General Anesthesia and the Premature Baby: Identifying Risks for Poor Neurodevelopmental Outcomes.

Preterm birth affects 1 in every 10 infants born in the United States. Importantly, more preterm infants are surviving to discharge from hospital, including those born at the cusp of viability (eg, 22 to 24 wk gestation). Such improvements, however, come at a cost as those delivered at less than 28 weeks gestation have the highest rates of morbidity and mortality.

Detection of circulating extracellular mRNAs by modified small-RNA-sequencing analysis.

Extracellular mRNAs (ex-mRNAs) potentially supersede extracellular miRNAs (ex-miRNAs) and other RNA classes as biomarkers. We performed conventional small-RNA-sequencing (sRNA-seq) and sRNA-seq with T4 polynucleotide kinase (PNK) end-treatment of total exRNA isolated from serum and platelet-poor EDTA, ACD, and heparin plasma to study the effect on ex-mRNA capture. Compared to conventional sRNA-seq PNK-treatment increased the detection of informative ex-mRNAs reads up to 50-fold.

Outcomes Research in Vulnerable Pediatric Populations.

The Pediatric Anesthesia and Neurodevelopment Assessment (PANDA) study team held its biennial symposium in April 2018 to discuss issues on anesthetic neurotoxicity in the developing brain. One of the sessions invited speakers with different areas of expertise to discuss "Outcomes Research in Vulnerable Pediatric Populations." The vulnerable populations included neonates, children with congenital heart disease, children from low socioeconomic status, and children with incarcerated parents.

Human plasma and serum extracellular small RNA reference profiles and their clinical utility.

Circulating extracellular RNAs (exRNAs) have the potential to serve as biomarkers for a wide range of medical conditions. However, limitations in existing exRNA isolation methods and a lack of knowledge on parameters affecting exRNA variability in human samples may hinder their successful discovery and clinical implementation. Using combinations of denaturants, reducing agents, proteolysis, and revised organic extraction, we developed an automated, high-throughput approach for recovery of exRNAs and exDNA from the same biofluid sample.

P2Y Receptor Function and Response to Cangrelor in Neonates With Cyanotic Congenital Heart Disease.

Shunt thrombosis remains a major cause of morbidity and mortality, especially during the initial palliation for single-ventricle physiology. The authors present evidence that the P2Y inhibitor cangrelor may fill a therapeutic void in thromboprophylaxis. They base this theory on results showing that platelets from neonatal patients with cyanotic congenital heart disease have a robust response to adenosine diphosphate and are amenable to P2Y inhibition with cangrelor.

PI3Kγ/δ and NOTCH1 Cross-Regulate Pathways That Define the T-cell Acute Lymphoblastic Leukemia Disease Signature.

PI3K/AKT and NOTCH1 signaling pathways are frequently dysregulated in T-cell acute lymphoblastic leukemias (T-ALL). Although we have shown that the combined activities of the class I PI3K isoforms p110γ and p110δ play a major role in the development and progression of PTEN-null T-ALL, it has yet to be determined whether their contribution to leukemogenic programing is unique from that associated with NOTCH1 activation. Using an -driven mouse model of T-ALL in which both the PI3K/AKT and NOTCH1 pathways are aberrantly upregulated, we now demonstrate that the combined activities of PI3Kγ/δ have both overlapping and distinct roles from NOTCH1 in generating T-ALL disease signature and in promoting tumor cell growth.

Acalabrutinib (ACP-196) in Relapsed Chronic Lymphocytic Leukemia.

Background: Irreversible inhibition of Bruton's tyrosine kinase (BTK) by ibrutinib represents an important therapeutic advance for the treatment of chronic lymphocytic leukemia (CLL). However, ibrutinib also irreversibly inhibits alternative kinase targets, which potentially compromises its therapeutic index. Acalabrutinib (ACP-196) is a more selective, irreversible BTK inhibitor that is specifically designed to improve on the safety and efficacy of first-generation BTK inhibitors.

Exploiting the kinetic interplay between GPIbα-VWF binding interfaces to regulate hemostasis and thrombosis.

Platelet-von Willebrand factor (VWF) interactions must be tightly regulated in order to promote effective hemostasis and prevent occlusive thrombus formation. However, it is unclear what role the inherent properties of the bond formed between the platelet receptor glycoprotein Ibα and the A1 domain of VWF play in these processes. Using VWF-A1 knock-in mice with mutations that enhance (I1309V) or disrupt (R1326H) platelet receptor glycoprotein Ibα binding, we now demonstrate that the kinetic interplay between two distinct contact surfaces influences the site and extent to which platelets bind VWF.

RUC-4: a novel αIIbβ3 antagonist for prehospital therapy of myocardial infarction.

Objective: Treatment of myocardial infarction within the first 1 to 2 hours with a thrombolytic agent, percutaneous coronary intervention, or an αIIbβ3 antagonist decreases mortality and the later development of heart failure. We previously reported on a novel small molecule αIIbβ3 antagonist, RUC-2, that has a unique mechanism of action. We have now developed a more potent and more soluble congener of RUC-2, RUC-4, designed to be easily administered intramuscularly by autoinjector to facilitate its use in the prehospital setting.

A selective microRNA-based strategy inhibits restenosis while preserving endothelial function.

Drugs currently approved to coat stents used in percutaneous coronary interventions do not discriminate between proliferating vascular smooth muscle cells (VSMCs) and endothelial cells (ECs). This lack of discrimination delays reendothelialization and vascular healing, increasing the risk of late thrombosis following angioplasty. We developed a microRNA-based (miRNA-based) approach to inhibit proliferative VSMCs, thus preventing restenosis, while selectively promoting reendothelialization and preserving EC function.

Analysis of early thrombus dynamics in a humanized mouse laser injury model.

Platelet aggregation and thrombus formation at the site of injury is a dynamic process that involves the continuous addition of new platelets as well as thrombus rupture. In the early stages of hemostasis (within minutes after vessel injury) this process can be visualized by transfusing fluorescently labeled human platelets and observing their deposition and detachment. These two counterbalancing events help the developing thrombus reach a steady-state morphology, where it is large enough to cover the injured vessel surface but not too large to form a severe thrombotic occlusion.

Simulation of platelet, thrombus and erythrocyte hydrodynamic interactions in a 3D arteriole with in vivo comparison.

Cylindrical blood vessels, ellipsoid platelets and biconcave-shaped deformable erythrocytes (RBCs) are important participants in hemostasis and thrombosis. However, due to the challenge of combining these components in simulation tools, few simulation studies have included all of them in realistic three-dimensional models. In the present study, we apply a recently developed simulation model to incorporate these components and analyze the flow in a thrombotic tubular arteriole, particularly the detailed hydrodynamic interactions between the thrombus shape, RBCs and platelets.

Targeting nonclassical oncogenes for therapy in T-ALL.

Constitutive phosphoinositide 3-kinase (PI3K)/Akt activation is common in T cell acute lymphoblastic leukemia (T-ALL). Although four distinct class I PI3K isoforms (α, β, γ, δ) could participate in T-ALL pathogenesis, none has been implicated in this process. We report that in the absence of PTEN phosphatase tumor suppressor function, PI3Kγ or PI3Kδ alone can support leukemogenesis, whereas inactivation of both isoforms suppressed tumor formation.

Humanizing thrombi in mice.

Antiplatelet therapies form the cornerstone of atherothrombosis prevention, reducing the morbidity and mortality associated with cardiovascular disease. Despite these benefits, there is still an unmet need for more effective and safer pharmacological agents. To expedite this process, biological platforms that better reflect the intravascular environment in humans will be required in order to shorten drug development time, enable better determination of dosing regimes, and aid in the design of clinical studies.

Histamine deficiency promotes inflammation-associated carcinogenesis through reduced myeloid maturation and accumulation of CD11b+Ly6G+ immature myeloid cells.

Histidine decarboxylase (HDC), the unique enzyme responsible for histamine generation, is highly expressed in myeloid cells, but its function in these cells is poorly understood. Here we show that Hdc-knockout mice show a high rate of colon and skin carcinogenesis. Using Hdc-EGFP bacterial artificial chromosome (BAC) transgenic mice in which EGFP expression is controlled by the Hdc promoter, we show that Hdc is expressed primarily in CD11b(+)Ly6G(+) immature myeloid cells (IMCs) that are recruited early on in chemical carcinogenesis.

A novel molecular interaction for the adhesion of follicular CD4 T cells to follicular DC.

Nectins and Nectin-like molecules (Necl) play a critical role in cell polarity within epithelia and in the nervous and reproductive systems. Recently, immune receptors specific for Nectins/Necl have been described. Since the expression and distribution of Nectins/Necl is often subverted during tumorigenesis, it has been suggested that the immune system may use these receptors to recognize and eliminate tumors.

Role of PI3Kdelta and PI3Kgamma in inflammatory arthritis and tissue localization of neutrophils.

The p110delta isoform of class I phosphoinositide 3-kinase (PI3K) plays a major role in B cell receptor signaling, while its p110gamma counterpart is thought to predominate in leukocyte chemotaxis. Consequently, emphasis has been placed on developing PI3Kgamma selective inhibitors to treat disease states that result from inappropriate tissue accumulation of leukocytes. We now demonstrate that PI3Kdelta blockade is effective in treating an autoimmune disorder in which neutrophil infiltration is required for tissue injury.

Modifying murine von Willebrand factor A1 domain for in vivo assessment of human platelet therapies.

The A1 domain of von Willebrand factor (VWF-A1) plays a crucial role in hemostasis and thrombosis by initiating platelet adhesion at sites of arterial injury through interactions with the platelet receptor glycoprotein Ib alpha (GPIbalpha). Here we report that murine VWF-A1 supports limited binding of human platelets. However, atomic models of GPIbalpha-VWF-A1 complexes identified an electrostatic 'hot-spot' that, when mutated in murine VWF-A1, switches its binding specificity from mouse to human GPIbalpha.

p110gamma and p110delta phosphoinositide 3-kinase signaling pathways synergize to control development and functions of murine NK cells.

Phosphoinositide 3-kinases (PI-3Ks) are key enzymes for cell development, activation, and survival. Here we showed that PI-3K class IB and class IA catalytic subunits, p110gamma and p110delta, played a crucial role in the development and functions of murine NK cells. p110gamma deficiency and impairment of G protein-coupled receptor (GPRC) signaling prevented full NK cell maturation.

Essential role of PI3Kdelta and PI3Kgamma in thymocyte survival.

Class 1 phosphoinositide 3-kinases (PI3Ks), consisting of PI3Kalpha, beta, gamma, and delta, are a family of intracellular signaling molecules that play important roles in cell-mediated immune responses. In thymocytes, however, their role is less clear, although PI3Kgamma is postulated to partially contribute to pre-TCR-dependent differentiation. We now report that PI3Kdelta, in conjunction with PI3Kgamma, is required for thymocyte survival and ultimately for T-cell production.

Adhesive mechanisms governing interferon-producing cell recruitment into lymph nodes.

Natural interferon-producing cells (IPCs) are found in peripheral lymph nodes (PLNs), where they support NK cell, T cell, and B cell responses to pathogens. However, their route of entry and the adhesive mechanisms used to gain access to PLNs remain poorly defined. We report that IPCs can enter PLNs via a hematogenous route, which involves a multistep adhesive process, and that transmigration is enhanced by inflammation.

The role of endothelial PI3Kgamma activity in neutrophil trafficking.

Phosphoinositide 3-kinase gamma (PI3Kgamma) in neutrophils plays a critical role in the directed migration of these cells into inflamed tissues. In this study, we demonstrate the importance of the endothelial component of PI3Kgamma activity relative to its leukocyte counterpart in supporting neutrophil interactions with the inflamed vessel wall. Despite the reconstitution of class-Ib PI3K function in neutrophils of p110gamma-/- mice, we observed a 45% reduction in accumulation of these cells in an acute lung injury model.

The snake venom protein botrocetin acts as a biological brace to promote dysfunctional platelet aggregation.

Botrocetin is a snake venom protein that enhances the affinity of the A1 domain of plasma von Willebrand factor (vWF) for the platelet receptor glycoprotein Ibalpha (GPIbalpha), an event that contributes to bleeding and host death. Here we describe a kinetic and crystallographic analysis of this interaction that reveals a novel mechanism of affinity enhancement. Using high-temporal-resolution microscopy, we show that botrocetin decreases the GPIbalpha off-rate two-fold in both human and mouse complexes without affecting the on-rate.

Mechanics of transient platelet adhesion to von Willebrand factor under flow.

A primary and critical step in platelet attachment to injured vascular endothelium is the formation of reversible tether bonds between the platelet glycoprotein receptor Ibalpha and the A1 domain of surface-bound von Willebrand factor (vWF). Due to the platelet's unique ellipsoidal shape, the force mechanics involved in its tether bond formation differs significantly from that of leukocytes and other spherical cells. We have investigated the mechanics of platelet tethering to surface-immobilized vWF-A1 under hydrodynamic shear flow.