Apolipoprotein A-IV binds αIIbβ3 integrin and inhibits thrombosis.

Platelet αIIbβ3 integrin and its ligands are essential for thrombosis and hemostasis, and play key roles in myocardial infarction and stroke. Here we show that apolipoprotein A-IV (apoA-IV) can be isolated from human blood plasma using platelet β3 integrin-coated beads. Binding of apoA-IV to platelets requires activation of αIIbβ3 integrin, and the direct apoA-IV-αIIbβ3 interaction can be detected using a single-molecule Biomembrane Force Probe.

Improved resolution in extracellular vesicle populations using 405 instead of 488 nm side scatter.

Improvements in identification and assessment of extracellular vesicles (EVs) have fuelled a recent surge in EV publications investigating their roles as biomarkers and mediators of disease. Meaningful scientific comparisons are, however, hampered by difficulties in accurate, reproducible enumeration and characterization of EVs in biological fluids. High-sensitivity flow cytometry (FCM) is presently the most commonly applied strategy to assess EVs, yet its utility is limited by variant ability to resolve smaller EVs.

Novel nuclear hENT2 isoforms regulate cell cycle progression via controlling nucleoside transport and nuclear reservoir.

Nucleosides participate in many cellular processes and are the fundamental building blocks of nucleic acids. Nucleoside transporters translocate nucleosides across plasma membranes although the mechanism by which nucleos(t)ides are translocated into the nucleus during DNA replication is unknown. Here, we identify two novel functional splice variants of equilibrative nucleoside transporter 2 (ENT2), which are present at the nuclear envelope.

Microparticles as biomarkers of lung disease: enumeration in biological fluids using lipid bilayer microspheres.

Extracellular vesicles, specifically microparticles (MPs), are rapidly gaining attention for their capacity to act as biomarkers for diagnosis, prognosis, or responsiveness to therapy in lung disease, in keeping with the concept of precision medicine. However, MP analysis by high-sensitivity flow cytometry (FCM) is complicated by a lack of accurate means for MP enumeration. To address this gap, we report here an enhanced FCM MP gating and enumeration technique based on the use of novel engineered lipid bilayer microspheres (LBMs).

Plasma fibronectin supports hemostasis and regulates thrombosis.

Plasma fibronectin (pFn) has long been suspected to be involved in hemostasis; however, direct evidence has been lacking. Here, we demonstrated that pFn is vital to control bleeding in fibrinogen-deficient mice and in WT mice given anticoagulants. At the site of vessel injury, pFn was rapidly deposited and initiated hemostasis, even before platelet accumulation, which is considered the first wave of hemostasis.

Dok-2 adaptor protein regulates the shear-dependent adhesive function of platelet integrin αIIbβ3 in mice.

The Dok proteins are a family of adaptor molecules that have a well defined role in regulating cellular migration, immune responses, and tumor progression. Previous studies have demonstrated that Doks-1 to 3 are expressed in platelets and that Dok-2 is tyrosine-phosphorylated downstream of integrin αIIbβ3, raising the possibility that it participates in integrin αIIbβ3 outside-in signaling. We demonstrate that Dok-2 in platelets is primarily phosphorylated by Lyn kinase.

Cadherin 6 has a functional role in platelet aggregation and thrombus formation.

Objective: Thrombosis occurs at sites of vascular injury when platelets adhere to subendothelial matrix proteins and to each other. Platelets express many surface receptor proteins, the function of several of these remains poorly characterized. Cadherin 6 is expressed on the platelet surface and contains an arginine-glycine-aspartic acid motif, suggesting that it might have a supportive role in thrombus formation.

Relative efficacy of steroid therapy in immune thrombocytopenia mediated by anti-platelet GPIIbIIIa versus GPIbα antibodies.

Immune thrombocytopenia (ITP) is characterized by platelet clearance mediated primarily by autoantibodies against the platelet GPIIbIIIa and/or GPIbα. Steroid therapy is a first-line treatment for ITP. However, some patients are refractory to this therapy and currently no method can predict which patients will respond.

Animal model of fetal and neonatal immune thrombocytopenia: role of neonatal Fc receptor in the pathogenesis and therapy.

Fetal and neonatal immune thrombocytopenia (FNIT) is a severe bleeding disorder in which maternal antibodies cross the placenta and destroy fetal/neonatal platelets. It has been demonstrated that the neonatal Fc receptor (FcRn) regulates immunoglobulin G (IgG) homeostasis and plays an important role in transplacental IgG transport. However, the role of FcRn in the pathogenesis and therapy of FNIT has not been studied.

Fibrinogen is required for maintenance of platelet intracellular and cell-surface P-selectin expression.

Platelet P-selectin plays important roles in inflammation and contributes to thrombosis and hemostasis. Although it has been reported that von Willebrand factor (VWF) affects P-selectin expression on endothelial cells, little information is available regarding regulation of platelet P-selectin expression. Here, we first observed that P-selectin expression was significantly decreased on platelets of fibrinogen and VWF double-deficient mice.

Plasma fibronectin depletion enhances platelet aggregation and thrombus formation in mice lacking fibrinogen and von Willebrand factor.

We previously showed that platelet aggregation and thrombus formation occurred in mice lacking both fibrinogen (Fg) and von Willebrand factor (VWF) and that plasma fibronectin (pFn) promoted thrombus growth and stability in injured arterioles in wild-type mice. To examine whether pFn is required for Fg/VWF-independent thrombosis, we generated Fg/VWF/conditional pFn triple-deficient (TKO; Cre(+), Fn(flox/flox), Fg/VWF(-/-)) mice and littermate control (Cre(-), Fn(flox/flox), Fg/VWF(-/-)) mice. Surprisingly, TKO platelet aggregation was not abolished, but instead was enhanced in both heparinized platelet-rich plasma and gel-filtered platelets.

A novel murine model of fetal and neonatal alloimmune thrombocytopenia: response to intravenous IgG therapy.

Fetal and neonatal alloimmune thrombo cytopenia (FNAITP) is a life-threatening bleeding disorder caused by maternal antibodies directed against fetal platelet antigens. The immunoreactive epitopes in FNAITP are primarily located in the extracellular regions of the platelet glycoprotein IIIa (beta3 integrin). Here we have established a novel animal model of FNAITP using beta3 integrin-deficient (beta3-/-) mice.

The catenin p120ctn inhibits Kaiso-mediated transcriptional repression of the beta-catenin/TCF target gene matrilysin.

The POZ-zinc finger transcription factor Kaiso was first identified as a specific binding partner for the Armadillo catenin and cell adhesion cofactor, p120ctn. Kaiso is a unique POZ protein with bi-modal DNA-binding properties; it associates with a sequence-specific DNA consensus Kaiso binding site (KBS) or methylated CpG dinucleotides, and regulates transcription of artificial promoters containing either site. Interestingly, the promoter of the Wnt/beta-catenin/TCF target gene matrilysin possesses two conserved copies of the KBS, which suggested that Kaiso might regulate matrilysin expression.

Non-canonical Wnt signals are modulated by the Kaiso transcriptional repressor and p120-catenin.

Gastrulation movements are critical for establishing the three principal germ layers and the basic architecture of vertebrate embryos. Although the individual molecules and pathways involved are not clearly understood, non-canonical Wnt signals are known to participate in developmental processes, including planar cell polarity and directed cell rearrangements. Here we demonstrate that the dual-specificity transcriptional repressor Kaiso, first identified in association with p120-catenin, is required for Xenopus gastrulation movements.

NLS-dependent nuclear localization of p120ctn is necessary to relieve Kaiso-mediated transcriptional repression.

The Armadillo catenin p120(ctn) regulates cadherin adhesive strength at the plasma membrane and interacts with the novel BTB/POZ transcriptional repressor Kaiso in the nucleus. The dual localization of p120(ctn) at cell-cell junctions and in the nucleus suggests that its nucleocytoplasmic trafficking is tightly regulated. Here we report on the identification of a specific and highly basic nuclear localization signal (NLS) in p120(ctn).

The p120(ctn)-binding partner Kaiso is a bi-modal DNA-binding protein that recognizes both a sequence-specific consensus and methylated CpG dinucleotides.

The p120(ctn)-binding partner Kaiso is a new member of the POZ-zinc finger family of transcription factors implicated in development and cancer. To understand the role of Kaiso in gene regulation and p120(ctn)-mediated signaling and adhesion, we sought to identify Kaiso-specific DNA binding sequences and potential target genes. Here we demonstrate that Kaiso is a dual specificity DNA-binding protein that recognizes the specific consensus sequence TCCTGCNA as well as methyl-CpG dinucleotides.