Objective. Free radical-mediated reactions have been implicated as contributors in a number of autoimmune diseases, including systemic lupus erythematosus (SLE). However, the potential for oxidative/ nitrosative stress to elicit an autoimmune response or to contribute to disease pathogenesis, and thus be useful when determining a prognosis, remains largely unexplored in humans. This study was undertaken to investigate the status and contribution of oxidative/ nitrosative stress in patients with SLE.Methods. Sera from 72 SLE patients with varying levels of disease activity according to the SLE Disease Activity Index (SLEDAI) and 36 age-and sex-matched healthy controls were evaluated for serum levels of oxidative/nitrosative stress markers, including antibodies to malondialdehyde (anti-MDA) protein adducts and to 4-hydroxynonenal (anti-HNE) protein adducts, MDA/HNE protein adducts, superoxide dismutase (SOD), nitrotyrosine (NT), and inducible nitric oxide synthase (iNOS).Results. Serum analysis showed significantly higher levels of both anti-MDA/anti-HNE protein adduct antibodies and MDA/HNE protein adducts in SLE patients compared with healthy controls. Interestingly, not only was there an increased number of subjects positive for anti-MDA or anti-HNE antibodies, but also the levels of both of these antibodies were statistically significantly higher among SLE patients whose SLEDAI scores were >6 as compared with SLE patients with lower SLEDAI scores (SLEDAI score <6). In addition, a significant correlation was observed between the levels of anti-MDA or anti-HNE antibodies and the SLEDAI score (r ‫؍‬ 0.734 and r ‫؍‬ 0.647, respectively), suggesting a possible causal relationship between these antibodies and SLE. Furthermore, sera from SLE patients had lower levels of SOD and higher levels of iNOS and NT compared with healthy control sera.Conclusion. These findings support an association between oxidative/nitrosative stress and SLE. The stronger response observed in serum samples from patients with higher SLEDAI scores suggests that markers of oxidative/nitrosative stress may be useful in evaluating the progression of SLE and in elucidating the mechanisms of disease pathogenesis.

Objective We sought to determine the effect of hydroxychloroquine therapy on the levels proinflammatory/prothrombotic markers and disease activity scores in patients with systemic lupus erythematosus (SLE) in a multiethnic, multi-center cohort (LUMINA). Methods Plasma/serum samples from SLE patients (n=35) were evaluated at baseline and after hydroxychloroquine treatment. Disease activity was assessed using SLAM-R scores. Interferon (IFN)-α2, interleukin (IL)-1β, IL-6, IL-8, inducible protein (IP)-10, monocyte chemotactic protein-1, tumor necrosis factor (TNF)-α and soluble CD40 ligand (sCD40L) levels were determined by a multiplex immunoassay. Anticardiolipin antibodies were evaluated using ELISA assays. Thirty-two frequency-matched plasma/serum samples from healthy donors were used as controls. Results Levels of IL-6, IP-10, sCD40L, IFN-α and TNF-α were significantly elevated in SLE patients versus controls. There was a positive but moderate correlation between SLAM-R scores at baseline and levels of IFN-α (p=0.0546). Hydroxychloroquine therapy resulted in a significant decrease in SLAM-R scores (p=0.0157), and the decrease in SLAM-R after hydroxychloroquine therapy strongly correlated with decreases in IFN-α (p=0.0087). Conclusions Hydroxychloroquine therapy resulted in significant clinical improvement in SLE patients, which strongly correlated with reductions in IFN-α levels. This indicates an important role for the inhibition of endogenous TLR activation in the action of hydroxychloroquine in SLE and provides additional evidence for the importance of type I interferons in the pathogenesis of SLE. This study underscores the use of hydroxychloroquine in the treatment of SLE.

Antiphospholipid (aPL)/anti-␤ 2 glycoprotein I (anti-␤ 2 GPI) antibodies stimulates tissue factor (TF) expression within vasculature and in blood cells, thereby leading to increased thrombosis. Several cellular receptors have been proposed to mediate these effects, but no convincing evidence for the involvement of a specific one has been provided. We investigated the role of Apolipoprotein E receptor 2 (ApoER2) on the pathogenic effects of a patient-derived polyclonal aPL IgG preparation (IgG-APS), a murine anti-␤ 2 GPI monoclonal antibody (E7) and of a constructed dimeric ␤ 2 GPI I (dimer), which in vitro mimics ␤ 2 GPI-antibody immune complexes, using an animal model of thrombosis, and ApoER2-deficient (؊/؊) mice. In wild type mice, IgG-APS, E7 and the dimer increased thrombus formation, carotid artery TF activity as well as peritoneal macrophage TF activity/expression. Those pathogenic effects were significantly reduced in ApoER2 (؊/؊) mice. In addition, those effects induced by the IgG-APS, by E7 and by the dimer were inhibited by treatment of wild-type mice with soluble binding domain 1 of ApoER2 (sBD1). Altogether these data show that ApoER2 is involved in pathogenesis of antiphospholipids antibodies. (Blood. 2011;117(4):1408-1414) IntroductionThe association between persistently present antiphospholipid (aPL) antibodies and the clinical manifestations of thrombosis or pregnancy morbidity is known as the antiphospholipid syndrome (APS). 1 aPL antibodies are heterogeneous and recognize a wide variety of plasma proteins with phospholipid-binding properties, such as prothrombin 2 and ␤ 2 glycoprotein I (␤ 2 GPI). 3,4 aPL antibodies directed against ␤ 2 GPI, a plasma protein without known physiologic function, are considered the most pathologically relevant antibodies.There is strong experimental evidence that anti-␤ 2 GPI antibodies have thrombogenic properties. In studies on endothelial cell activation, 5-8 authors have shown the induction of a prothrombotic and proinflammatory phenotype upon exposure to anti-␤ 2 GPI antibodies, indicated by expression of tissue factor (TF) and increased surface expression of adhesion molecules, such as intercellular adhesion molecule-1, vascular-cell adhesion molecule-1, and E-selectin. Activation of monocytes by anti-␤ 2 GPI antibodies leads to TF expression as well. 9 Furthermore, anti-␤ 2 GPI antibodies, or recombinant dimers of ␤ 2 GPI that mimic ␤ 2 GPI-antibody immune complexes, increase platelet deposition to extracellular matrix components in in vitro flow models. 10 Injection of anti-␤ 2 GPI antibodies in murine 11 or hamster 12 thrombosis models leads to increased thrombus formation.Several receptors were postulated to mediate the prothrombotic cellular effects of anti-␤ 2 GPI antibodies. The interaction between annexin A2 and ␤ 2 GPI-antibody immune complexes has been reported to lead to endothelial cell activation. 13 It seems unlikely, however, that annexin A2 is able to convey activation signals across the cell membrane because this phospholipid-binding protei...

Summary. Objectives: In the antiphospholipid syndrome (APS), the immunodominant epitope for the majority of circulating pathogenic antiphospholipid antibodies (aPLs) is the N-terminal domain I (DI) of b 2 -glycoprotein I. We have previously shown that recombinant DI inhibits the binding of aPLs in fluid phase to immobilized native antigen, and that this inhibition is greater with the DI(D8S/D9G) mutant and absent with the DI(R39S) mutant. Hence, we hypothesized that DI and DI(D8S/D9G) would inhibit aPL-induced pathogenicity in vivo. Methods: C57BL/6 mice (n = 5, each group) were injected with purified IgG derived from APS patients (IgG-APS, 500 lg) or IgG from normal healthy serum (IgG-NHS) and either recombinant DI, DI(R39S), DI(D8S/D9G), or an irrelevant control peptide (at 10-40 lg). Outcome variables measured were femoral vein thrombus dynamics in treated and control groups following standardized vessel injury, expression of vascular cell adhesion molecule-1 (VCAM-1) on the aortic endothelial surface, and tissue factor (TF) activity in murine macrophages. Results: IgG-APS significantly increased thrombus size as compared with IgG-NHS. The IgG-APS thrombus enhancement effect was abolished in mice pretreated with recombinant DI (P £ 0.0001) and DI(D8S/D9G) (P £ 0.0001), but not in those treated with DI(R39S) or control peptide. This inhibitory effect by DI was dose-dependent, and at lower doses DI(D8S/D9G) was a more potent inhibitor of thrombosis than wild-type DI (P £ 0.01). DI also inhibited IgG-APS induction of VCAM-1 on the aortic endothelial surface and TF production by murine macrophages. Conclusion: Our findings in this proof-of-concept study support the development of recombinant DI or the novel variant DI(D8S/D9G) as a potential future therapeutic agent for APS.

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