrate, integrated at the bottom of a cell culture reservoir. During measurement, the structure is excited with square wave current pulses. Meanwhile, the temperature-induced voltage change measured over the structure is used to derive variations in the number of cells at the interface. This principle is demonstrated first by performing cell sedimentation measurements to quantify the presence of cells at the sensor interface in the absence of cell growth. Later, cell proliferation experiments were performed, whereby parameters such as the available nutrient content and the cell starting concentration were modified. Results from these experiments show that the thermal-based sensor is able to accurately measure variations in the number of cells at the interface. Moreover, the influence of the modified parameters could be observed in the obtained proliferation curves. These findings highlight the potential for the presented thermal method to be incorporated in a standardized well plate format for high-throughput monitoring of cell proliferation.

Objective: This study was undertaken to identify novel autoantibodies in axial spondyloarthritis (SpA) and determine their diagnostic potential in patients with early axial SpA and controls from 2 independent cohorts., Methods: An axial SpA complementary DNA phage display library was used to screen for novel IgG antibodies in plasma from patients with early axial SpA. The presence of these antibodies against novel peptides (i.e., peptides identified in an early axial SpA cohort from Hasselt University, designated UH-axSpA) was determined by enzyme-linked immunosorbent assay in 76 patients with early axial SpA, 75 controls with nonspecific chronic low back pain, 60 patients with rheumatoid arthritis, and 94 healthy controls from the UH cohort. Antibody reactivity to these novel peptides was further validated in 174 patients with axial SpA (of whom 79 had early axial SpA) from the University Hospitals Leuven (Bio)SPAR (Spondyloarthritis [Biologics]) cohort., Results: We identified antibodies to 9 novel UH-axSpA peptides, corresponding to randomly formed peptides and to a novel axial SpA autoantigen, double homeobox protein 4. Antibodies to 3 UH-axSpA peptides with the highest positive likelihood ratio (LR) for a diagnosis of axial SpA were present in significantly more patients with early axial SpA from the UH and (Bio)SPAR cohorts (14.2% [22/155]) compared to controls with chronic low back pain (5% [4/75]), resulting in 95% specificity. The positive LR for confirming axial SpA using antibodies to these 3 UH-axSpA peptides was 2.7, which is higher than the LR obtained with the currently used laboratory marker C-reactive protein. Testing for antibodies to these 3 UH-axSpA peptides in patients with chronic low back pain increased the posttest probability of a diagnosis of axial SpA from 79% to 91%., Conclusion: Antibodies to 3 UH-axSpA peptides could provide a novel tool in the diagnosis of a subset of axial SpA patients., (© 2020, American College of Rheumatology.)

Microglia, the immunocompetent cells of the central nervous system (CNS), play an important role in maintaining cellular homeostasis in the CNS. These cells secrete immunomodulatory factors including nanovesicles and participate in the removal of cellular debris by phagocytosis or autophagy. Accumulating evidence indicates that specifically the cellular exchange of small extracellular vesicles (EVs), participates in physiology and disease through intercellular communication. However, the contribution of microglial-derived extracellular vesicles (M-EVs) to the maintenance of microglia homeostasis and how M-EVs could influence the phenotype and gene function of other microglia subtypes is unclear. In addition, knowledge of canonical signalling pathways of inflammation and immunity gene expression patterns in human microglia exposed to M-EVs is limited. Here, we analysed the effects of M-EVs produced in vitro by either tumour necrosis factor alpha (TNFα) activated or non-activated microglia BV2 cells. We showed that M-EVs are internalized by both mouse and human C20 microglia cells and that the uptake of M-EVs in microglia induced autophagic vesicles at various stages of degradation including autophagosomes and autolysosomes. Consistently, stimulation of microglia with M-EVs increased the protein expression of the autophagy marker, microtubule-associated proteins 1A/1B light chain 3B isoform II (LC3B-II), and promoted autophagic flux in live cells. To elucidate the biological activities occurring at the transcriptional level in C20 microglia stimulated with M-EVs, the gene expression profiles, potential upstream regulators, and enrichment pathways were characterized using targeted RNA sequencing. Inflammation and immunity transcriptome gene panel sequencing of both activated and normal microglia stimulated with M-EVs showed involvement of several canonical pathways and reduced expression of key genes involved in neuroinflammation, inflammasome and apoptosis signalling pathways compared to control cells. In this study, we provide the perspective that a beneficial activity of in vitro cell culture produced EVs could be the modulation of autophagy during cellular stress. Therefore, we use a monoculture system to study microglia-microglia crosstalk which is important in the prevention and propagation of inflammation in the brain. We demonstrate that in vitro produced microglial EVs are able to influence multiple biological pathways and promote activation of autophagy in order to maintain microglia survival and homeostasis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The authors declare that they have no conflict of interest., (© 2020 The Authors. Journal of Extracellular Vesicles published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.)

Irreproducibility of research results is one of the major contributing factors to the failure of translating basic research results into tangible bedside progress. To address this, the University Biobank Limburg (UBiLim) was founded by a collaboration between Hasselt University, the Hospital East-Limburg, and the Jessa Hospital. This paper describes the evolution of this process and the barriers encountered on the way. UBiLim evolved from an archival collection over a single-site biobank into a federated structure, supporting translational research at the founding institutions. Currently, UBiLim is a federated biobank, with an established organizational structure and processing, and storage facilities at each of the three sites. All activities are integrated in an ISO15189-accredited Quality Management System and based on (inter)national biobank guidelines. Common methods for processing and storage of a plethora of sample types, suitable for state-of-the-art applications, were validated and implemented. Because the biobank is embedded in two hospitals, the request of researchers to include certain sample types or enroll specific patient groups can quickly be met. Funding has been a major challenge in each step of its evolution and remains the biggest issue for long-term biobank sustainability. To a lesser extent, the Belgian legislation and the operational cost of information management system are also concerns for smooth biobank operations. Nonetheless, UBiLim serves as a facilitator and accelerator for translational research in the Limburg area of Belgium that, given the fields of research, may have an impact on international patient care., (Copyright © 2019 Linsen, Vanhees, Vanoppen, Ulenaers, Driessens, Penders, Somers, Stinissen and Rummens.)

IgD - CD27 - double negative (DN) B cells with proinflammatory characteristics are abnormally elevated in a proportion of multiple sclerosis (MS) patients. In this study, the origin and selection characteristics of DN B cells were studied in MS patients and healthy controls (HC). Expression of developmental markers on peripheral blood DN, IgD - CD27 + class-switched memory (CSM) and IgD + CD27 - naive B cells of HC ( n = 48) and MS patients ( n = 96) was determined by flow cytometry. High-throughput adaptive immune receptor repertoire sequencing was performed on peripheral blood DN and CSM B cells of HC and MS patients ( n = 3 each). DN B cells from HC and MS patients showed similar phenotypic and Ig repertoire characteristics. Phenotypic analysis indicated a mature state of DN B cells by low CD5, CD10, and CD38 expression. However, the frequency of CD95 + and IgA + cells was lower in DN versus CSM B cells. DN B cells are Ag experienced, as shown by somatic hypermutation of their Ig genes in adaptive immune receptor repertoire sequencing, although they showed a lower mutation load than CSM B cells. Shared clones were found between DN and CSM B cells, although >95% of the clones were unique to each population, and differences in V(D)J usage and CDR3 physicochemical properties were found. Thus, DN B cells arise in HC and MS patients via a common developmental pathway that is probably linked to immune aging. However, DN and CSM B cells develop through unique differentiation pathways, with most DN B cells representing an earlier maturation state., (Copyright © 2019 by The American Association of Immunologists, Inc.)

Autism spectrum disorder (ASD) is a neurodevelopmental disorder whose behavioral symptoms become apparent in early childhood. The underlying pathophysiological mechanisms are only partially understood and the clinical manifestations are heterogeneous in nature, which poses a major challenge for diagnosis, prognosis and intervention. In the last years, an important role of a dysregulated immune system in ASD has emerged, but the mechanisms connecting this to a disruption of brain development are still largely unknown. Although ASD is not considered as a typical autoimmune disease, self-reactive antibodies or autoantibodies against a wide variety of targets have been found in a subset of ASD patients. In addition, autoantibodies reactive to fetal brain proteins have also been described in the prenatal stage of neurodevelopment, where they can be transferred from the mother to the fetus by transplacental transport. In this review, we give an extensive overview of the antibodies described in ASD according to their target antigens, their different origins, and timing of exposure during neurodevelopment.

Multiple sclerosis (MS) is a demyelinating autoimmune disease in which innate and adaptive immune cells infiltrate the central nervous system (CNS) and damage the myelin sheaths surrounding the axons. Upon activation, infiltrated macrophages, CNS-resident microglia, and astrocytes switch their metabolism toward glycolysis, resulting in the formation of α-dicarbonyls, such as methylglyoxal (MGO) and glyoxal (GO). These potent glycating agents lead to the formation of advanced glycation endproducts (AGEs) after reaction with amino acids. We hypothesize that AGE levels are increased in MS lesions due to the inflammatory activation of macrophages and astrocytes. First, we measured tissue levels of AGEs in brain samples of MS patients and controls. Analysis of MS patient and non-demented control (NDC) specimens showed a significant increase in protein-bound N δ -(5-hydro-5-methyl-4-imidazolon-2-yl)-ornithine (MG-H1), the major AGE, compared to white matter of NDCs (107 ± 11 vs. 154 ± 21, p < 0.05). In addition, immunohistochemistry revealed that MGO-derived AGEs were specifically present in astrocytes, whereas the receptor for AGEs, RAGE, was detected on microglia/macrophages. Moreover, in cerebrospinal fluid from MS patients, α-dicarbonyls and free AGEs correlated with their respective levels in the plasma, whereas this was not observed for protein-bound AGEs. Taken together, our data show that MG-H1 is produced by astrocytes. This suggests that AGEs secreted by astrocytes have paracrine effects on RAGE-positive macrophages/microglia and thereby contribute to the pathology of MS.

Purpose: The purpose of this study was to evaluate objectively the effectiveness of photobiomodulation therapy (PBMT) for the prevention of acute radiation dermatitis (ARD) by using biophysical skin measurements., Methods: A randomized, placebo-controlled trial with 120 breast cancer patients who underwent an identical radiotherapy (RT) regimen post-lumpectomy was performed (TRANSDERMIS trial). Patients were randomized to receive PBM (808 nm CW/905 nm pulsed, 168 mW/cm 2 , spot size 19.6 cm 2 , fluence 4 J/cm 2 ) or placebo treatments from the first day of RT (2×/week). Biophysical skin measurements were collected to assess the skin pigmentation and barrier function. Measurements were collected at the first day of RT, a RT dose of 40 Gray (Gy), and the end of RT (66 Gy)., Results: The incidence of moist desquamation was significantly higher in the control than in the PBMT group at the end of RT (30 vs. 7%, respectively, odds ratio = 6, p = 0.004). The biophysical skin measures showed that the mean percentage change from the baseline transepidermal water loss (TEWL), erythema, and melanin values was significantly higher in the control than in the PBMT group at the end of RT (ps < 0.05). Logistic regression analysis revealed that the risk on moist desquamation was significantly increased for patients with a large (> 800 cc) breast volume (odds ratio = 4, p = 0.017)., Conclusions: This is the first randomized controlled trial demonstrating by objective measurements that PBMT is effective in reducing the incidence of moist desquamation in breast cancer patients undergoing RT. Additionally, a large breast volume is an important risk factor for the development of moist desquamation.

Multiple sclerosis (MS) is a chronic inflammatory autoimmune disease of the central nervous system (CNS) in which demyelination and neurodegeneration occurs. The immune system of MS patients is characterized by a dysregulation in the balance between pro- and anti-inflammatory immune cells, whereby both the innate and adaptive immune system are involved. Dimethyl fumarate (DMF) was licensed in 2013 as an oral first-line therapy for relapsing-remitting (RR)MS patients. It has a strong efficacy with neuroprotective and immunomodulatory effects and a favourable benefit-risk profile. However, the effects of DMF on the immune system of MS patients were not clear before entering the market. During the last years, numerous in vitro and ex vivo studies have clarified the working mechanism of DMF in MS. Here, we discuss the pharmacokinetics of DMF and its effect on molecular immune-related pathways, which is further linked to the clinical and immunological effects of DMF treatment. The efficacy and safety of DMF treatment for RRMS is discussed as reported from clinical trials. Further, the immunological effects of DMF treatment in RRMS patients are addressed in more detail, including the distribution and function of immune cells. Taken together, evidence from recent studies points to a multifactorial working mechanism of DMF treatment in MS which leads to a restored immune balance favouring a more tolerogenic or anti-inflammatory immune profile., (Copyright © 2018 Elsevier B.V. All rights reserved.)

As seen in this CME online activity (available at http://journal.cme.chestnet.org/sv-sleep-disorder), central sleep apnea (CSA) is associated with increased mortality in patients with heart failure (HF), and it has been thought that treatment of CSA may improve underlying HF. Positive airway pressure therapy, specifically auto-servoventilation (ASV), can not only suppress abnormal breathing patterns but has been reported to improve cardiac function in HF patients with CSA. In patients with HF and with CSA unsuppressed with CPAP, newer ASV use has been associated with significant CSA improvement; in addition, several studies have reported efficacy of ASV in the treatment of underlying cardiac dysfunction in HF patients with CSA. However, results from the large randomized Treatment of Sleep-Disordered Breathing with Predominant Central Sleep Apnea by Adaptive Servo-Ventilation in Patients with Heart Failure (SERVE-HF) trial (Cowie MR, Woehrle H, Wegscheider K, et al. Adaptive servo-ventilation for central sleep apnea in systolic heart failure. New Engl J Med. 2015;373[12]:1095-1105) showed no significant effect on the primary end point in patients with stable HF with reduced ejection fraction and predominantly CSA, but all-cause and cardiovascular mortality were both increased in the ASV arm. These results are surprising and inconsistent with earlier smaller studies reporting positive surrogate outcomes, and they require additional study and resolution. However, until this is done, there is an urgent educational need for review of the approved labeling and validated clinical use of ASV within the medical community. The purpose of this educational activity is to review the appropriate use of ASV for the treatment of sleep-disordered breathing, including Cheyne-Stokes respiration, treatment-emergent central apnea, and opioid-induced periodic breathing. Emphasis will be placed on proper patient and therapy selection, especially in patients with HF., (Copyright © 2018 American College of Chest Physicians. Published by Elsevier Inc. All rights reserved.)

The effects of dimethyl fumarate (DMF) on the immune system in multiple sclerosis (MS) are not completely elucidated. In this study, an extensive immunophenotypic analysis of innate and adaptive immune cells of DMF-treated MS patients was performed. Peripheral blood immune cell phenotypes were determined using flow cytometry in a follow-up study of 12 MS patients before, after 3 and 12 months of DMF treatment and a cross-sectional study of 25 untreated and 64 DMF-treated MS patients. Direct effects of DMF on B cells were analyzed in vitro. After 12 months of DMF treatment, percentages of monocytes, natural killer cells, naive T and B cells and transitional B cells increased. Percentages of (effector) memory T cells, (non) class-switched memory B cells and double negative B cells decreased together with CD4 + T cells expressing interferon-γ (IFN-γ), granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin-17 (IL-17). DMF treatment was fully effective as of 6 months and directly induced apoptosis and decreased expression of costimulatory CD40, antigen presentation molecule MHCII and B cell activating factor receptor (BAFFR) on B cells. DMF induced a persistent change of the immune system of MS patients, directly induced apoptosis and reduced expression of functional markers on B cells.

B cells possess a predominant role in adaptive immune responses via antibody-dependent and -independent functions. The microbiome of the gastrointestinal tract is currently being intensively investigated due to its profound impact on various immune responses, including B cell maturation, activation, and IgA antibody responses. Recent findings have demonstrated the interplay between dietary components, gut microbiome, and autoantibody production. "Western" dietary patterns, such as high fat and high salt diets, can induce alterations in the gut microbiome that in turn affects IgA responses and the production of autoantibodies. This could contribute to multiple pathologies including autoimmune and inflammatory diseases. Here, we summarize current knowledge on the influence of various dietary components on B cell function and (auto)antibody production in relation to the gut microbiota, with a particular focus on the gut-brain axis in the pathogenesis of multiple sclerosis.

Objective: Acute radiodermatitis (RD) is a distressing and painful skin reaction that occurs in 95% of the patients undergoing radiotherapy (RT). The aim of this study was to evaluate the effectiveness of photobiomodulation therapy (PBMT) in the prevention of acute RD in breast cancer (BC) patients undergoing RT., Methods: This study was a randomized, placebo-controlled trial including 120 BC patients that underwent an identical RT regimen post-lumpectomy. Patients were randomly assigned to the laser therapy (LT) or placebo group, with 60 patients in each group. Laser or placebo treatments were applied 2 days a week, immediately after the RT session, starting at the first day of RT. PBMT was delivered using a class IV MLS ® M6 laser that combines two synchronized laser diodes in the infrared range (808-905 nm) with a fixed energy density (4 J/cm 2 ). Skin reactions were scored based on the criteria of the Radiation Therapy Oncology Group (RTOG) and the Radiation-Induced Skin Reaction Assessment Scale (RISRAS). The patients completed the Skindex-16 questionnaire to evaluate their quality of life. All the measurements were collected at the first day, at a RT dose of 40 Gray (Gy), and at the end of RT (total dose 66 Gy)., Results: At a RT dose of 40 Gy, there was no significant difference between the groups in the distribution of RTOG grades. However, at the end of RT the severity of the skin reactions significantly differed between the two groups (P = 0.004), with a larger percentage of patients experiencing RTOG grade 2 or higher (e.g., moist desquamation) in the placebo group (30% vs. 6.7%, for the placebo and laser group, resp.). The objective RISRAS score confirmed these results. In addition, the Skindex-16 and RISRAS subjective score demonstrated that the patients' quality of life was significantly better in the LT than in the control group., Conclusions: The results of this trial show that PBMT is an effective tool to prevent the development of grade 2 acute RD or higher in BC patients. In addition, it also reduces the patients' symptoms related to RD. Lasers Surg. Med. © 2018 Wiley Periodicals, Inc., (© 2018 Wiley Periodicals, Inc.)

Multiple sclerosis (MS) is an inflammatory auto-immune disease of the central nervous system (CNS). Serum glucose alterations and impaired glucose tolerance (IGT) are reported in MS patients, and are commonly associated with the development of cardio-metabolic co-morbidities. We previously found that a subgroup of MS patients shows alterations in their lipoprotein profile that are similar to a pre-cardiovascular risk profile. In addition, we showed that a high-intensity exercise training has a positive effect on IGT in MS patients. In this study, we hypothesize that exercise training positively influences the lipoprotein profile of MS patients. To this end, we performed a pilot study and determined the lipoprotein profile before (controls, n = 40; MS patients, n = 41) and after ( n = 41 MS only) 12 weeks of medium-intensity continuous training (MIT, n = 21, ~60% of VO 2max ) or high-intensity interval training (HIT, n = 20, ~100-200% of VO 2max ) using nuclear magnetic resonance spectroscopy (NMR). Twelve weeks of MIT reduced intermediate-density lipoprotein particle count ((nmol/L); -43.4%; p < 0.01), low-density lipoprotein cholesterol (LDL-c (mg/dL); -7.6%; p < 0.05) and VLDL size ((nm); -6.6%; p < 0.05), whereas HIT did not influence the lipoprotein profile. These results show that MIT partially normalizes lipoprotein alterations in MS patients. Future studies including larger patient and control groups should determine whether MIT can reverse other lipoprotein levels and function and if these alterations are related to MS disease progression and the development of co-morbidities., Competing Interests: The authors declare no conflict of interest. The funding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, and in the decision to publish the results.

Multiple sclerosis (MS) is the leading cause of chronic neurological disability in young adults. The clinical disease course of MS varies greatly between individuals, with some patients progressing much more rapidly than others, making prognosis almost impossible. We previously discovered that cytotoxic CD4+ T cells (CD4+ CTL), identified by the loss of CD28, are able to migrate to sites of inflammation and that they contribute to tissue damage. Furthermore, in an animal model for MS, we showed that these cells are correlated with inflammation, demyelination, and disability. Therefore, we hypothesize that CD4+ CTL drive progression of MS and have prognostic value. To support this hypothesis, we investigated whether CD4+ CTL are correlated with worse clinical outcome and evaluated the prognostic value of these cells in MS. To this end, the percentage of CD4+CD28null T cells was measured in the blood of 176 patients with relapsing-remitting MS (=baseline). Multimodal evoked potentials (EP) combining information on motoric, visual, and somatosensoric EP, as well as Kurtzke expanded disability status scale (EDSS) were used as outcome measurements at baseline and after 3 and 5 years. The baseline CD4+CD28null T cell percentage is associated with EP ( P  = 0.003, R 2  = 0.28), indicating a link between these cells and disease severity. In addition, the baseline CD4+CD28null T cell percentage has a prognostic value since it is associated with EP after 3 years ( P  = 0.005, R 2  = 0.29) and with EP and EDSS after 5 years ( P  = 0.008, R 2  = 0.42 and P  = 0.003, R 2  = 0.27). To the best of our knowledge, this study provides the first direct link between the presence of CD4+ CTL and MS disease severity, as well as its prognostic value. Therefore, we further elaborate on two important research perspectives: 1° investigating strategies to block or reverse pathways in the formation of these cells resulting in new treatments that slow down MS disease progression, 2° including immunophenotyping in prediction modeling studies to aim for personalized medicine.

Cytomegalovirus (CMV) is a latent virus which causes chronic activation of the immune system. Here, we demonstrate that cytotoxic and pro-inflammatory CD4 + CD28 null T cells are only present in CMV seropositive donors and that CMV-specific Immunoglobulin (Ig) G titers correlate with the percentage of these cells. In vitro stimulation of peripheral blood mononuclear cells with CMVpp65 peptide resulted in the expansion of pre-existing CD4 + CD28 null T cells. In vivo, we observed de novo formation, as well as expansion of CD4 + CD28 null T cells in two different chronic inflammation models, namely the murine CMV (MCMV) model and the experimental autoimmune encephalomyelitis (EAE) model for multiple sclerosis (MS). In EAE, the percentage of peripheral CD4 + CD28 null T cells correlated with disease severity. Pre-exposure to MCMV further aggravated EAE symptoms, which was paralleled by peripheral expansion of CD4 + CD28 null T cells, increased splenocyte MOG reactivity and higher levels of spinal cord demyelination. Cytotoxic CD4 + T cells were identified in demyelinated spinal cord regions, suggesting that peripherally expanded CD4 + CD28 null T cells migrate towards the central nervous system to inflict damage. Taken together, we demonstrate that CMV drives the expansion of CD4 + CD28 null T cells, thereby boosting the activation of disease-specific CD4 + T cells and aggravating autoimmune mediated inflammation and demyelination.

In traumatic spinal cord injury (SCI) patients, the assessment of the exact degree of lesion severity and neurological prognosis has proven to be extremely challenging. The current tools for predicting functional outcome in SCI patients such as clinical examination and magnetic resonance imaging are often inaccessible to unstable or polytraumatized patients, lack sensitivity, and are unreliable in the acute phase of the injury. Multiple candidate protein biomarkers known to be linked to the pathology have been studied for their potential to predict neurological outcome over time. This hypothesis-driven approach, however, has yielded only minimal success, and the reported individual markers lack sensitivity and correlation with specific outcome measures, which highlights the need for an unbiased high-throughput screening approach to identify novel candidate biomarkers. Antibodies were suggested to represent better biomarkers as their counterpart antigens, as they are highly specific and abundantly present in blood due to their inherent amplification and long half-life. Moreover, antibodies are easily accessible and are amenable to high-throughput screening. We therefore suggest an unbiased, high-throughput, and powerful antibody profiling procedure, named Serological antigen selection, based on complementary DNA (cDNA) phage display to combine the multiple benefits of stable and frequent antibodies and those of an unbiased method. The application of such an innovative and unbiased approach can complement the more traditional approaches to aid in the discovery of novel SCI-associated biomarkers.

Lipoproteins modulate innate and adaptive immune responses. In the chronic inflammatory disease multiple sclerosis (MS), reports on lipoprotein level alterations are inconsistent and it is unclear whether lipoprotein function is affected. Using nuclear magnetic resonance (NMR) spectroscopy, we analysed the lipoprotein profile of relapsing-remitting (RR) MS patients, progressive MS patients and healthy controls (HC). We observed smaller LDL in RRMS patients compared to healthy controls and to progressive MS patients. Furthermore, low-BMI (BMI ≤ 23 kg/m 2 ) RRMS patients show increased levels of small HDL (sHDL), accompanied by larger, triglyceride (TG)-rich VLDL, and a higher lipoprotein insulin resistance (LP-IR) index. These alterations coincide with a reduced serum capacity to accept cholesterol via ATP-binding cassette (ABC) transporter G1, an impaired ability of HDL 3 to suppress inflammatory activity of human monocytes, and modifications of HDL 3 's main protein component ApoA-I. In summary, lipoprotein levels and function are altered in RRMS patients, especially in low-BMI patients, which may contribute to disease progression in these patients.

Objectives: To provide evidence-based recommendations and guidance to the public regarding indicators of good sleep quality across the life-span., Methods: The National Sleep Foundation assembled a panel of experts from the sleep community and representatives appointed by stakeholder organizations (Sleep Quality Consensus Panel). A systematic literature review identified 277 studies meeting inclusion criteria. Abstracts and full-text articles were provided to the panelists for review and discussion. A modified Delphi RAND/UCLA Appropriateness Method with 3 rounds of voting was used to determine agreement., Results: For most of the sleep continuity variables (sleep latency, number of awakenings >5minutes, wake after sleep onset, and sleep efficiency), the panel members agreed that these measures were appropriate indicators of good sleep quality across the life-span. However, overall, there was less or no consensus regarding sleep architecture or nap-related variables as elements of good sleep quality., Conclusions: There is consensus among experts regarding some indicators of sleep quality among otherwise healthy individuals. Education and public health initiatives regarding good sleep quality will require sustained and collaborative efforts from multiple stakeholders. Future research should explore how sleep architecture and naps relate to sleep quality. Implications and limitations of the consensus recommendations are discussed., (Copyright © 2016 National Sleep Foundation. Published by Elsevier Inc. All rights reserved.)

Rheumatoid arthritis (RA) is the world's most common autoimmune disease mainly characterized by a chronic inflammation of multiple synovial joints. Rheumatologists now have a whole range of treatment options including glucocorticoids (GCs), classical synthetic and biological disease-modifying antirheumatic drugs (cs- and bDMARDS), resulting in a tremendous improvement in treatment outcomes for RA patients over the last two decades. Despite this progress, the choice of treatment regimen to achieve stable remission at the individual patient level still largely depends on trial and error. In this review, the need for novel theranostic markers that can predict a patient's response to methotrexate, the standard first-line csDMARD treatment, is discussed. Like in many autoimmune diseases, the majority of RA patients form a whole range of autoantibodies. We aim to find novel theranostic autoantibody markers using serological antigen selection, a high-throughput technique that uses cDNA phage display to identify novel antigen targets. We have constructed a barcoded cDNA phage display library from the synovial tissue of three RA patients by fusing cDNA products to the filamentous phage minor coat protein VI. This library contains a large proportion of full-length genes and gene fragments that are cloned in frame with the phage gene VI. By screening this library for antibody reactivity in serum samples of patients from the CareRA trial, which compared different intensive treatment strategies based on csDMARDs and a step-down GC schedule, our cDNA phage display library has great potential for the discovery of novel theranostic autoantibody biomarkers.

Introduction: Estimating body fat content has shown to be a better predictor of adiposity-related cardiovascular risk than the commonly used body mass index (BMI). The white-light 3D body volume index (BVI) scanner is a non-invasive device normally used in the clothing industry to assess body shapes and sizes. We assessed the hypothesis that volume obtained by BVI is comparable to the volume obtained by air displacement plethysmography (Bod-Pod) and thus capable of assessing body fat mass using the bi-compartmental principles of body composition., Methods: We compared BVI to Bod-pod, a validated bicompartmental method to assess body fat percent that uses pressure/volume relationships in isothermal conditions to estimate body volume. Volume is then used to calculate body density (BD) applying the formula density=Body Mass/Volume. Body fat mass percentage is then calculated using the Siri formula (4.95/BD - 4.50) × 100. Subjects were undergoing a wellness evaluation. Measurements from both devices were obtained the same day. A prediction model for total Bod-pod volume was developed using linear regression based on 80% of the observations (N=971), as follows: Predicted Bod-pod Volume (L)=9.498+0.805*(BVI volume, L)-0.0411*(Age, years)-3.295*(Male=0, Female=1)+0.0554*(BVI volume, L)*(Male=0, Female=1)+0.0282*(Age, years)*(Male=0, Female=1). Predictions for Bod-pod volume based on the estimated model were then calculated for the remaining 20% (N=243) and compared to the volume measured by the Bod-pod., Results: Mean age among the 971 individuals was 41.5 ± 12.9 years, 39.4% were men, weight 81.6 ± 20.9 kg, BMI was 27.8 ± 6.3kg/m 2 . Average difference between volume measured by Bod-pod- predicted volume by BVI was 0.0 L, median: -0.4 L, IQR: -1.8 L to 1.5 L, R2=0.9845. Average difference between body fat measured-predicted was-1%, median: -2.7%, IQR: -13.2 to 9.9, R2=0.9236., Conclusion: Volume and BFM can be estimated by using volume measurements obtained by a white- light 3D body scanner and the prediction model developed in this study., Competing Interests: Conflicts of interest: All authors declare no conflict of interest. Select Research (Worcester, UK), the company that designed the 3D BVI body scanner and software provided without charge the scanner used for this and other studies taking place at Mayo Clinic.

Immune aging occurs in the elderly and in autoimmune diseases. Recently, IgD - CD27 - (double negative, DN) and CD21 - CD11c + (CD21 low ) B cells were described as age-associated B cells with proinflammatory characteristics. This study investigated the prevalence and functional characteristics of DN and CD21 low B cells in multiple sclerosis (MS) patients. Using flow cytometry, we demonstrated a higher proportion of MS patients younger than 60 y with peripheral expansions of DN (8/41) and CD21 low (9/41) B cells compared with age-matched healthy donors (1/33 and 2/33, respectively), which indicates an increase in age-associated B cells in MS patients. The majority of DN B cells had an IgG + memory phenotype, whereas CD21 low B cells consisted of a mixed population of CD27 - naive, CD27 + memory, IgG + , and IgM + cells. DN B cells showed similar (MS patients) or increased (healthy donors) MHC-II expression as class-switched memory B cells and intermediate costimulatory molecule expression between naive and class-switched memory B cells, indicating their potential to induce (proinflammatory) T cell responses. Further, DN B cells produced proinflammatory and cytotoxic cytokines following ex vivo stimulation. Increased frequencies of DN and CD21 low B cells were found in the cerebrospinal fluid of MS patients compared with paired peripheral blood. In conclusion, a proportion of MS patients showed increased peripheral expansions of age-associated B cells. DN and CD21 low B cell frequencies were further increased in MS cerebrospinal fluid. These cells could contribute to inflammation by induction of T cell responses and the production of proinflammatory cytokines., (Copyright © 2016 by The American Association of Immunologists, Inc.)

Antibody-independent B cell functions play an important role in multiple sclerosis (MS) pathogenesis. In this study, B cell antigen presentation and costimulation in MS were studied. Peripheral blood B cells of MS patients showed increased expression of costimulatory CD86 and CD80 molecules compared with healthy controls (HC). In MS cerebrospinal fluid (CSF), 12-fold and 2-fold increases in CD86 + and CD80 + B cells, respectively, were evidenced compared with peripheral blood. Further, B cells from MS patients induced proinflammatory T cells in response to myelin basic protein (MBP). Immunomodulatory treatment restored B cell costimulatory molecule expression and caused significantly reduced B cell induced T cell responses. Together, these results demonstrate the potential of B cells from MS patients to induce autoreactive proinflammatory T cell responses. Immunomodulatory therapy abrogated this effect, emphasizing the importance of B cell antigen presentation and costimulation in MS pathology., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Recent evidence implicates antibody responses as pivotal damaging factors in spinal cord injury (SCI)-induced neuroinflammation. To date, only a limited number of the antibody targets have been uncovered, and the discovery of novel targets with pathologic and clinical relevance still represents a major challenge., Methods: In this study, we, therefore, applied an unbiased, innovative and powerful strategy, called serological antigen selection (SAS), to fully identify the complex information present within the antibody repertoire of SCI patients., Results: We constructed a high-quality cDNA phage display library derived from human spinal cord tissue to screen for antibody reactivity in pooled plasma samples from traumatic SCI patients (n = 10, identification cohort). By performing SAS, we identified a panel of 19 antigenic targets to which the individual samples of the plasma pool showed antibody reactivity. Sequence analysis to identify the selected antigenic targets uncovered 5 known proteins, to which antibody reactivity has not been associated with SCI before, as well as linear peptides. Immunoreactivity against 9 of the 19 novel identified targets was validated in 41 additional SCI patients and an equal number of age- and gender-matched healthy subjects. Overall, we found elevated antibody levels to at least 1 of the 9 targets in 51 % of our total SCI patient cohort (n = 51) with a specificity of 73 %. By combining 6 of these 9 targets into a panel, an overall reactivity of approximately half of the SCI patients could be maintained while increasing the specificity to 82 %., Conclusions: In conclusion, our innovative high-throughput approach resulted in the identification of previously unexplored antigenic targets with elevated immunoreactivity in more than 50 % of the SCI patients. Characterization of the validated antibody responses and their targets will not only provide new insight into the underlying disease processes of SCI pathology but also significantly contribute to uncovering potential antibody biomarkers for SCI patients.

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) characterized by demyelination, axonal degeneration and gliosis. The progressive form of MS is an important research topic as not much is known about its underlying mechanisms and no therapy is available. Although progressive MS is traditionally considered to be driven by neurodegeneration, compartmentalized CNS inflammation is currently accepted as one of the driving processes behind neurodegeneration and progression. In this review, the involvement of B cells and antibodies in progressive MS is discussed. The identification of meningeal ectopic B cell follicles in secondary progressive MS (SPMS) patients and the successful use of B cell-depleting therapy in primary progressive MS (PPMS) patients have underlined the importance of B cells in progressive MS. Proof is also available for the role of antibodies in neurodegeneration and progression in MS. Here, oligoclonal immunoglobulin M (IgM) production and autoreactive antibodies are described, with a focus on antibodies directed against sperm-associated antigen 16 (SPAG16). Further research into the role of B cells and autoantibodies in MS progression can lead to novel prognostic and theranostic opportunities., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Objectives: Despite recent progress in biomarker discovery for RA diagnostics, still over one-third of RA patients-and even more in early disease-present without RF or ACPA. The aim of this study was to confirm the presence of previously identified autoantibodies to novel Hasselt University (UH) peptides in early and seronegative RA., Methods: Screening for antibodies against novel UH peptides UH-RA.1, UH-RA.9, UH-RA.14 and UH-RA.21, was performed in two large independent cohorts. Peptide ELISAs were developed to screen for the presence of antibodies to UH-RA peptides. First, 292 RA patients (including 39 early patients), 90 rheumatic and 97 healthy controls from UH were studied. Antibody reactivity to two peptides (UH-RA.1 and UH-RA.21) was also evaluated in 600 RA patients, 309 patients with undifferentiated arthritis and 157 rheumatic controls from the Leiden Early Arthritis Clinic cohort., Results: In both cohorts, 38% of RA patients were seronegative for RF and ACPA. Testing for autoantibodies to UH-RA.1 and UH-RA.21 reduced the serological gap from 38% to 29% in the UH cohort (P = 0.03) and from 38% to 32% in the Leiden Early Arthritis Clinic cohort (P = 0.01). Furthermore, 19-33% of early RA patients carried antibodies to these peptides. Specificities in rheumatic controls ranged from 82 to 96%. Whereas antibodies against UH-RA.1 were related to remission, anti-UH-RA.21 antibodies were associated with inflammation, joint erosion and higher tender and swollen joint counts., Conclusion: This study validates the presence of antibody reactivity to novel UH-RA peptides in seronegative and early RA. This might reinforce current diagnostics and improve early diagnosis and intervention in RA., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Ankylosing spondylitis (AS) is a debilitating, chronic, rheumatic disease characterized by inflammation and new bone formation resulting in fusion of the spine and sacroiliac joints. Since early treatment is impeded by a delayed diagnosis, it is highly important to find new biomarkers that improve early diagnosis and may also contribute to a better assessment of disease activity, prognosis and therapy response in AS. Because of the absence of rheumatoid factor, AS was long assumed to have a seronegative character and antibodies are thus not considered a hallmark of the disease. However, emerging evidence suggests plasma cells and autoantibodies to be involved in the disease course. In this review, the role of B cells and antibodies in AS is discussed. Furthermore, an overview is provided of antibodies identified in AS up till now, and their diagnostic potential. Many of these antibody responses were based on small study populations and further validation is lacking. Moreover, most were identified by a hypothesis-driven approach and thus limited to antibodies against targets that are already known to be involved in AS pathogenesis. Hence, we propose an unbiased approach to identify novel diagnostic antibodies. The already successfully applied techniques cDNA phage display and serological antigen selection will be used to identify antibodies against both known and new antigen targets in AS plasma. These newly identified antibodies will enhance early diagnosis of AS and provide more insight into the underlying disease pathology, resulting in a more effective treatment strategy and eventually an improved disease outcome., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: Recently, autoantibodies against novel UH-RA peptides (UH-RA.1 and UH-RA.21) were identified as candidate biomarkers for patients with rheumatoid arthritis (RA) who are seronegative for the current diagnostic markers rheumatoid factor and anticitrullinated protein antibodies. Previously, screening for anti-UH-RA autoantibodies was based on measuring the immunoglobulin (Ig) G response. We aimed to investigate whether measurement of other isotypes could improve the performance of diagnostic testing. In addition, assigning the isotype profile might provide valuable information on effector functions of the antibodies., Methods: The isotype profile of antibodies against UH-RA.1 and UH-RA.21 was studied. The IgG, IgM, and IgA classes, together with the 4 different IgG subclasses, were determined in 285 patients with RA, 88 rheumatic control subjects, and 90 healthy control subjects., Results: Anti-UH-RA.1 antibodies were primarily of the IgM isotype and twice as prevalent as IgG (IgG3-dominated) and IgA. RA sensitivity when testing for anti-UH-RA.1 IgM was shown to be higher than when testing for the IgG isotype: 18 % versus 9 % sensitivity when RA specificity was set to 90 %. Within antibodies against UH-RA.21, IgG and IgA were more common than IgM. Different anti-UH-RA.21 IgG subclasses were found, with the highest prevalence found for IgG2. Combined testing for IgG and IgA slightly increased RA sensitivity of UH-RA.21-specific antibody testing to 27 % compared with solely testing for IgG (23 %). Notably, a higher number of anti-UH-RA.21 antibody isotypes was related to increased levels of erythrocyte sedimentation rate. Finally, for both antibody responses, the full antibody isotype use was demonstrated in early and seronegative disease., Conclusions: The isotype distribution of anti-UH-RA.1 and anti-UH-RA.21 antibodies was successfully outlined, and, for antibodies against UH-RA.1, we found that isotype-specific testing might have implications for diagnostic testing. The exact mechanisms by which the different antibody isotypes act still have to be unraveled.

Background: Body composition changes with aging lead to increased adiposity and decreased muscle mass, making the diagnosis of obesity challenging. Conventional anthropometry, including body mass index (BMI), while easy to use clinically may misrepresent adiposity. We determined the diagnostic accuracy of BMI using dual-energy X-ray absorptiometry (DEXA) in assessing the degree of obesity in older adults., Methods: The National Health and Nutrition Examination Surveys 1999-2004 were used to identify adults aged ⩾60 years with DEXA measures. They were categorized (yes/no) as having elevated body fat by gender (men: ⩾25%; women ⩾35%) and by BMI ⩾25 and ⩾30 kg m(-)(2). The diagnostic performance of BMI was assessed. Metabolic characteristics were compared in discordant cases of BMI/body fat. Weighting and analyses were performed per NHANES (National Health and Nutrition Examination Survey) guidelines., Results: We identified 4984 subjects (men: 2453; women: 2531). Mean BMI and % body fat was 28.0 kg m(-2) and 30.8% in men, and 28.5 kg m(-)(2) and 42.1% in women. A BMI ⩾30 kg m(-)(2) had a low sensitivity and moderately high specificity (men: 32.9 and 80.8%, concordance index 0.66; women: 38.5 and 78.5%, concordance 0.69) correctly classifying 41.0 and 45.1% of obese subjects. A BMI ⩾25 kg m(-2) had a moderately high sensitivity and specificity (men: 80.7 and 99.6%, concordance 0.81; women: 76.9 and 98.8%, concordance 0.84) correctly classifying 80.8 and 78.5% of obese subjects. In subjects with BMI <30 kg m(-)(2), body fat was considered elevated in 67.1% and 61.5% of men and women, respectively. For a BMI ⩾30 kg m(-)(2), sensitivity drops from 40.3% to 14.5% and 44.5% to 23.4%, whereas specificity remains elevated (>98%), in men and women, respectively, in those 60-69.9 years to subjects aged ⩾80 years. Correct classification of obesity using a cutoff of 30 kg m(-)(2) drops from 48.1 to 23.9% and 49.0 to 19.6%, in men and women in these two age groups., Conclusions: Traditional measures poorly identify obesity in the elderly. In older adults, BMI may be a suboptimal marker for adiposity.