Studies have shown significantly increased thromboembolic events at high-altitude. We recently reported that transferrin could potentiate blood coagulation, but the underlying mechanism for high-altitude-related thromboembolism is still poorly understood. Here, we examined the activity and concentration of plasma coagulation factors and transferrin in plasma collected from long-term human residents and short-stay mice exposed to varying altitudes. We found that the activities of thrombin and FXIIa along with the concentrations of transferrin were significantly increased in the plasma of humans and mice at high altitudes. Furthermore, both hypoxia (6% O2) and low temperature (0 °C), two critical high-altitude factors, enhanced hypoxia-inducible factor (HIF)-1a levels to promote the expression of the transferrin gene, whose enhancer region contains HIF-1a binding site, and consequently to induce hypercoagulability by potentiating thrombin and FXIIa. Importantly, thromboembolic disorders and pathological insults in mouse models induced by both hypoxia and low temperature were ameliorated by transferrin interferences, including transferrin antibody treatment, transferrin down-regulation, and the administration of our designed peptides that inhibit the potentiation of transferrin on thrombin and FXIIa. Thus, low temperature and hypoxia up-regulated transferrin expression promoted hypercoagulability. Our data suggest that targeting the transferrin-coagulation pathway is a novel and may be a powerful strategy against thromboembolic events caused by harmful environmental factors under high-altitude conditions.

Several adaptor molecules bind to cytoplasmic tails of β-integrins and facilitate bidirectional signaling, which is critical in thrombosis and hemostasis. Interfering with integrin-adaptor interactions spatially or temporally to inhibit thrombosis without affecting hemostasis is an attractive strategy for the development of safe anti-thrombotics. Here we show for the first time that 14-3-3ζ-c-Src-integrin-β3 complex is formed during platelet activation. 14-3-3ζ-c-Src interaction is mediated by -pirlglalnfsvfyye- fragment (PE16) on 14-3-3ζ and SH2-domain on c-Src, while 14-3-3ζ-integrin β3 interaction is mediated by -eskvfylkmkgdyyrYL- fragment (EL17) on 14-3-3ζ and -keatstf- fragment (KF7) on β3 integrin cytoplasmic tail. EL17-motif inhibitor or KF7 peptide interferes with the formation of 14-3-3ζ-c-Src-integrin-β3 complex and selectively inhibits β3 outside-in signaling without affecting the integrin-fibrinogen interaction, which suppresses thrombosis without causing significant bleeding. This study characterizes a previously unidentified 14-3-3ζ-c-Src-integrin-β3 complex in platelets and provides a novel strategy for the development of safe and effective anti-thrombotic therapies.

: Platelets are small blood cells known primarily for their ability to adhere and aggregate at injured vessels to arrest bleeding. However, when triggered under pathological conditions, the same adaptive mechanism of platelet adhesion and aggregation may cause thrombosis, a primary cause of heart attack and stroke. Over recent decades, research has made considerable progress in uncovering the intricate and dynamic interactions that regulate these processes. Integrins are heterodimeric cell surface receptors expressed on all metazoan cells that facilitate cell adhesion, movement, and signaling, to drive biological and pathological processes such as thrombosis and hemostasis. Recently, our group discovered that the plexinsemaphorin-integrin (PSI) domains of the integrin β subunits exert endogenous thiol isomerase activity derived from their two highly conserved CXXC active site motifs. Given the importance of redox reactions in integrin activation and its location in the knee region, this PSI domain activity may be critically involved in facilitating the interconversions between integrin conformations. Our monoclonal antibodies against the β3 PSI domain inhibited its thiol isomerase activity and proportionally attenuated fibrinogen binding and platelet aggregation. Notably, these antibodies inhibited thrombosis without significantly impairing hemostasis or causing platelet clearance. In this review, we will update mechanisms of thrombosis and hemostasis including platelet versatilities and immune-mediated thrombocytopenia, discuss critical contributions of the newly discovered PSI domain thiol isomerase activity, and its potential as a novel target for anti-thrombotic therapies and beyond.

Kawasaki disease (KD) is an acute systemic vasculitis primarily affecting infants and children. Activated platelets predispose patients to coronary artery structural lesions that may lead to thrombotic cardiovascular events. To discover potential proteins underlying platelet activation in KD, we conducted a protein chip assay of 34 cytokines and discovered thymic stromal lymphopoietin (TSLP) was aberrantly expressed, which remained elevated after intravenous immunoglobulin G (IVIG) treatment and during convalescence in KD patients in comparison to healthy controls. Enzymelinked immunosorbent assay (ELISA) corroborated the upregulation of TSLP in KD patients, which was exacerbated in convalescent patients complicated with thrombosis. TSLP receptors on platelets were also significantly upregulated in KD patients complicated with thrombosis. Platelet activation, apoptosis, and mitochondrial autophagy (mitophagy) were increased in convalescence KD patients complicated with thrombosis. In vitro, TSLP induced platelet activation and platelet mitophagy in healthy blood donors, as observed in KD patients. TSLP, similar to mitophagy agonist carbonyl cyanide 3-chlorophenyl hydrazone (CCCP), promoted thrombosis, which was attenuated by the mitophagy inhibitor Mdivi-1. Co-immunoprecipitation