Interferon and anti-TNF therapies differentially modulate amygdala reactivity which predicts associated bidirectional changes in depressive symptoms.

A third of patients receiving Interferon-α (IFN-α) treatment for Hepatitis-C develop major depressive disorder (MDD). Conversely, anti-Tumor Necrosis Factor (TNF) therapies improve depression providing key empirical support for the "inflammatory theory" of depression. Heightened amygdala reactivity (particularly to negatively valanced stimuli) is a consistent finding within MDD; can predict treatment efficacy and reverses following successful treatment. However, whether IFN-α and anti-TNF enhance/attenuate depressive symptoms through modulation of amygdala emotional reactivity is unknown. Utilizing a prospective study design, we recruited 30 patients (mean 48.0 ± 10.5 years, 21 male) initiating IFN-α treatment for Hepatitis-C and 30 (mean 50.4 ± 15.7 years, 10 male) anti-TNF therapy for inflammatory arthritis. All completed an emotional face-processing task during fMRI and blood sampling before and after their first IFN-α (4-h) or anti-TNF (24-h) injection and follow-up psychiatric assessments for 3 months of treatment. IFN-α significantly increased depression symptoms (Hamilton Depression Rating Scale HAM-D) at 4 weeks (p < 0.001) but not 4-h after first dose (p > 0.1). Conversely, anti-TNF significantly improved depressive symptoms (Hospital Anxiety and Depression Rating Scale HADS) at both 24-h (P = 0.015) and 12 weeks (p = 0.018). In support of our a-priori hypothesis, both IFN-α and anti-TNF significantly modulated amygdala reactivity with IFN-α acutely enhancing right amygdala responses to sad (compared with neutral) faces (p = 0.032) and anti-TNF conversely decreasing right amygdala reactivity (across emotional valence) (p = 0.033). Furthermore, these changes predicted IFN-induced increases in HAM-D 4 weeks later (R2 = 0.17, p = 0.022) and anti-TNF-associated decreases in HADS at 24-h (R2 = 0.23, p = 0.01) suggesting that actions of systemic inflammation on amygdala emotional reactivity play a mechanistic role in inflammation-associated depressive symptoms.

Expression of sterile-α and armadillo motif containing protein (SARM) in rheumatoid arthritis monocytes correlates with TLR2-induced IL-1ß and disease activity.

OBJECTIVE: Cartilage and bone damage in RA are associated with elevated IL-1ß. The effects of IL-1ß can be reduced by biological therapies that target IL-1ß or TNF-α. However, the mechanisms responsible for increased IL-1ß and the effect of anti-TNF-α have not been fully elucidated. Recently, sterile-α and armadillo motif containing protein (SARM) was identified as a negative regulator of toll-like receptor (TLR) induced IL-1ß secretion through an interaction with the inflammasome. This study set out to investigate SARM during TLR-induced IL-1ß secretion in RA peripheral blood monocytes and in patients commencing anti-TNF-α treatment. METHODS: Monocytes were isolated from RA patients and healthy controls; disease activity was measured by DAS28. IL-1ß secretion was measured by ELISA following TLR1/2, TLR4 and TLR7/8 stimulation. The mRNA expression of SARM1, IL-1ß and the components of the NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome were measured by quantitative PCR. SARM protein expression was measured by western blotting. RESULTS: TLR1/2 activation induced elevated IL-1ß in RA monocytes compared with healthy controls (P = 0.0009), which negatively correlated with SARM1 expression (P = 0.0086). Lower SARM expression also correlated with higher disease activity (P = 0.0246). Additionally, patients responding to anti-TNF-α treatment demonstrated a rapid upregulation of SARM, which was not observed in non-responders. CONCLUSION: Together, these data highlight a potential contribution from SARM to RA pathophysiology where decreased SARM may lead to elevated IL-1ß associated with RA pathogenesis. Furthermore, the data additionally present a potential mechanism by which TNF-α blockade can modify IL-1ß secretion.

TLR1/2 and 5 induce elevated cytokine levels from rheumatoid arthritis monocytes independent of ACPA or RF autoantibody status.

OBJECTIVE: RA is an autoimmune inflammatory joint disease. Both RF and ACPA are associated with more progressive disease and higher levels of systemic inflammation. Monocyte activation of toll-like receptors (TLRs) by endogenous ligands is a potential source of increased production of systemic cytokines. RA monocytes have elevated TLRs, some of which are associated with the disease activity score using 28 joints (DAS28). The aim of this study was to measure TLR-induced cytokine production from monocytes, stratified by autoantibody status, to assess if their capacity to induce cytokines is related to autoantibody status or DAS28. METHODS: Peripheral blood monocytes isolated from RA patients and healthy controls were stimulated with TLR1/2, TLR2/6, TLR4, TLR5, TLR7, TLR8 and TLR9 ligands for 18 h before measuring IL-6, TNFα and IL-10. Serum was used to confirm the autoantibody status. Cytokine levels were compared with RF, ACPA and DAS28. RESULTS: RA monocytes demonstrated significantly increased IL-6 and TNFα upon TLR1/2 stimulation and IL-6 and IL-10 upon TLR5 activation. TLR7 and TLR9 activation did not induce cytokines and no significant differences were observed between RA and healthy control monocytes upon TLR2/6, TLR4 or TLR8 activation. When stratified by ACPA or RF status there were no correlations between autoantibody status and elevated cytokine levels. However, TLR1/2-induced IL-6 did correlate with DAS28. CONCLUSIONS: Elevated TLR-induced cytokines in RA monocytes were not related to ACPA or RF status. However, TLR1/2-induced IL-6 was associated with disease activity.

Effect of rituximab on a salivary gland ultrasound score in primary Sjögren's syndrome: results of the TRACTISS randomised double-blind multicentre substudy.

OBJECTIVES: To compare the effects of rituximab versus placebo on salivary gland ultrasound (SGUS) in primary Sjögren's syndrome (PSS) in a multicentre, multiobserver phase III trial substudy. METHODS: Subjects consenting to SGUS were randomised to rituximab or placebo given at weeks 0, 2, 24 and 26, and scanned at baseline and weeks 16 and 48. Sonographers completed a 0-11 total ultrasound score (TUS) comprising domains of echogenicity, homogeneity, glandular definition, glands involved and hypoechoic foci size. Baseline-adjusted TUS values were analysed over time, modelling change from baseline at each time point. For each TUS domain, we fitted a repeated-measures logistic regression model to model the odds of a response in the rituximab arm (≥1-point improvement) as a function of the baseline score, age category, disease duration and time point. RESULTS: 52 patients (n=26 rituximab and n=26 placebo) from nine centres completed baseline and one or more follow-up visits. Estimated between-group differences (rituximab-placebo) in baseline-adjusted TUS were -1.2 (95% CI -2.1 to -0.3; P=0.0099) and -1.2 (95% CI -2.0 to -0.5; P=0.0023) at weeks 16 and 48. Glandular definition improved in the rituximab arm with an OR of 6.8 (95% CI 1.1 to 43.0; P=0.043) at week 16 and 10.3 (95% CI 1.0 to 105.9; P=0.050) at week 48. CONCLUSIONS: We demonstrated statistically significant improvement in TUS after rituximab compared with placebo. This encourages further research into both B cell depletion therapies in PSS and SGUS as an imaging biomarker. TRIAL REGISTRATION NUMBER: 65360827, 2010-021430-64; Results.

CMV-Specific T-cell Responses at Older Ages: Broad Responses With a Large Central Memory Component May Be Key to Long-term Survival.

Cytomegalovirus (CMV) infection sometimes causes large expansions of CMV-specific T cells, particularly in older people. This is believed to undermine immunity to other pathogens and to accelerate immunosenescence. While multiple different CMV proteins are recognized, most publications on age-related T-cell expansions have focused on dominant target proteins UL83 or UL123, and the T-cell activation marker interferon-γ (IFN-γ). We were concerned that this narrow approach might have skewed our understanding of CMV-specific immunity at older ages. We have, therefore, widened the scope of analysis to include in vitro-induced T-cell responses to 19 frequently recognized CMV proteins in "young" and "older" healthy volunteers and a group of "oldest old" long-term survivors (>85 years of age). Polychromatic flow cytometry was used to analyze T-cell activation markers (CD107, CD154, interleukin-2 [IL-2], tumor necrosis factor [TNF], and IFN-γ) and memory phenotypes (CD27, CD45RA). The older group had, on average, larger T-cell responses than the young, but, interestingly, response size differences were relatively smaller when all activation markers were considered rather than IFN-γ or TNF alone. The oldest old group recognized more proteins on average than the other groups, and had even bigger T-cell responses than the older group with a significantly larger central memory CD4 T-cell component.

Functional Diversity of Cytomegalovirus-Specific T Cells Is Maintained in Older People and Significantly Associated With Protein Specificity and Response Size.

BACKGROUND: Parallel upregulation of several T-cell effector functions (ie, polyfunctionality) is believed to be critical for the protection against viruses but thought to decrease in large T-cell expansions, in particular at older ages. The factors determining T-cell polyfunctionality are incompletely understood. Here we revisit the question of cytomegalovirus (CMV)-specific T-cell polyfunctionality, including a wide range of T-cell target proteins, response sizes, and participant ages. METHODS: Polychromatic flow cytometry was used to analyze the functional diversity (ie, CD107, CD154, interleukin 2, tumor necrosis factor, and interferon γ expression) of CD4+ and CD8+ T-cell responses to 19 CMV proteins in a large group of young and older United Kingdom participants. A group of oldest old people (age >85 years) was included to explore these parameters in exceptional survivors. Polyfunctionality was assessed for each protein-specific response subset, by subset and in aggregate, across all proteins by using the novel polyfunctionality index. RESULTS: Polyfunctionality was not reduced in healthy older people as compared to young people. However, it was significantly related to target protein specificity. For each protein, it increased with response size. In the oldest old group, overall T-cell polyfunctionality was significantly lower. DISCUSSION: Our results give a new perspective on T-cell polyfunctionality and raise the question of whether maintaining polyfunctionality of CMV-specific T cells at older ages is necessarily beneficial.

Optimising the use of electronic health records to estimate the incidence of rheumatoid arthritis in primary care: what information is hidden in free text?

BACKGROUND: Primary care databases are a major source of data for epidemiological and health services research. However, most studies are based on coded information, ignoring information stored in free text. Using the early presentation of rheumatoid arthritis (RA) as an exemplar, our objective was to estimate the extent of data hidden within free text, using a keyword search. METHODS: We examined the electronic health records (EHRs) of 6,387 patients from the UK, aged 30 years and older, with a first coded diagnosis of RA between 2005 and 2008. We listed indicators for RA which were present in coded format and ran keyword searches for similar information held in free text. The frequency of indicator code groups and keywords from one year before to 14 days after RA diagnosis were compared, and temporal relationships examined. RESULTS: One or more keyword for RA was found in the free text in 29% of patients prior to the RA diagnostic code. Keywords for inflammatory arthritis diagnoses were present for 14% of patients whereas only 11% had a diagnostic code. Codes for synovitis were found in 3% of patients, but keywords were identified in an additional 17%. In 13% of patients there was evidence of a positive rheumatoid factor test in text only, uncoded. No gender differences were found. Keywords generally occurred close in time to the coded diagnosis of rheumatoid arthritis. They were often found under codes indicating letters and communications. CONCLUSIONS: Potential cases may be missed or wrongly dated when coded data alone are used to identify patients with RA, as diagnostic suspicions are frequently confined to text. The use of EHRs to create disease registers or assess quality of care will be misleading if free text information is not taken into account. Methods to facilitate the automated processing of text need to be developed and implemented.

Variant connective tissue (joint hypermobility) and its relevance to depression and anxiety in adolescents: a cohort-based case-control study.

OBJECTIVE: To test whether variant connective tissue structure, as indicated by the presence of joint hypermobility, poses a developmental risk for mood disorders in adolescence. DESIGN: Cohort-based case-control study. SETTING: Data from the Avon Longitudinal Study of Parents and Children (ALSPAC) were interrogated. PARTICIPANTS: 6105 children of the ALSPAC cohort at age 14 years old, of whom 3803 also were assessed when aged 18 years. MAIN OUTCOME MEASURES: In a risk analysis, we examined the relationship between generalised joint hypermobility (GJH) at age 14 years with psychiatric symptoms at age 18 years. In an association analysis, we examined the relationship between presence of symptomatic joint hypermobility syndrome (JHS) and International Classification of Diseases-10 indication of depression and anxiety (Clinical Interview Schedule Revised (CIS-R), Anxiety Sensitivity Index) at age 18 years. RESULTS: GJH was more common in females (n=856, 28%) compared with males (n=319, 11%; OR: 3.20 (95% CI: 2.78 to 3.68); p<0.001). In males, GJH at age 14 years was associated with depression at 18 years (OR: 2.10 (95% CI: 1.17 to 3.76); p=0.013). An index of basal physiological arousal, elevated resting heart rate, mediated this effect. Across genders, the diagnosis of JHS at age 18 years was associated with the presence of depressive disorder (adjusted OR: 3.53 (95% CI: 1.67 to 7.40); p=0.001), anxiety disorder (adjusted OR: 3.14 (95% CI: 1.52 to 6.46); p=0.002), level of anxiety (B=8.08, t(3278)=3.95; p<0.001) and degree of psychiatric symptomatology (B=5.89, t(3442)=5.50; p<0.001). CONCLUSIONS: Variant collagen, indexed by joint hypermobility, is linked to the emergence of depression and anxiety in adolescence, an effect mediated by autonomic factors in males. Recognition of this association may motivate further evaluation, screening and interventions to mitigate development of psychiatric disorders and improve health outcomes.

The challenges of chronic pain and fatigue.

In this review, we explore the challenges of chronic pain and fatigue in clinical practice. Both pain and fatigue are common, troubling and frequently overlapping symptoms, and we describe both the clinical burden and the 'clinical problem'. We explore commonly associated symptoms and possible pathological associations, including variant connective tissue (joint hypermobility), small fibre neuropathy, mast cell activation, dysregulated inflammatory and interoceptive processes, which may inform treatment targets. We suggest a multidisciplinary management approach.

Contribution of Toll-Like Receptors and the NLRP3 Inflammasome in Rheumatoid Arthritis Pathophysiology.

Rheumatoid arthritis (RA) is a progressive autoimmune disease that is characterized by inflammation of the synovial joints leading to cartilage and bone damage. The pathogenesis is sustained by the production of pro-inflammatory cytokines including tumor necrosis factor (TNF), interleukin (IL)-1 and IL-6, which can be targeted therapeutically to alleviate disease severity. Several innate immune receptors are suggested to contribute to the chronic inflammation in RA, through the production of pro-inflammatory factors in response to endogenous danger signals. Much research has focused on toll-like receptors and more recently the nucleotide-binding domain and leucine-rich repeat pyrin containing protein-3 (NLRP3) inflammasome, which is required for the processing and release of IL-1ß. This review summarizes the current understanding of the potential involvement of these receptors in the initiation and maintenance of inflammation and tissue damage in RA and experimental arthritis models.

Streptococcus-associated vasculitis: A role for antibiotic therapy?

Numerous post-Streptococcal syndromes (PSS) have been described in the literature. The role of antibiotic therapy in the management of PSS is best established with acute rheumatic fever. We present a patient with streptococcus-associated medium vessel vasculitis with multiple flares despite immunosuppressive therapy that achieved a sustained remission with long term oral penicillin V 250 mg twice daily.

Cytomegalovirus infection is associated with an increase in aortic stiffness in older men which may be mediated in part by CD4 memory T-cells.

Human Cytomegalovirus (CMV) infection is associated with atherosclerosis, higher cardiovascular disease (CVD) risk, and an increase in memory T-cells (Tmem). T-cells have also been implicated in CVD, independently of CMV infection. To better understand the CMV-associated CVD risk, we examined the association between CMV (IgG) serostatus and central aortic (carotid-to-femoral) pulse wave velocity (cfPWV), an early, independent predictor of CVD. We also investigated if such an association might be reflected by the distribution of Tmem and/or other T-cell subsets. Methods: Healthy older volunteers (60-93 years) underwent routine clinical and laboratory evaluation, including assessment of cfPWV in eligible participants. Flow-cytometry was used to assess proportions of memory T-cells, CD28null T-cells, and CMV-specific T-cells. The following associations were examined; CMV serostatus/cfPWV, CMV serostatus/proportion of Tmem, proportion of Tmem/cfPWV, CD28null T-cells/cfPWV, and CMV-specific T-cells/cfPWV. Linear regression models were used to adjust for age, sex, socioeconomic status, smoking, waist-to-hip ratio, cholesterol, and blood pressure as required. Results: Statistically significant positive associations were found (P-values for the fully adjusted models are given); CMV serostatus/cfPWV in men (P ≤ 0.01) but not in women, CMV serostatus/proportions of CD4 Tmem in men (P ≤ 0.05) but not in women; proportions of CD4 Tmem/cfPWV among CMV seropositive (CMV+) people (P ≤ 0.05) but not CMV seronegative (CMV-) people. Conclusion: CMV infection increases the CVD risk of older men by increasing cfPWV. This may be mediated in part by increased proportions of CD4 Tmem, higher numbers of which are found in CMV+ older people and more so among men than women. Given the high prevalence of CMV worldwide, our findings point to a significant global health issue. Novel strategies to mitigate the increased CVD risk associated with CMV may be required.

Beyond bones: The relevance of variants of connective tissue (hypermobility) to fibromyalgia, ME/CFS and controversies surrounding diagnostic classification: an observational study.

BACKGROUND: Fibromyalgia and myalgic encephalomyelitis / chronic fatigue syndrome (ME/CFS) are poorly understood conditions with overlapping symptoms, fuelling debate as to whether they are manifestations of the same spectrum or separate entities. Both are associated with hypermobility, but this remains significantly undiagnosed, despite impact on quality of life. OBJECTIVE: We planned to understand the relevance of hypermobility to symptoms in fibromyalgia and ME/CFS. METHOD: Sixty-three patient participants presented with a confirmed diagnosis of fibromyalgia and/or ME/CFS; 24 participants were healthy controls. Patients were assessed for symptomatic hypermobility. RESULTS: Evaluations showed exceptional overlap in patients between fibromyalgia and ME/CFS, plus 81% met Brighton criteria for hypermobility syndrome (odds ratio 7.08) and 18% met 2017 hypermobile Ehlers-Danlos syndrome (hEDS) criteria. Hypermobility scores significantly predicted symptom levels. CONCLUSION: Symptomatic hypermobility is particularly relevant to fibromyalgia and ME/CFS, and our findings highlight high rates of mis-/underdiagnosis. These poorly understood conditions have a considerable impact on quality of life and our observations have implications for diagnosis and treatment targets.

Validity of the emotional intelligence scale for use in sport.

This study investigated the factorial validity of the 33-item self-rated Emotional Intelligence Scale (EIS: Schutte et al., 1998) for use with athletes. In stage 1, content validity of the EIS was assessed by a panel of experts (n = 9). Items were evaluated in terms of whether they assessed EI related to oneself and EI focused on others. Content validity further examined items in terms of awareness, regulation, and utilization of emotions. Content validity results indicated items describe 6-factors: appraisal of own emotions, regulation of own emotions, utilization of own emotions, optimism, social skills, and appraisal of others emotions. Results highlighted 13-items which make no direct reference to emotional experiences, and therefore, it is questionable whether such items should be retained. Stage 2 tested two competing models: a single factor model, which is the typical way researchers use the EIS and the 5-factor model (optimism was discarded as it become a single-item scale fiolliwng stage 1) identified in stage 1. Confirmatory factor analysis (CFA) results on EIS data from 1,681 athletes demonstrated unacceptable fit indices for the 33-item single factor model and acceptable fit indices for the 6-factor model. Data were re-analyzed after removing the 13-items lacking emotional content, and CFA results indicate partial support for single factor model, and further support for a five-factor model (optimism was discarded as a factor during item removal). Despite encouraging results for a reduced item version of the EIS, we suggest further validation work is needed. Key pointsGiven the inherent link between construct measurement and theory testing, it is imperative for researchers to pay close attention to measurement issues showed poor fit indices. The present study investigated a self-report emotional intelligence for use in sportResults indicate that a single-item model shows poor fit with acceptable fit indices for a 6-factor model.A revised 5-factor and 19-item model showed improved model fit.Despite encouraging results, we suggest further validation work is needed.

CD28 null CD4 T-cell expansions in autoimmune disease suggest a link with cytomegalovirus infection.

Immunosenescence is thought to contribute to the increase of autoimmune diseases in older people. Immunosenescence is often associated with the presence of an expanded population of CD4 T cells lacking expression of CD28 (CD28 null). These highly cytotoxic CD4 T cells were isolated from disease-affected tissues in patients with rheumatoid arthritis, systemic lupus erythematosus, multiple sclerosis, or other chronic inflammatory diseases and their numbers appeared to be linked to disease severity. However, we recently demonstrated that the common herpes virus, cytomegalovirus (CMV), not ageing, is the major driver of this subset of cytotoxic T cells. In this review, we discuss how CMV might potentiate and exacerbate autoimmune disease through the expansion of CD28 null CD4 T cells.

Impact of musculoskeletal symptoms on physical functioning and quality of life among treated people with HIV in high and low resource settings: A case study of the UK and Zambia.

BACKGROUND: Musculoskeletal symptoms in people living with HIV (PLWH) such as pain, joint stiffness, and fatigue are commonly reported. Prevalence rates of up to 45%, 79% and 88% respectively have been reported. However, very little is known about differences in prevalence and impact of musculoskeletal symptoms on physical functioning and quality of life of PLWH on effective combined antiretroviral treatment (cART) in high and low-resource settings. METHODS: A cross-sectional study of PLWH on effective cART enrolled from two large urban clinics in the UK and Zambia was conducted in 2016. Eligible participants had no history of trauma to the joints within 4 weeks of recruitment, or documented evidence of previous rheumatic disease. Current musculoskeletal symptoms, functional ability, and health-related quality of life were evaluated using the health assessment (HAQ) and quality-of-life short form (SF-36) self-reported questionnaires. RESULTS: 214 patients were enrolled (108:UK and 106:Zambia). Participants from Zambia were younger (47 vs 44 years) and had significantly lower CD4 counts (640 vs 439 cells/mL p = 0.018) compared to those from the UK, while the UK group had lived with HIV longer (11 vs 6 years; p<0.001) and reported more comorbidities than the Zambian group (66% vs 26%; p<0.001). Musculoskeletal pain was common in both groups (UK:69% vs Zambia:61% p = 0.263) but no significant differences in physical functional capacity between the groups were observed. However, the UK group had significantly worse quality of life measurements (general health, vitality, mental health, emotional, and social functioning) associated with musculoskeletal symptoms compared to the Zambian group (p<0.001). CONCLUSIONS: Musculoskeletal symptoms in PLWH from both the UK and Zambia were common. PLWH in the UK reported worse quality of life measures associated with musculoskeletal symptoms compared to those in Zambia, suggesting that factors such as mental health, patient expectations and multimorbidity might play a role in determining well-being and quality of life of PLWH with musculoskeletal symptoms.

Secretion of IL-1ß From Monocytes in Gout Is Redox Independent.

The pro-inflammatory cytokine interleukin-1ß (IL-1ß) plays important roles in immunity but is also implicated in autoimmune disease. The most well-established mechanism of IL-1ß secretion is via activation of the NOD-like receptor family pyrin domain containing-3 (NLRP3) inflammasome which requires an initial priming signal followed by an activating signal. However, the precise mechanism by which the inflammasome is activated remains unclear. The role of reactive oxygen species (ROS) in this process is contradictory, with some studies suggesting that ROS are crucial while others describe opposite effects. In this study, we evaluated the effects of oxidative stress on IL-1ß secretion. Gout is a disease driven solely by IL-1ß secretion in response to monosodium urate (MSU) crystals which form during periods of hyperuricemia and thus presents an opportunity to study factors contributing to IL-1ß secretion. Sera and monocytes were isolated from patients with gout to determine whether differences in antioxidant status could explain the susceptibility of these individuals to gout attacks. In addition, sera and monocytes were collected from patients with chronic kidney disease (CKD) for comparison as this condition is associated with high levels of oxidative stress and disturbances in serum uric acid levels. There were differences in some aspects of antioxidant defenses in gout patients and these were mainly due to higher serum uric acid. Monocytes from gout patients were more responsive to priming, but not activation, of the NLRP3 inflammasome. However, expression of the components of the NLRP3 inflammasome were unaffected by priming or activation of the inflammasome, nor were these expression levels differentially regulated in gout patients. Inhibition of ROS by N-Acetyl Cysteine inhibited TLR2-induced priming of the NLRP3 inflammasome, but had no effect on MSU-induced activation. Together these findings demonstrate that oxidative stress only affects priming of the NLRP3 inflammasome but does not influence activation.

Precipitation of Soluble Uric Acid Is Necessary for In Vitro Activation of the NLRP3 Inflammasome in Primary Human Monocytes.

OBJECTIVE: To investigate the effects of soluble uric acid (UA) on expression and activation of the NOD-like receptor (NLR) pyrin domain containing protein 3 (NLRP3) inflammasome in human monocytes to elucidate the role of hyperuricemia in the pathogenesis of gout. METHODS: Primary human monocytes and the THP-1 human monocyte cell line were used to determine the effects of short- and longterm exposure to UA on activation of the NLRP3 inflammasome and subsequent interleukin 1ß (IL-1ß) secretion by ELISA and cell-based assays. Expression of key NLRP3 components in monocytes from patients with a history of gout were analyzed by quantitative PCR. RESULTS: Precipitation of UA was required for activation of the NLRP3 inflammasome and subsequent release of IL-1ß in human monocytes. Neither monosodium urate (MSU) crystals nor soluble UA had any effect on activation of the transcription factor, nuclear factor-κB. Prolonged exposure of monocytes to soluble UA did not alter these responses. However, both MSU crystals and soluble UA did result in a 2-fold increase in reactive oxygen species. Patients with gout (n = 15) had significantly elevated serum UA concentrations compared to healthy individuals (n = 16), yet secretion of IL-1ß and expression of NLRP3 inflammasome components in monocytes isolated from these patients were not different from those of healthy controls. CONCLUSION: Despite reports indicating that soluble UA can prime and activate the NLRP3 inflammasome in human peripheral blood mononuclear cells, precipitation of soluble UA into MSU crystals is essential for in vitro NLRP3 signaling in primary human monocytes.

CD28null pro-atherogenic CD4 T-cells explain the link between CMV infection and an increased risk of cardiovascular death.

An increased risk of cardiovascular death in Cytomegalovirus (CMV)-infected individuals remains unexplained, although it might partly result from the fact that CMV infection is closely associated with the accumulation of CD28null T-cells, in particular CD28null CD4 T-cells. These cells can directly damage endothelium and precipitate cardiovascular events. However, the current paradigm holds that the accumulation of CD28null T-cells is a normal consequence of aging, whereas the link between these T-cell populations and CMV infection is explained by the increased prevalence of this infection in older people. Resolving whether CMV infection or aging triggers CD28null T-cell expansions is of critical importance because, unlike aging, CMV infection can be treated. Methods: We used multi-color flow-cytometry, antigen-specific activation assays, and HLA-typing to dissect the contributions of CMV infection and aging to the accumulation of CD28null CD4 and CD8 T-cells in CMV+ and CMV- individuals aged 19 to 94 years. Linear/logistic regression was used to test the effect of sex, age, CMV infection, and HLA-type on CD28null T-cell frequencies. Results: The median frequencies of CD28null CD4 T-cells and CD28null CD8 T-cells were >12-fold (p=0.000) but only approximately 2-fold higher (p=0.000), respectively, in CMV+ (n=136) compared with CMV- individuals (n=106). The effect of CMV infection on these T-cell subsets was confirmed by linear regression. Unexpectedly, aging contributed only marginally to an increase in CD28null T-cell frequencies, and only in CMV+ individuals. Interestingly, the presence of HLA-DRB1*0301 led to an approximately 9-fold reduction of the risk of having CD28null CD4 T-cell expansions (OR=0.108, p=0.003). Over 75% of CMV-reactive CD4 T-cells were CD28null. Conclusion: CMV infection and HLA type are major risk factors for CD28null CD4 T-cell-associated cardiovascular pathology. Increased numbers of CD28null CD8 T-cells are also associated with CMV infection, but to a lesser extent. Aging, however, makes only a negligible contribution to the expansion of these T-cell subsets, and only in the presence of CMV infection. Our results open up new avenues for risk assessment, prevention, and treatment.