Endoplasmic reticulum protein 29 negatively regulates platelet functions and thrombosis in mice.

Background: Several members of protein disulfide isomerase (PDI) family with the CXYC active motif such as PDI, ERp57, ERp72, ERp46, ERp5 and TMX1 have important roles in platelet functions and thrombosis. These members contribute to the network of redox regulation of platelet activities. However, whether other PDI family members without the CXYC motif such as ERp29, have a role in these processes remains unknown.

Aims: To determine the role of ERp29 in platelet functions and thrombosis.

Methods: The phenotypes of platelet-specific ERp29-deficient (Pf4-Cre/ERp29^fl/fl) mice were evaluated using tail bleeding assay and laser-induced and FeCl₃-induced arterial injury models, as well as venous thrombosis model. In vitro, the functions of ERp29-deficient platelets were assessed in respect to aggregation, adhesion, spreading, clot retraction, granule secretion and integrin αIIbβ3 activation measured by flow cytometry. Redox state of integrin αIIbβ3 thiols was detected using 3-(N-maleimido-propionyl) biotin (MPB) labeling.

Results: Compared with WT mice, Pf4-Cre/ERp29^fl/fl mice exhibited shortened tail-bleeding times, increased platelet accumulation in the two arterial thrombosis models, and enhanced thrombogenesis in the venous thrombosis model. ERp29-deficient platelets had enhanced response in aggregation, ATP release, spreading, clot retraction, αIIbβ3 activation, fibrinogen binding and P-selectin expression. As detected by MPB labeling, the free thiol content of integrin αIIbβ3 in ERp29-deficient platelets were increased compared with WT platelets, suggesting that the role of ERp29 is associated with oxidation of the functional disulfides of integrin αIIb and/or β3 subunits.

Conclusion(s): ERp29 is the first disulfide isomerase without the CXYC motif that negatively regulates platelet function. This study provides new insight into the redox network controlling thrombosis.