Evaluating the conformation of recombinant domain I of β2-glycoprotein I and its interaction with human monoclonal antibodies

Pericleous, Charis; Miles, Jennifer A.; Esposito, Diego; Garza-Garcia, Acely; Driscoll, Paul C.; Lambrianides, Anastasia; Latchman, David S.; Isenberg, David; Rahman, Anisur; Ioannou, Yiannis; Giles, Ian

doi:10.1016/j.molimm.2011.07.024

Cited by 16 publications

(16 citation statements)

References 39 publications

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“…Polyclonal IgG from this patient has previously been shown to cause thrombosis in our mouse model [ 13 ]. Patient serum was fractionated into aDI-rich and aDI-poor fractions as follows: human recombinant hexahistidine-tagged DI was produced and purified as described previously [ 10 , 14 ]. Immobilized-nickel resin beads (Promega, Madison, WI, USA) were equilibrated with PBS (pH 7.4), followed by incubation with purified DI (50 μg DI/100 μl resin) for 30 min.…”

Section: Methodsmentioning

confidence: 99%

“…We previously demonstrated that human recombinant DI abrogated the thrombogenic ability of polyclonal human IgG purified from a patient with APS in our well-characterized in vivo mouse model of thrombotic APS [ 9 ], while a human monoclonal IgG aPL, IS4, both bound DI [ 10 ] and enhanced clot formation in the same model [ 11 , 12 ]. Critically, however, no study has shown that affinity-purified IgG aDI from patients with APS has a direct prothrombotic effect in vivo .…”

Section: Introductionmentioning

confidence: 99%

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Proof-of-concept study demonstrating the pathogenicity of affinity-purified IgG antibodies directed to domain I of 2-glycoprotein I in a mouse model of anti-phospholipid antibody-induced thrombosis

et al. 2014

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Objective. IgG aPL against domain I of β2-glycoprotein I (β2GPI) [anti-DI (aDI)] is associated with the pathogenesis of APS, an autoimmune disease defined by thrombosis and pregnancy morbidity. To date, however, no study has demonstrated direct pathogenicity of IgG aDI in vivo. In this proof-of-concept study, we designed a novel system to affinity purify polyclonal aDI aPL in order to assess its prothrombotic ability in a well-characterized mouse microcirculation model for APS.Methods. Two polyclonal IgG fractions were isolated from serum of a patient with APS, both with high aPL activity but differing in aDI activity (aDI-rich and aDI-poor). These IgG fractions were tested for their pathogenic ability in an in vivo mouse model of thrombosis. Male CD1 mice were injected intraperitoneally with either aDI-rich or aDI-poor IgG; as a control, IgG isolated from healthy serum was used. A pinch injury was applied to the right femoral vein and thrombus dynamics and tissue factor activity in isolated tissue were evaluated.Results. Both aDI-rich and aDI-poor IgG retained aCL and anti-β2GPI activity, while only aDI-rich IgG displayed high aDI activity, as defined by our in-house cut-offs for positivity in each assay. aDI-rich IgG induced significantly larger thrombi in vivo compared with aDI-poor IgG (P < 0.0001). Similarly, aDI-rich IgG significantly enhanced the procoagulant activity of carotid artery endothelium and peritoneal macrophages isolated from experimental animals (P < 0.01).Conclusion. These data directly demonstrate that the ability to cause thrombosis in vivo is concentrated in the aDI fraction of aPL.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Proof-of-concept study demonstrating the pathogenicity of affinity-purified IgG antibodies directed to domain I of 2-glycoprotein I in a mouse model of anti-phospholipid antibody-induced thrombosis

et al. 2014

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“…42 Subsequent cleavage with factor Xa removed the hexahistidine tag, resulting in native DI peptide.…”

Section: Methodsmentioning

confidence: 99%

Antiphospholipid antibodies enhance rat neonatal cardiomyocyte apoptosis in an in vitro hypoxia/reoxygenation injury model via p38 MAPK

et al. 2017

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A significant amount of myocardial damage during a myocardial infarction (MI) occurs during the reperfusion stage, termed ischaemia/reperfusion (I/R) injury, and accounts for up to 50% of total infarcted tissue post-MI. During the reperfusion phase, a complex interplay of multiple pathways and mechanisms is activated, which ultimately leads to cell death, primarily through apoptosis. There is some evidence from a lupus mouse model that lupus IgG, specifically the antiphospholipid (aPL) antibody subset, is pathogenic in mesenteric I/R injury. Furthermore, it has previously been shown that the immunodominant epitope for the majority of circulating pathogenic aPLs resides in the N-terminal domain I (DI) of beta-2 glycoprotein I (β2GPI). This study describes the enhanced pathogenic effect of purified IgG derived from patients with lupus and/or the antiphospholipid syndrome in a cardiomyocyte H/R in vitro model. Furthermore, we have demonstrated a pathogenic role for aPL containing samples, mediated via aPL–β2GPI interactions, resulting in activation of the pro-apoptotic p38 MAPK pathway. This was shown to be inhibited using a recombinant human peptide of domain I of β2GPI in the fluid phase, suggesting that the pathogenic anti-β2GPI antibodies in this in vitro model target this domain.

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“…The anti-DI antibodies were measured [40] in the same manner as human 2GPI, except the plates were coated with bacterially-expressed, conformationally correct human recombinant DI, produced by bacterial expression at University College London [41,42]. Commercially sourced positive control sera (Louisville APL Diagnostics, USA) for IgG and IgM anti-2GPI antibodies were used as calibrators for anti-DI assay.…”

Section: Elisa For Detecting Igg and Igm Anti-dimentioning

confidence: 99%

The association between IgG and IgM antibodies against cardiolipin, β2-glycoprotein I and Domain I of β2-glycoprotein I with disease profile in patients with multiple sclerosis

Filippidou

Krashias

Pericleous

et al. 2016

Molecular Immunology

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Antiphospholipid antibodies (aPL) occur in patients with multiple sclerosis (MS) with a number of studies reporting elevated levels; their exact prevalence and pathogenic role remain unclear. Epidemiological studies associate MS with an increased risk of deep venous thromboembolism and stroke; overlapping clinical features with APS. Antibodies against the first domain - Domain I (DI) - of β2glycoprotein I (β2GPI), show the most clinical significance and evidence for pathogenicity in the antiphospholipid syndrome (APS), but have not yet been investigated in MS. Serum from a well-defined cohort of 127 MS patients and 92 healthy controls were tested for IgM and IgG antibodies against cardiolipin (CL), β2GPI and DI. Higher frequency of IgM and IgG anti-CL were found in MS patients (18.1% and 21.3%), compared to controls (1.1% in both cases, p<0.0001). We report that anti-DI antibodies were associated with MS patients, with 6.3% and 7.1% positive for IgM and IgG, respectively, compared to controls, 1.1% (p<0.05). IgM anti-CL antibodies were elevated in secondary progressive MS and primary progressive MS compared to relapse-remitting MS, (p<0.005). This study enrolled the largest number of patients with definite MS for studying the association with aPL. Although we confirmed IgM and IgG anti-CL antibodies occur in patients with MS, this is the first study that identified anti-DI antibodies in MS patients. This new finding may prove valuable and future studies are required to evaluate its role as a potential risk factor of thromboembolic phenomena in MS.

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Evaluating the conformation of recombinant domain I of β2-glycoprotein I and its interaction with human monoclonal antibodies

Cited by 16 publications

References 39 publications

Proof-of-concept study demonstrating the pathogenicity of affinity-purified IgG antibodies directed to domain I of 2-glycoprotein I in a mouse model of anti-phospholipid antibody-induced thrombosis

Proof-of-concept study demonstrating the pathogenicity of affinity-purified IgG antibodies directed to domain I of 2-glycoprotein I in a mouse model of anti-phospholipid antibody-induced thrombosis

Antiphospholipid antibodies enhance rat neonatal cardiomyocyte apoptosis in an in vitro hypoxia/reoxygenation injury model via p38 MAPK

The association between IgG and IgM antibodies against cardiolipin, β2-glycoprotein I and Domain I of β2-glycoprotein I with disease profile in patients with multiple sclerosis

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