Session Type: Abstract Submissions (ACR)
Antiphospholipid antibodies have been shown to bind serine proteases (SP) involved in the coagulation cascade. Previously, we found that IgG anti-Factor(F)Xa antibody levels were higher in patients with the antiphospholipid syndrome (APS) and patients with systemic lupus erythematosus (SLE) but no APS compared with disease and healthy control (HC) groups. Given that FXa has important haemostatic and cellular effects we hypothesized that anti-FXa antibodies may be important in the pathogenesis of APS and SLE. Therefore, we investigated whether differences exist in the avidity and functional effects of APS and SLE anti-FXa IgG upon the coagulant and cellular functions of FXa.
IgG was purified from patients with APS (n=15) and SLE (n=15) who had medium or high serum levels of anti-FXa binding and HC (n=10) negative for anti-FXa IgG. The avidity of IgG-FXa binding was measured under chaotropic conditions using a NaCl gradient. We measured effects of anti-FXa IgG on FXa-activated clotting time (ACT) and on FXa enzymatic activity in a chromogenic assay in the absence and presence of antithrombin (AT)III. Cellular effects of IgG on FXa-protease-activated receptor (PAR) mediated intracellular calcium release in human umbilical vein endothelial cells (HUVEC) were measured using the fluorescent image plate reader (FLIPR).
All SLE-IgG displayed significantly lower (less than 25 % binding) avidity compared to APS-IgG (25-70% binding) to FXa at 0.13 to 1M concentrations of NaCl (p<0.05). The mean residual binding of APS-IgG to FXa was significantly higher than that of SLE-IgG below 2M NaCl (26 vs 13 %; p<0.05 at 1 M). FXa enzymatic activity was significantly reduced by APS-IgG (90%) and SLE-IgG (92 %) compared to HC-IgG (98 %) (APS vs HC p<0.05, SLE vs HC p<0.05, APS vs SLE p=0.04). ATIII mediated inhibition of FXa however, was significantly reduced by APS-IgG (62 %) compared with HC (79 %) and SLE (81 %) (p<0.05). The greatest prolongation of FXa-ACT was observed with APS-IgG followed by SLE-IgG and HC-IgG (74, 63 and 26 sec respectively). In cultured HUVEC, APS IgG caused significantly greater enhancement of FXa-mediated Ca2+ flux compared to SLE and HC-IgG at the same concentration (400 μg/ml) (APS vs HC p<0.05, APS vs SLE p=0.03) and this effect was reduced at lower IgG concentration (200 μg/ml).
FXa reactive IgG isolated from patients with APS displayed higher avidity binding to FXa and had greater functional effects upon FXa activity, ATIII mediated inhibition of FXa and FXa-PAR mediated signalling in cultured endothelial cells than SLE-IgG. Further work is now underway to further characterise the cellular effects of these anti-FXa IgG.
V. M. Ripoll,
D. A. Isenberg,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/factor-xa-reactive-antibodies-in-patients-with-systemic-lupus-erythematosus-and-antiphospholipid-syndrome-have-differential-effects-upon-coagulation-assays-and-endothelial-cells/