Heparin-induced thrombocytopenia (HIT) is a serious adverse reaction to heparin, associated with increased risk of thromboembolic complications. Pathogenic antibodies form immunogenic complexes with PF4 and heparin, which activate platelets and leukocytes mainly through the low affinity IgG Fc receptor FcγRIIa. Intravenous immunoglobulins (IVIG) have been used as a therapeutic for HIT and are believed to alleviate thrombocytopenia and reduce thrombosis risk, possibly through competition with pathogenic IgG. Yet the anti-thrombotic effects of IVIG in HIT remain underexplored. The aim of our study was to investigate the effect of IVIG on thrombus formation in a disease model of HIT.

Microfluidic channels were coated with a confluent monolayer of human umbilical vein endothelial cells (HUVECs). Cells were primed with low-dose TNF-α to induce sub-thrombotic endothelial injury. Whole blood was preincubated with unfractionated heparin (UFH, 0.2 IU/mL), with or without IVIG, or IV.3 anti-FcγRIIa IgG, respectively. Monoclonal anti-PF4/heparin antibodies, or serum from thrombotic HIT patients, were introduced to the whole blood mixture. Blood was recalcified and perfused over HUVECs at a venous shear stress. Thrombus structure and growth dynamics were investigated by immunofluorescence microscopy.

Monoclonal HIT-IgG and HIT patient sera induced thrombus formation in presence of prophylactic dose heparin, only when endothelial cells were challenged with TNF-α. HIT thrombi were enriched with fibrin, phosphatidylserine, and leukocyte aggregations, differing markedly from the two-dimensional platelet-fibrin rich adhesions formed in absence of pathogenic HIT IgG. Pre-treatment of blood with IVIG or IV.3 fully inhibited the HIT thrombosis, showing no thrombus formation and improved flow rate through microfluidic channels. Deposition of fibrin and blood cells under inhibitory conditions resembled those observed in heparinized blood in absence of HIT-IgG.

We demonstrate that HIT-IgG induce thrombosis on minimally activated endothelial cells in a heparin-dependent manner. IVIG inhibited thrombus formation, improving blood flow through the microfluidic system. The FcγRIIa inhibitor achieved similar inhibition to competitive replacement of pathogenic IgG with IVIG. In conclusion, these results show that IVIG has a preventative effect in a model of HIT-thrombosis, similar to direct FcγRIIa inhibition.

The authors have no conflicts of interests to declare.