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Protein microarray analysis reveals BAFF-binding autoantibodies in systemic lupus erythematosus.

ABSTRACT: Autoantibodies against cytokines, chemokines, and growth factors inhibit normal immunity and are implicated in inflammatory autoimmune disease and diseases of immune deficiency. In an effort to evaluate serum from autoimmune and immunodeficient patients for Abs against cytokines, chemokines, and growth factors in a high-throughput and unbiased manner, we constructed a multiplex protein microarray for detection of serum factor-binding Abs and used the microarray to detect autoantibody targets in SLE. We designed a nitrocellulose-surface microarray containing human cytokines, chemokines, and other circulating proteins and demonstrated that the array permitted specific detection of serum factor-binding probes. We used the arrays to detect previously described autoantibodies against cytokines in samples from individuals with autoimmune polyendocrine syndrome type 1 and chronic mycobacterial infection. Serum profiling from individuals with SLE revealed that among several targets, elevated IgG autoantibody reactivity to B cell-activating factor (BAFF) was associated with SLE compared with control samples. BAFF reactivity correlated with the severity of disease-associated features, including IFN-?-driven SLE pathology. Our results showed that serum factor protein microarrays facilitate detection of autoantibody reactivity to serum factors in human samples and that BAFF-reactive autoantibodies may be associated with an elevated inflammatory disease state within the spectrum of SLE.

SUBMITTER: Price JV

PROVIDER: S-EPMC3859403 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature

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Protein microarray analysis reveals BAFF-binding autoantibodies in systemic lupus erythematosus.

Price Jordan V JV Haddon David J DJ Kemmer Dodge D Delepine Guillaume G Mandelbaum Gil G Jarrell Justin A JA Gupta Rohit R Balboni Imelda I Chakravarty Eliza F EF Sokolove Jeremy J Shum Anthony K AK Anderson Mark S MS Cheng Mickie H MH Robinson William H WH Browne Sarah K SK Holland Steven M SM Baechler Emily C EC Utz Paul J PJ

The Journal of clinical investigation 20131125 12

Autoantibodies against cytokines, chemokines, and growth factors inhibit normal immunity and are implicated in inflammatory autoimmune disease and diseases of immune deficiency. In an effort to evaluate serum from autoimmune and immunodeficient patients for Abs against cytokines, chemokines, and growth factors in a high-throughput and unbiased manner, we constructed a multiplex protein microarray for detection of serum factor-binding Abs and used the microarray to detect autoantibody targets in ...[more]

PMID: 24270423

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Project description:The mitochondrion supplies energy to the cell and regulates apoptosis. Unlike other mammalian organelles, mitochondria are formed by binary fission and cannot be directly produced by the cell. They contain numerous copies of a compact circular genome that encodes RNA molecules and proteins involved in mitochondrial oxidative phosphorylation. Whereas, mitochondrial DNA (mtDNA) activates the innate immune system if present in the cytosol or the extracellular milieu, it is also the target of circulating autoantibodies in systemic lupus erythematosus (SLE). However, it is not known whether mitochondrial RNA is also recognized by autoantibodies in SLE. In the present study, we evaluated the presence of autoantibodies targeting mitochondrial RNA (AmtRNA) in SLE. We quantified AmtRNA in an inducible model of murine SLE. The AmtRNA were also determined in SLE patients and healthy volunteers. AmtRNA titers were measured in both our induced model of murine SLE and in human SLE, and biostatistical analyses were performed to determine whether the presence and/or levels of AmtRNA were associated with clinical features expressed by SLE patients. Both IgG and IgM classes of AmtRNA were increased in SLE patients (n = 86) compared to healthy controls (n = 30) (p < 0.0001 and p = 0.0493, respectively). AmtRNA IgG levels correlated with anti-mtDNA-IgG titers (r s = 0.54, p < 0.0001) as well as with both IgG and IgM against β-2-glycoprotein I (anti-β2GPI; r s = 0.22, p = 0.05), and AmtRNA-IgG antibodies were present at higher levels when patients were positive for autoantibodies to double-stranded-genomic DNA (p < 0.0001). AmtRNA-IgG were able to specifically discriminate SLE patients from healthy controls, and were negatively associated with plaque formation (p = 0.04) and lupus nephritis (p = 0.03). Conversely, AmtRNA-IgM titers correlated with those of anti-β2GPI-IgM (r s = 0.48, p < 0.0001). AmtRNA-IgM were higher when patients were positive for anticardiolipin antibodies (aCL-IgG: p = 0.01; aCL-IgM: p = 0.002), but AmtRNA-IgM were not associated with any of the clinical manifestations assessed. These findings identify mtRNA as a novel mitochondrial antigen target in SLE, and support the concept that mitochondria may provide an important source of circulating autoantigens in SLE.

Project description:ObjectiveTo determine whether systemic lupus erythematosus (SLE) can develop in the absence of BAFF in an SLE-prone host.MethodsStarting with C57BL/6 mice that express a human BCL2 transgene (Tg) in their B cells (thereby rendering B cell survival largely independent of BAFF-triggered signals), we introgressed this Tg into NZM 2328 mice genetically deficient in BAFF (NZM.Baff-/- ) to generate NZM.Baff-/- .Bcl2Tg mice. Expression of human Bcl-2 and lymphocyte profiles were assessed by fluorescence-activated cell sorting, and serologic profiles were determined by enzyme-linked immunosorbent assay. Immunofluorescence and histologic analyses were performed to assess renal immunopathologic features in the mice, and clinical disease was assessed according to the outcomes of severe proteinuria and death.ResultsIn comparison to their non-Tg NZM.Baff-/- littermates (n ≥ 7), NZM.Baff-/- .Bcl2Tg mice (n ≥ 8) overexpressed Bcl-2 in their B cells and developed significantly increased percentages and numbers of B cells and plasma cells, serum levels of IgG autoantibodies, glomerular deposition of IgG and C3, and severity of glomerular and tubulointerstitial inflammation, culminating in severe proteinuria and death (all P < 0.05 versus NZM.Baff-/- littermates). The time course for development of SLE-like features in NZM.Baff-/- .Bcl2Tg mice was more rapid than has been previously observed in NZM 2328 wild-type mice (median age at death 4.5 months versus 7.5 months). NZM.Baff-/- .Bcl2Tg mice remained responsive to BAFF, since reintroduction of the Baff gene into these mice further accelerated the course of disease (median age at death 3 months).ConclusionThe role of BAFF in the development of SLE-like disease may be dispensable as long as B cell survival is preserved via a BAFF-independent pathway. This may help explain the limited and variable clinical success with BAFF antagonists in human SLE. Thus, NZM.Baff-/- .Bcl2Tg mice may serve as a powerful murine model for the study of BAFF-independent SLE.

Project description:ObjectiveAnticytokine autoantibodies occur across a range of hematologic, pulmonary, and infectious diseases. However, systematic investigation of their presence and significance in autoimmune diseases is lacking. This study was undertaken to examine the distinct functions of anticytokine autoantibodies in patients with systemic lupus erythematosus (SLE) compared to patients with other rheumatic diseases and healthy controls.MethodsSerum samples from patients with SLE (n = 199), patients with primary Sjögren's syndrome (SS) (n = 150), patients with rheumatoid arthritis (RA) (n = 149), and healthy controls (n = 200) were screened for 24 anticytokine autoantibodies using a multiplex bead-based assay. To evaluate the biologic activity of anticytokine autoantibodies, their ability to block cytokine-induced signal transduction or protein expression was measured. RNA sequencing was performed on whole blood in a subset of healthy controls and patients with SLE.ResultsPatients with SLE and those with SS had a striking excess of autoantibodies against interferons and the interferon-responsive chemokine interferon-inducible protein 10 (IP-10). Only autoantibodies against type I interferon, interleukin-12 (IL-12), and IL-22 exhibited neutralizing activity. In SLE, the presence of anti-interferon-γ autoantibodies was correlated with more severe disease activity, higher levels of anti-double-stranded DNA antibodies, and elevated expression of interferon-α/β-inducible genes. Conversely, in SLE patients with blocking anti-interferon-α autoantibodies, the type I interferon gene expression signature was normalized. Anti-type III interferon autoantibodies (λ2, λ3) and anti-IP-10 autoantibodies were newly recognized in SLE patient serum, and autoantibodies against macrophage-colony stimulating factor, IL-4, IL-7, IL-17, and IL-22, none of which have been previously identified in rheumatic conditions, were discovered.ConclusionAnticytokine autoantibodies are associated with distinct patterns of disease in SLE, SS, and RA. Anti-interferon autoantibodies are overrepresented in patients with SLE and those with SS, and fall into distinct functional classes, with only a subset of anti-type I interferon antibodies exhibiting neutralizing activity. Anti-interferon-γ autoantibodies are correlated with increased disease activity and interferon-related gene expression, suggesting that such autoantibodies may contribute to the pathogenesis of SLE.

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Protein microarray analysis reveals BAFF-binding autoantibodies in systemic lupus erythematosus.

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Protein microarray analysis reveals BAFF-binding autoantibodies in systemic lupus erythematosus.

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