• We determined the location of gal3bp and gal3 and their role in promoting VT and leukocyte/endothelial cell interactions for the first time.• Gal3bp and gal3 have the potential to be used as targets for future VT therapies.Galectin-3-binding protein (gal3bp) and its receptor/ligand, galectin-3 (gal3), are secreted proteins that initiate signaling cascades in several diseases, and recent human proteomic data suggest they may play a role in venous thrombosis (VT). We hypothesized that gal3bp and gal3 may promote VT. Using a mouse stasis model of VT, we found that gal3bp and gal3 were localized on vein wall, red blood cells, platelets, and microparticles, whereas leukocytes expressed gal3 only. Gal3 was dramatically increased during early VT and gal3bp:gal3 colocalized in the leukocyte/endothelial cell interface, where leukocytes were partially attached to the vein wall. Thrombus size correlated with elevated gal3 and interleukin-6 (IL-6) vein wall levels. Recombinant gal3 promoted VT and increased vein wall IL-6 mRNA. Although recombinant gal3 restored the VT size in gal3 2/2 mice, it had no effect on IL6 2/2 mice, suggesting that gal3:gal3bp promotes VT through IL-6. Moreover, significantly fewer activated neutrophils were present in the gal3 2/2 vein walls. In a group of human patients, elevated circulating gal3bp correlated with acute VT. In conclusion, gal3bp:gal3 play a critical role in VT, likely via IL-6 and PMN-mediated thrombotic mechanisms, and may be a potential biomarker in human VT. (Blood. 2015;125(11):1813-1821

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Galectin-3 binding protein (Gal-3BP) was detected in procoagulant microparticles from mice and patients with venous thrombosis (VT), in prior studies from our laboratory. However, the mechanistic role of Gal-3BP and Galactin-3 (Gal-3) in the pathophysiology of thrombosis is unknown. Our hypothesis is that Gal-3BP and Gal-3 are critical to thrombus formation. Methods: Mice : Using our inferior vena cava ligation mouse model of VT, microparticle samples from thrombosed and non-thrombosed wild type mice were analyzed for Gal-3BP. We then either neutralized Gal-3BP, using an antibody (Anti-M-Gal-3BP), or utilized Gal-3 knock-out (Gal-3 KO) mice, compared to mice given saline as controls (NaCl). We evaluated thrombus weight (TW), inflammatory cell counts, and the cell source of the molecules Gal-3BP and Gal-3 using western blot. Patients: Microparticles and plasma samples from patients who were positive or negative for VT were tested for Gal-3BP using ELISA. Results: Mice : Two days after thrombosis, blocking Gal-3BP significantly reduced both vein wall inflammatory cells ( p =0.0369) and TW ( p =0.0024), compared to NaCl. In Gal-3 KO mice, TW was significantly smaller versus controls ( p =0.0061). We found Gal-3BP on microparticles and platelets while Gal-3 was on monocytes. Patients : Gal-3BP was significantly elevated in microparticles ( p =0.0495) and in plasma ( p =0.0400), versus patients negative for VT. Conclusions: We demonstrated a clear link between Gal-3BP and VT, in both mice and patients. This is the first time that Gal-3BP has been shown to be involved with thrombus formation, making it a promising biomarker and potential target for therapeutic interventions of VT.