Elsevier

Journal of Autoimmunity

Volume 74, November 2016, Pages 182-193
Journal of Autoimmunity

Dysregulation of innate and adaptive serum mediators precedes systemic lupus erythematosus classification and improves prognostic accuracy of autoantibodies

https://doi.org/10.1016/j.jaut.2016.06.001Get rights and content

Highlights

  • Distinct T-helper type mediators are elevated >3.5 years before SLE classification.

  • Elevated levels of IL-4, IL-5, IL-6, and IFN-γ precede autoantibody positivity.

  • Immune dysregulation amplifies as patients approach SLE classification.

  • Models incorporating autoantibodies and immune mediators accurately predict SLE risk.

Abstract

Systemic lupus erythematosus (SLE) is a complex autoimmune disease with a poorly understood preclinical stage of immune dysregulation and symptom accrual. Accumulation of antinuclear autoantibody (ANA) specificities is a hallmark of impending clinical disease. Yet, many ANA-positive individuals remain healthy, suggesting that additional immune dysregulation underlies SLE pathogenesis. Indeed, we have recently demonstrated that interferon (IFN) pathways are dysregulated in preclinical SLE. To determine if other forms of immune dysregulation contribute to preclinical SLE pathogenesis, we measured SLE-associated autoantibodies and soluble mediators in samples from 84 individuals collected prior to SLE classification (average timespan = 5.98 years), compared to unaffected, healthy control samples matched by race, gender, age (±5 years), and time of sample procurement. We found that multiple soluble mediators, including interleukin (IL)-5, IL-6, and IFN-γ, were significantly elevated in cases compared to controls more than 3.5 years pre-classification, prior to or concurrent with autoantibody positivity. Additional mediators, including innate cytokines, IFN-associated chemokines, and soluble tumor necrosis factor (TNF) superfamily mediators increased longitudinally in cases approaching SLE classification, but not in controls. In particular, levels of B lymphocyte stimulator (BLyS) and a proliferation-inducing ligand (APRIL) were comparable in cases and controls until less than 10 months pre-classification. Over the entire pre-classification period, random forest models incorporating ANA and anti-Ro/SSA positivity with levels of IL-5, IL-6, and the IFN-γ-induced chemokine, MIG, distinguished future SLE patients with 92% (±1.8%) accuracy, compared to 78% accuracy utilizing ANA positivity alone. These data suggest that immune dysregulation involving multiple pathways contributes to SLE pathogenesis. Importantly, distinct immunological profiles are predictive for individuals who will develop clinical SLE and may be useful for delineating early pathogenesis, discovering therapeutic targets, and designing prevention trials.

Introduction

Systemic lupus erythematosus (SLE) is a clinically and serologically heterogeneous systemic autoimmune disease which causes significant morbidity and early mortality, especially in young women and minorities (1). Immune dysregulation in the form of pathogenic autoantibodies and chronic inflammation contributes to a wide range of clinical manifestations, including skin rashes, arthritis, and life-threatening renal and/or central nervous system damage [1]. A number of antinuclear autoantibody (ANA) specificities have been shown to accumulate prior to SLE classification [2], [3], [4]; preclinical use of hydroxychloroquine may abrogate autoantibody accumulation and delay clinical disease onset [4]. Early intervention is an attractive approach to SLE treatment. However, our understanding of pathogenic mechanisms in preclinical SLE is inadequate. Closing this knowledge gap would improve our ability to identify individuals with preclinical SLE, define windows of opportunity for early intervention, and facilitate the development of pathway-targeted treatments.

Current biomarkers in preclinical SLE have limited utility for forecasting the transition to classified disease [2], [3], [5]. Although SLE-associated autoantibody specificities such as anti-dsDNA, anti-spliceosome and anti-Ro/SSA, accumulate in SLE patients years before classification [3], their presence is not sufficient to predict SLE. ANAs are also found in sera from patients with other systemic rheumatic diseases [6], and from healthy individuals who do not go on to develop SLE, including some unaffected family members of SLE patients [7], and up to 14% of the general population [8]. Because individuals may remain healthy despite being ANA-positive, ANA positivity alone is likely not the sole pathogenic driver of SLE [2], [9], [10]. In addition to ANA positivity, the dysregulation of various immune pathways driven by soluble mediators may contribute to the development of clinical disease. High expression of type I interferon (IFN)-related genes has been associated with SLE, yet an elevated IFN signature is not present in all patients [5]. Evidence stemming from lupus-like animal models and SLE patients suggests that breaks in tolerance leading to the activation and persistence of autoreactive B cells arise from amplified crosstalk between innate and adaptive immunity [11], [12]. Key mediators of such crosstalk, including Th-type cytokines IFN-γ (Th1), interleukin (IL)-4 and IL-5 (Th2), and IL-17 and IL-21 (Th17) facilitate lymphocyte recruitment to germinal centers [13], [14], [15] and pathogenic autoantibody production [16], [17] with the help of T-follicular helper (Tfh) cells [17]. We have recently demonstrated that type II IFN (IFN-γ) becomes elevated prior to and concurrent with the development of lupus-associated autoantibodies [18]. The tumor necrosis factor (TNF) superfamily member B lymphocyte stimulator (BLyS), secreted in response to type I and type II IFNs [19], [20], further supports and propagates autoantibody production as a survival factor for self-reactive B-lymphocytes [21]. In addition to driving the production of pathogenic autoantibodies, these mediators also contribute to inflammation associated with SLE disease flare [22] and organ damage [23]. Although these mediators contribute to SLE disease activity, their role in preclinical autoimmunity and transition to clinical disease are not well understood.

No single factor or mechanism is likely sufficient to explain the complexity and heterogeneity of SLE pathogenesis; thus a multivariate, longitudinal approach is warranted to delineate mechanisms of early disease pathogenesis and discern unique parameters that forecast SLE classification. In the current study, we leveraged longitudinal serum samples from the Department of Defense Serum Repository (DoDSR) to compare levels and determine temporal relationships between autoantibodies and immune mediators from multiple immune pathways in individuals who subsequently developed SLE compared to matched, healthy controls. Our findings shed light on potential mechanisms of early preclinical SLE immunopathogenesis, whereby dysregulation of immune mediators occurs prior to and concurrent with autoantibody accumulation, and is amplified leading up to SLE classification. Further, this study informs the design of reliable and sensitive tools to predict SLE onset. Such tools can be used to identify high risk patients in need of rheumatology referral and enrollment in prospective, preclinical intervention studies, as well as inform the development of novel treatment strategies to avert or delay tissue damage that often accompanies transition to classified disease [24], [25], [26], [27].

Section snippets

Study population and serum samples

Experiments were performed in accordance with the Helsinki Declaration and approved by the Institutional Review Boards of the Oklahoma Medical Research Foundation and the Walter Reed National Military Medical Center. Samples were obtained from the DoDSR. Demographic and clinical information, including medication history and American College of Rheumatology (ACR) criteria for SLE classification, were extracted from medical records by study personnel. All patients with available serum samples

Innate and adaptive soluble immune mediators are dysregulated more than 3.5 years before SLE classification

Altered levels of multiple adaptive-type soluble mediators, including inflammatory Th1-, Th2-, and Th17-type cytokines, as well as innate and regulatory mediators, have been observed in established SLE [18], [40], [41]. To elucidate the possible involvement of soluble mediators in various stages of preclinical SLE pathogenesis, longitudinal changes in serum cytokine levels were compared in samples spanning pre- and post-classification time periods in cases and controls matched by demographics

Discussion

Deciphering immune dysregulation that contributes to early lupus pathogenesis is essential for efforts to thwart the development of tissue and organ damage and ensuing morbidity and early mortality associated with progression to clinical SLE. The goal of the current study was to expand and clarify our understanding of SLE pathogenesis prior to and concurrent with the development of clinical disease by determining the nature and temporal relationship of immune pathway dysregulation and the

Conclusions

In conclusion, data from our study delineate a complex and cumulative pathogenic process in preclinical SLE, involving a number risk factors and gradual dysregulation of innate and T-helper, adaptive immune pathways (Fig. 5). Abnormalities in multiple Th-type cytokines arise in early preclinical SLE pathogenesis and could be leveraged to identify individuals at highest risk of future SLE clinical onset with >90% accuracy. This study also describes multifactorial models that improve the

Acknowledgements

The authors thank the collaborating military rheumatologists for assistance in identifying cases and extracting clinical information; Tim Gross, Nicolas Dominguez, Susan Macwana, Jeannie Te, Jeannette Osban, Wendy Klein, Virginia Roberts, and Jourdan Anderson for technical assistance; Rebecka Bourn, PhD and Angela Andersen, PhD (Life Science Editors) for scientific editing. This work was supported by the National Institute of Allergy, Immunology and Infectious Diseases; National Institute of

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