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Proof-of-concept study demonstrating the pathogenicity of affinity-purified IgG antibodies directed to domain I of β2-glycoprotein I in a mouse model of anti-phospholipid antibody-induced thrombosis.

Pericleous C, Ruiz-Limón P, Romay-Penabad Z, Marín AC, Garza-Garcia A, Murfitt L, Driscoll PC, Latchman DS, Isenberg DA, Giles I, Ioannou Y, Rahman A, Pierangeli SS - Rheumatology (Oxford) (2014)

Bottom Line: A pinch injury was applied to the right femoral vein and thrombus dynamics and tissue factor activity in isolated tissue were evaluated.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).

View Article: PubMed Central - PubMed

Affiliation: Centre for Rheumatology Research, Division of Medicine, University College London, London, UK, Division of Rheumatology, Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX, USA, Structural Biology, Medical Research Council National Institute for Medical Research and Arthritis Research UK Centre for Adolescent Rheumatology, University College London Hospital and Great Ormond Street Hospital, London, UK.

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Binding properties of purified IgG fractions to cardiolipin, β2GPI and DIPurified IgG from unfractionated serum, aDI-rich and aDI-poor IgG from fractionated serum were tested at 100 µg/ml for (A) aCL, (B) aβ2GPI and (C) aDI activity in a dose-dependent manner. Results are expressed in IgG phospholipid units (GPLU), β2GPI units (GBU) and DI units (GDIU), respectively. The avidity of purified IgG (D) to CL, (E) β2GPI and (F) DI was determined by incubating IgG (at 100 µg/ml) with increasing NaCl concentrations (0.15 M original concentration in PBS, 0.25 M, 0.5 M, 1 M, 2 M and 4 M). Results are shown as a percentage of aPL activity retained, where 100% represents activity at 0.15 M NaCl. β2GPI: β2-glycoprotein I; DI: domain I; aDI: anti-DI; aβ2GPI: anti-β2GPI.
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keu360-F1: Binding properties of purified IgG fractions to cardiolipin, β2GPI and DIPurified IgG from unfractionated serum, aDI-rich and aDI-poor IgG from fractionated serum were tested at 100 µg/ml for (A) aCL, (B) aβ2GPI and (C) aDI activity in a dose-dependent manner. Results are expressed in IgG phospholipid units (GPLU), β2GPI units (GBU) and DI units (GDIU), respectively. The avidity of purified IgG (D) to CL, (E) β2GPI and (F) DI was determined by incubating IgG (at 100 µg/ml) with increasing NaCl concentrations (0.15 M original concentration in PBS, 0.25 M, 0.5 M, 1 M, 2 M and 4 M). Results are shown as a percentage of aPL activity retained, where 100% represents activity at 0.15 M NaCl. β2GPI: β2-glycoprotein I; DI: domain I; aDI: anti-DI; aβ2GPI: anti-β2GPI.

Mentions: aCL, aβ2GPI and aDI titres for APS serum were >96 GPLU, 93.9 GBU and 50.2 GDIU, respectively. As illustrated in Fig. 1, purified IgG (100 µg/ml) from unfractionated serum was highly positive in all three assays, with values >96 GPLU, >100 GBU and 89.4 GDIU. aDI-rich IgG maintained high activity against all three antigens (>96 GPLU, >100 GBU, >100 GDIU); as expected, aDI activity was enriched compared with unfractionated IgG. Conversely, aDI-poor IgG confirmed reduced binding to these antigens (89.3 GPLU, 46.8 GBU, 17 GDIU); the reduction in aDI activity was by far the greatest (Fig. 1A–C). Both aDI-rich and aDI-poor IgG retained aCL, aβ2GPI and aDI activity above the cut-off values for positivity in each assay. All samples from healthy controls were negative in all three assays.Fig. 1


Proof-of-concept study demonstrating the pathogenicity of affinity-purified IgG antibodies directed to domain I of β2-glycoprotein I in a mouse model of anti-phospholipid antibody-induced thrombosis.

Pericleous C, Ruiz-Limón P, Romay-Penabad Z, Marín AC, Garza-Garcia A, Murfitt L, Driscoll PC, Latchman DS, Isenberg DA, Giles I, Ioannou Y, Rahman A, Pierangeli SS - Rheumatology (Oxford) (2014)

Binding properties of purified IgG fractions to cardiolipin, β2GPI and DIPurified IgG from unfractionated serum, aDI-rich and aDI-poor IgG from fractionated serum were tested at 100 µg/ml for (A) aCL, (B) aβ2GPI and (C) aDI activity in a dose-dependent manner. Results are expressed in IgG phospholipid units (GPLU), β2GPI units (GBU) and DI units (GDIU), respectively. The avidity of purified IgG (D) to CL, (E) β2GPI and (F) DI was determined by incubating IgG (at 100 µg/ml) with increasing NaCl concentrations (0.15 M original concentration in PBS, 0.25 M, 0.5 M, 1 M, 2 M and 4 M). Results are shown as a percentage of aPL activity retained, where 100% represents activity at 0.15 M NaCl. β2GPI: β2-glycoprotein I; DI: domain I; aDI: anti-DI; aβ2GPI: anti-β2GPI.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4372675&req=5

keu360-F1: Binding properties of purified IgG fractions to cardiolipin, β2GPI and DIPurified IgG from unfractionated serum, aDI-rich and aDI-poor IgG from fractionated serum were tested at 100 µg/ml for (A) aCL, (B) aβ2GPI and (C) aDI activity in a dose-dependent manner. Results are expressed in IgG phospholipid units (GPLU), β2GPI units (GBU) and DI units (GDIU), respectively. The avidity of purified IgG (D) to CL, (E) β2GPI and (F) DI was determined by incubating IgG (at 100 µg/ml) with increasing NaCl concentrations (0.15 M original concentration in PBS, 0.25 M, 0.5 M, 1 M, 2 M and 4 M). Results are shown as a percentage of aPL activity retained, where 100% represents activity at 0.15 M NaCl. β2GPI: β2-glycoprotein I; DI: domain I; aDI: anti-DI; aβ2GPI: anti-β2GPI.
Mentions: aCL, aβ2GPI and aDI titres for APS serum were >96 GPLU, 93.9 GBU and 50.2 GDIU, respectively. As illustrated in Fig. 1, purified IgG (100 µg/ml) from unfractionated serum was highly positive in all three assays, with values >96 GPLU, >100 GBU and 89.4 GDIU. aDI-rich IgG maintained high activity against all three antigens (>96 GPLU, >100 GBU, >100 GDIU); as expected, aDI activity was enriched compared with unfractionated IgG. Conversely, aDI-poor IgG confirmed reduced binding to these antigens (89.3 GPLU, 46.8 GBU, 17 GDIU); the reduction in aDI activity was by far the greatest (Fig. 1A–C). Both aDI-rich and aDI-poor IgG retained aCL, aβ2GPI and aDI activity above the cut-off values for positivity in each assay. All samples from healthy controls were negative in all three assays.Fig. 1

Bottom Line: A pinch injury was applied to the right femoral vein and thrombus dynamics and tissue factor activity in isolated tissue were evaluated.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).

View Article: PubMed Central - PubMed

Affiliation: Centre for Rheumatology Research, Division of Medicine, University College London, London, UK, Division of Rheumatology, Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX, USA, Structural Biology, Medical Research Council National Institute for Medical Research and Arthritis Research UK Centre for Adolescent Rheumatology, University College London Hospital and Great Ormond Street Hospital, London, UK.

Show MeSH
Related in: MedlinePlus