No abstract

Background: Quercetin-3-rutinoside is an inhibitor of protein disulfide isomerase, a potential target for antithrombotic therapy. Results: Quercetin-3-rutinoside induces a compact conformation in PDI and binds to PDI with an IC 50 of about 10 M. Conclusion: Quercetin-3-rutinoside interacts with the bЈx domain of protein disulfide isomerase with a 1:1 stoichiometry. Significance: The bЈx domain reverses the antithrombotic properties of quercetin-3-rutinoside in a thrombosis model in a live mouse.

• ERp5, like its family members PDI and ERp57, accumulates at sites of vessel wall injury.• Both platelets and endothelium secrete ERp5 on activation and contribute ERp5 necessary for thrombus formation in vivo.Protein disulfide isomerase (PDI) and endoplasmic reticulum protein 57 (ERp57) are emerging as important regulators of thrombus formation. Another thiol isomerase, endoplasmic reticulum protein 5 (ERp5), is involved in platelet activation. We show here the involvement of ERp5 in thrombus formation using the mouse laser-injury model of thrombosis and a specific antibody raised against recombinant ERp5. Anti-ERp5 antibody inhibited ERp5-dependent platelet and endothelial cell disulfide reductase activity in vitro. ERp5 release at the thrombus site was detected after infusion of Alexa Fluor 488-labeled anti-ERp5 antibody at 0.05 mg/g body weight, a dose that does not inhibit thrombus formation. Anti-ERp5 at 3 mg/g body weight inhibited laser-induced thrombus formation in vivo by causing a 70% decrease in the deposition of platelets and a 62% decrease in fibrin accumulation compared to infusion of control antibody (P < .01). ERp5 binds to b3 integrin with an equilibrium dissociation constant (K D ) of 21 mM, measured by surface plasmon resonance. The cysteine residues in the ERp5 active sites are not required for binding to b3 integrin. These results provide evidence for a novel role of ERp5 in thrombus formation, a function that may be mediated through its association with aIIbb3.