Autoimmune rheumatic disease in Australian Aboriginal and Torres Strait Islander Peoples: What do we know?

Autoimmune rheumatic disease (AIRD) is a collective term, which comprises a group of multisystem inflammatory autoimmune diseases, including connective tissue disease, chronic inflammatory arthritis, sarcoidosis and systemic vasculitis. Some AIRD are prevalent in the general population, and all can cause significant morbidity and reduced quality of life, with some increasing the risk of premature mortality, such as systemic lupus erythematosus (SLE), a connective tissue disease that is more prevalent and severe in Australian Aboriginal and Torres Strait Islander Peoples with high mortality rates. To ensure that management of AIRD can be optimised for all Australians, it is important that we understand the prevalence and potential phenotypic variations of AIRD across the Australian population. However, to date there have been few described cases of AIRD other than SLE in Aboriginal and Torres Strait Islander Peoples. In this review, we summarise what is known about AIRD other than SLE in Aboriginal and Torres Strait Islander Peoples, particularly with regards to prevalence, phenotype and disease outcomes, and highlight the current gaps in knowledge.

Primary aldosteronism: an unsuspected culprit of hypertension in systemic lupus erythematosus?

Primary aldosteronism (PA) is the most common endocrine cause of secondary hypertension and is associated with a high risk of cardiovascular disease in the general population. Patients suffering from systemic lupus erythematosus (SLE), a multisystem and multifactorial autoimmune disease, experience a high burden of hypertension and cardiovascular disease. Importantly, cardiovascular disease is one of the leading causes of death in SLE. Very limited evidence suggests an increased proportion of autoimmune diseases such as SLE in patients with PA. However, studies evaluating the prevalence of PA in the SLE population are lacking. Despite the potential for curative or targeted treatments, guidelines for the management of hypertension in SLE do not currently recommend testing for PA. This review highlights PA as a potentially over-looked secondary cause of hypertension in SLE, and offers future directions in research to improve the detection of this highly modifiable cardiovascular risk factor in the SLE population.

Patterns and prevalence of cognitive dysfunction in systemic lupus erythematosus.

OBJECTIVE: Systemic lupus erythematosus (SLE) is a chronic autoimmune disease, in which cognitive dysfunction is common, but poorly understood. This study aims to characterize the prevalence and patterns of cognitive dysfunction in SLE. METHOD: SLE patients (n = 95) and demographically matched healthy controls (n = 48) underwent cross-sectional cognitive testing using the 1-hr conventional neuropsychological test battery recommended by the American College of Rheumatology for use in SLE. We used standard deviations (SD) from the healthy control group to define impairment. For each cognitive test we compared SLE and control groups using independent samples t-tests (or alternatives when needed). We performed cluster analysis using a machine learning algorithm to look for patterns of cognitive dysfunction. RESULTS: The SLE group performed significantly worse than healthy controls on every cognitive test. The largest differences were in the domains of verbal fluency, working memory and attention, while fine motor and psychomotor speed were the least affected domains. As expected, the prevalence of cognitive dysfunction varied depending on the SD cut-off used, with 49% of participants being >1.5 SD below the healthy control mean in at least two cognitive domains. Heat mapping showed variability in the pattern of dysfunction between individual patients and cluster analysis confirmed the presence of two clusters of patients, which were those significantly impaired versus those having preserved cognition. CONCLUSIONS: Cognitive dysfunction is common in SLE but markedly heterogeneous across both cognitive domains and across the SLE group. Cluster analysis supports the use of a binary definition of cognitive dysfunction in SLE.

Circulating Interleukin-37 Levels in Healthy Adult Humans - Establishing a Reference Range.

Interleukin (IL)-37 has an important function in limiting excessive inflammation. Its expression is increased in numerous inflammatory and autoimmune conditions and correlates with disease activity, suggesting it could have potential as a disease biomarker. Nevertheless, a reference range has yet to be determined. Our aim was to establish the first reference range of circulating IL-37 levels in healthy adult humans. PubMed was searched for studies reporting blood IL-37 concentrations in healthy adult subjects as measured by enzyme-linked immunosorbent assay. Nineteen studies were included in the analysis. Mean IL-37 levels were weighted by sample sizes, and weighted mean lower and upper levels ( ± 2SD of means) were calculated to provide a weighted mean and reference range. IL-37 levels were quantified in either serum or plasma from a total of 1035 (647 serum; 388 plasma) healthy subjects. The serum, plasma and combined matrix weighted means (reference ranges) were 72.9 (41.5 - 104.4) pg/mL, 83.9 (41.1 - 126.8) pg/mL, and 77.1 (41.4 - 112.8) pg/mL, respectively. There were no significant differences between serum and plasma means and upper and lower limits. Study means and upper IL-37 levels were significantly higher in Chinese population studies. From our analysis, a preliminary reference range for circulating IL-37 levels in healthy human adults has been established. In order to determine a reliable reference range for clinical application, large, prospective, multi-ethnic, healthy population studies are necessary. In addition, demographics, sample matrix, collection, processing and storage methods potentially affecting IL-37 detection levels should be thoroughly investigated.

Granulocyte colony-stimulating factor is not pathogenic in lupus nephritis.

Systemic lupus erythematosus (lupus) is an autoimmune disease characterized by autoantibodies that form immune complexes with self-antigens, which deposit in various tissues, leading to inflammation and disease. The etiology of disease is complex and still not completely elucidated. Dysregulated inflammation is an important disease feature, and the mainstay of lupus treatment still utilizes nonspecific anti-inflammatory drugs. Granulocyte colony-stimulating factor (G-CSF) is a growth, survival, and activation factor for neutrophils and a mobilizer of hematopoietic stem cells, both of which underlie inflammatory responses in lupus. To determine whether G-CSF has a causal role in lupus, we genetically deleted G-CSF from Lyn-deficient mice, an experimental model of lupus nephritis. Lyn-/- G-CSF-/- mice displayed many of the inflammatory features of Lyn-deficient mice; however, they had reduced bone marrow and tissue neutrophils, consistent with G-CSF's role in neutrophil development. Unexpectedly, in comparison to aged Lyn-deficient mice, matched Lyn-/- G-CSF-/- mice maintained neutrophil hyperactivation and exhibited exacerbated numbers of effector memory T cells, augmented autoantibody titers, and worsened lupus nephritis. In humans, serum G-CSF levels were not elevated in patients with lupus or with active renal disease. Thus, these studies suggest that G-CSF is not pathogenic in lupus, and therefore G-CSF blockade is an unsuitable therapeutic avenue.

GILZ Regulates the Expression of Pro-Inflammatory Cytokines and Protects Against End-Organ Damage in a Model of Lupus.

Glucocorticoid-induced leucine zipper (GILZ) mimics many of the anti-inflammatory effects of glucocorticoids, suggesting it as a point of therapeutic intervention that could bypass GC adverse effects. We previously reported that GILZ down-regulation is a feature of human SLE, and loss of GILZ permits the development of autoantibodies and lupus-like autoimmunity in mice. To further query the contribution of GILZ to protection against autoimmune inflammation, we studied the development of the lupus phenotype in Lyn-deficient (Lyn-/-) mice in which GILZ expression was genetically ablated. In Lyn-/- mice, splenomegaly, glomerulonephritis, anti-dsDNA antibody titres and cytokine expression were exacerbated by GILZ deficiency, while other autoantibody titres and glomerular immune complex deposition were unaffected. Likewise, in patients with SLE, GILZ was inversely correlated with IL23A, and in SLE patients not taking glucocorticoids, GILZ was also inversely correlated with BAFF and IL18. This suggests that at the onset of autoimmunity, GILZ protects against tissue injury by modulating pro-inflammatory pathways, downstream of antibodies, to regulate the cycle of inflammation in SLE.

Associations of serum soluble Fas and Fas ligand (FasL) with outcomes in systemic lupus erythematosus.

OBJECTIVE: Fas/Fas ligand (FasL) and B cell-activating factor (BAFF) signalling have pivotal roles in SLE pathogenesis. We investigated the clinical associations of serum concentrations of soluble Fas (sFas) and soluble FasL (sFasL) in SLE and their relationship with BAFF. METHODS: Serum sFas and sFasL were quantified by multiplex assay, and BAFF by ELISA, in 118 patients with SLE and 17 healthy controls (HC). SLE disease activity and organ damage were assessed using the Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) and the Systemic Lupus International Collaborating Clinics Damage Index. RESULTS: sFas, sFasL and BAFF were detectable in all samples. Serum sFas and sFasL were significantly higher in SLE compared with HC. In univariable regression analyses, patients with active renal disease and those with flare had significantly higher levels of sFas compared with those without. High serum BAFF in patients with SLE was associated with increased sFas but not sFasL. The association between sFas and renal disease remained significant after adjusting for BAFF, but the association with flare attenuated. High sFas levels were associated with increased time-adjusted mean SLEDAI-2K, even after adjusting for BAFF, and with higher odds of flare over time. In contrast, high sFasL was associated with reduced organ damage over time. Serum sFasL/sFas ratio was negatively associated with active overall disease, flare and organ damage. CONCLUSIONS: Serum sFas is associated with active renal SLE, and active disease and flare over time, while sFasL/sFas ratio is negatively associated with disease activity and organ damage accrual. Treatments correcting abnormal levels of sFas/FasL may be worthy of evaluation in SLE.

Measuring MIF in Biological Fluids.

MIF is a key regulator of host immune responses and increased levels secreted from cells, or found circulating systemically, have been implicated in the pathogenesis of many inflammatory and autoimmune disorders. Here, we describe methods for detecting and quantifying extracellular concentrations of MIF in both human- and murine-derived biological samples.

Novel Methods of Incorporating Time in Longitudinal Multivariate Analysis Reveals Hidden Associations With Disease Activity in Systemic Lupus Erythematosus.

Objective: Systemic lupus erythematosus (SLE) is a multisystem autoimmune disease. SLE is characterized by high inter-patient variability, including fluctuations over time, a factor which most biomarker studies omit from consideration. We investigated relationships between disease activity and biomarker expression in SLE, using novel methods to control for time-dependent variability, in a proof-of-concept study to evaluate whether doing so revealed additional information. Methods: We measured 4 serum biomarkers (MIF, CCL2, CCL19, and CXCL10) and 13 routine clinical laboratory parameters, alongside disease activity measured by the SLE disease activity index-2k (SLEDAI-2k), collected longitudinally. We analyzed these data with unsupervised learning methods via ensemble clustering, incorporating temporal relationships using dynamic time warping for distance metric calculation. Results: Data from 843 visits in 110 patients (median age 47, 83% female) demonstrated highly heterogeneous time-dependent relationships between disease activity and biomarkers. Unbiased magnitude-based hierarchical clustering of biomarker expression levels isolated a patient subset (n = 9) with distinctively heterogeneous expression of the 17 biological parameters, and who had MIF, CCL2, CCL19, and CXCL10 levels that were higher and more strongly associated with disease activity, based on leave-one-out cross-validated regression analysis. In the remaining subgroup, a time-dependent regression model revealed significantly stronger predictive power of biomarkers for disease activity, compared to a time-agnostic regression model. Despite no significant difference in simple magnitude, using dynamic time warping analysis to align longitudinal profiles revealed a large subset (n = 69) with significantly stronger associations between biological parameters and disease activity. This subgroup had significantly lower flare rates, disease activity and damage scores, suggesting this clustering is clinically meaningful. Conclusions: These results suggest associations between biological parameters and disease activity in SLE exist in a multi-dimensional time-dependent pattern, with implications for the analysis of biomarkers in SLE often used to identify therapeutic targets. Novel methods to analyse high-dimensional data and control for time-dependent variability may have broad utility in the study complex relationships between clinical and biological parameters.

Analysis of serum B cell-activating factor from the tumor necrosis factor family (BAFF) and its soluble receptors in systemic lupus erythematosus.

OBJECTIVES: To determine the presence and clinical associations of the soluble receptors of B cell-activating factor from the tumor necrosis factor family (BAFF) in serum of patients with systemic lupus erythematosus (SLE). METHODS: Serum BAFF and soluble BAFF receptor (sBAFF-R) were quantified using ELISA, and soluble B cell maturation antigen (sBCMA) and transmembrane activator and cyclophilin ligand interactor (sTACI) by Luminex, in 87 SLE patients and 17 healthy controls (HC). Disease activity and organ damage were assessed using SLE Disease Activity Index 2000 (SLEDAI-2K) and Systemic Lupus International Collaborating Clinics (SLICC) SLE Damage Index (SDI), respectively. RESULTS: BAFF and all receptors were detectable in all serum samples. Serum sBCMA and sTACI, but not sBAFF-R, were significantly higher in SLE than in HC. Serum BAFF was also increased in SLE, but this association was attenuated after adjusting for age and ethnicity. Increased serum BAFF was associated with flare and organ damage. Increased serum sBCMA was associated with the presence of anti-dsDNA, but not with overall or organ-specific disease activity, flare or organ damage. Neither sTACI nor sBAFF-R was associated with any SLE clinical parameters in multivariable analysis. While serum BAFF correlated negatively with sBAFF-R in HC, no statistically significant correlations were observed between BAFF and its receptors in SLE patients. CONCLUSION: Serum BAFF was associated with flare and organ damage independent of the presence of its soluble receptors. While sBCMA was associated with anti-dsDNA positivity, other soluble BAFF receptors were not associated with SLE clinical indicators.

Analysis of serum interleukin(IL)-1α, IL-1ß and IL-18 in patients with systemic sclerosis.

OBJECTIVES: Systemic sclerosis (SSc) is an autoimmune disease characterised by fibrosis, vascular dysfunction and immune dysregulation. The pathogenesis of SSc remains poorly understood, although studies have indicated a role for the innate immune response. METHODS: Here, we measured serum interleukin (IL)-1α, IL-1ß and IL-18 levels in 105 SSc patients and 47 healthy controls (HC) and analysed them with respect to multiple clinical parameters. RESULTS: Serum IL-18 concentrations were significantly higher in SSc patients than in HC, while no significant differences in concentrations of IL-1α and IL-1ß were observed between SSc and HC. In both SSc and HC serum, IL-1α and IL-1ß were positively correlated, while in SSc, both cytokines negatively correlated with IL-18. Serum IL-18 was significantly negatively correlated with both carbon monoxide transfer coefficient (KCO) and diffusing capacity of the lungs for carbon monoxide (DLCO). Serum IL-1ß was positively correlated with the modified Rodnan skin score (mRSS), particularly in patients with limited subtype. DLCO, KCO and tricuspid regurgitation (TR) velocity were significantly higher in patients with high serum IL-1ß. Serum IL-1α was significantly lower in SSc patients with low KCO and positively correlated with KCO. SSc patients with high serum IL-1α concentrations were more likely to have digital ulcers. CONCLUSIONS: Our data suggest that these IL-1 family cytokines may have different roles in the pathogenesis of SSc fibrotic complications.

Effect of storage duration on cytokine stability in human serum and plasma.

Quantification of analytes such as cytokines in serum samples is intrinsic to translational research in immune diseases. Optimising pre-analytical conditions is critical for ensuring study quality, including evaluation of cytokine stability. We aimed to evaluate the effect on cytokine stability of storage duration prior to freezing of serum, and compare to plasma samples obtained from patients with systemic lupus erythematosus (SLE). Protein stability was analysed by simultaneously quantifying 18 analytes using a custom multi-analyte profile in SLE patient serum and plasma samples that had been prospectively stored at 4 °C for pre-determined periods between 0 and 30 days, prior to freezing. Six analytes were excluded from analysis, because most tested samples were above or below the limit of detection. Amongst the 12 analysed proteins, 11 did not show significant signal degradation. Significant signal degradation was observed from the fourth day of storage for a single analyte, CCL19. Proteins levels were more stable in unseparated serum compared to plasma for most analytes, with the exception of IL-37 which appears slightly more stable in plasma. Based on this, a maximum 3 days of storage at 4 °C for unseparated serum samples is recommended for biobanked samples intended for cytokine analysis in studies of human immune disease.

Analysis of serum macrophage migration inhibitory factor and D-dopachrome tautomerase in systemic sclerosis.

OBJECTIVES: Macrophage migration inhibitory factor (MIF) and D-dopachrome tautomerase (DDT), members of the same cytokine superfamily, are linked to the pathogenesis of a number of inflammatory diseases. The aim of this study was to investigate their clinical relevance in systemic sclerosis (SSc). METHODS: Serum MIF and DDT were quantified in 105 SSc patients by ELISA and levels compared to healthy controls (HC) (47) and patients with systemic lupus erythematosus (SLE) (184). Clinical parameters included organ involvement, serum laboratory markers and results of pulmonary function tests, and overall disease activity assessed using the European Scleroderma Trials and Research group (EUSTAR) activity index. RESULTS: There was no significant difference in serum DDT concentrations between patients with SSc and HC. However, serum MIF was significantly increased in SSc compared to both HC and SLE cohorts. Serum MIF was increased in SSc patients with low forced vital capacity (FVC) and was also associated with the use of angiotensin II receptor blockers and beta blockers in SSc, confirmed after adjusting for the presence of systemic hypertension and low FVC. Serum DDT was significantly higher in SSc patients with low FEV1 and negatively correlated with EUSTAR score, particularly in patients with limited disease. CONCLUSION: Although not significantly linked to specific clinical parameters, serum MIF was significantly higher in SSc patients than in HC and SLE patients, suggesting a fundamental role for MIF in SSc. DDT, while closely related to MIF, did not show a similar expression profile, suggesting functional differences between these molecules.

Analysis of urinary macrophage migration inhibitory factor in systemic lupus erythematosus.

OBJECTIVE: To characterise the clinical relevance of urinary macrophage migration inhibitory factor (uMIF) concentrations in patients with systemic lupus erythematosus (SLE). METHODS: MIF, adjusted for urine creatinine, was quantified by ELISA in urine samples from 64 prospectively recruited patients with SLE. Serum MIF and urinary monocyte chemoattractant protein 1 (uMCP-1) were quantified by ELISA in a subset of patients (n = 39). Disease activity was assessed using the SLE Disease Activity Index-2000 (SLEDAI-2K) score. RESULTS: uMIF was detectable in all patients with SLE. uMIF was positively correlated with overall SLEDAI-2K, was significantly higher in patients with SLE with high disease activity (SLEDAI-2K≥10) compared with those with inactive disease (SLEDAI-2K<4), and this association remained significant after adjusting for ethnicity, flare and use of immunosuppressants. uMIF was also significantly higher in SLE patients with flare of disease, although not confirmed in multivariable analysis. No significant differences in uMIF levels were observed according to the presence of renal disease activity, as assessed by renal SLEDAI-2K or biopsy-confirmed lupus nephritis. In contrast, uMCP-1 was significantly higher in SLE patients with active renal disease. uMIF expression was not associated with irreversible organ damage accrual or glucocorticoid use. CONCLUSIONS: These data suggest uMIF as a potential overall but not renal-specific SLE biomarker, whereas uMCP-1 is a renal-specific SLE biomarker.

Analysis of Serum Interleukin (IL)-1ß and IL-18 in Systemic Lupus Erythematosus.

Systemic lupus erythematosus (SLE) is a chronic multisystem autoimmune disease characterized by biological and clinical heterogeneity. The interleukin (IL)-1 superfamily is a group of innate cytokines that contribute to pathogenesis in many autoimmune diseases. IL-1ß and IL-18 are two members that have been shown to play a role in murine lupus-like models, but their role in human SLE remains poorly understood. Here, IL-1ß and IL-18 were quantified by enzyme-linked immunosorbent assay in the serum of healthy controls (HCs) and SLE patients from a prospectively followed cohort. Disease activity and organ damage were assessed using SLE disease activity index 2000 (SLEDAI-2K) and SLE damage index scores (SDI), respectively. 184 SLE patients (mean age 44.9 years, 91% female, 56% double-stranded deoxyribonucleic acid positive) were compared to 52 HC. SLE patients had median [IQR] SLEDAI-2K of 4 [2,6], and SDI of 1 [0-2]. Serum IL-18 levels were statistically significantly higher in SLE patients compared to HCs. Univariable linear regression analyses showed that patients with active renal disease or irreversible organ damage had statistically significantly elevated serum IL-18 levels. The association between serum IL-18 and active renal disease was confirmed in multivariable analysis after adjusting for ethnicity and organ damage. High baseline serum IL-18 levels were associated with organ damage at the subsequent visit. Serum IL-1ß levels were not significantly elevated in SLE patients when compared to HCs and had no association with overall or organ-specific disease activity or organ damage in cross-sectional and longitudinal analyses. Our data suggest that serum IL-18 and IL-1ß have different clinical implications in SLE, with IL-18 being potentially associated with active renal disease.

BAFF-driven autoimmunity requires CD19 expression.

B cell activating factor of the tumor necrosis factor family (BAFF or BLyS) is a critical factor for B cell survival and maturation. BAFF-transgenic (BAFF-Tg) mice develop autoimmunity that resembles systemic lupus erythematosus (SLE) in a T cell-independent but MyD88-dependent manner, implicating toll-like receptor (TLR) signaling. The specific B cell subtypes that make pro-inflammatory autoantibodies in BAFF-Tg mice are TLR-activated innate B cells known as marginal zone (MZ) and B1 B cells. These cells infiltrate the salivary glands and kidneys of diseased BAFF-Tg mice. However, loss of B1a or MZ B cells does not protect BAFF-Tg mice against disease, suggesting that B1b B cells might be the important pathogenic B cell subset. To test this hypothesis, we have generated BAFF-Tg mice that retained follicular B cells, but are deficient in B1a, B1b and MZ B cells, by crossing BAFF-Tg mice to CD19-deficient mice (BTg-CD19(-/-)). The BTg-CD19(-/-) mice did not produce autoantibodies and were protected from splenomegaly, kidney pathology and all signs of autoimmunity. This work suggests that B1b B cells, rather than MZ or B1a B cells, are sufficient and possibly required for the development of autoimmunity. Loss of the majority of innate-like B cells was able to protect BAFF-Tg mice from developing disease, so we can now conclude that autoimmunity induced by excessive BAFF production requires B1b B cells and CD19 signaling.