PLATELET FUNCTION ASSESSMENT IN A MICROFABRICATED DEVICE

Introduction Although platelets are small and simple in shape, they are complicated in their physiologhy. Their alpha granules and dense bodies secrete a large number of agents that are involved in haemostasis, and the glycoproteins on thei r surfaces form the linkages with proteins like fibrinogen, fibronectin, collagen and von Willebrand factor that are necessary for adhesion and aggregation A simple way to test platelet function is to immobilize substrates on a flow channel, induce flow over the substrate, and measure the percentage of surface covered by platelet adhesion. The use of several substrates can allow the assessment of a variety of platelet functions. A practical platelet analyzer should 1) be capable of distinguishing multiple aspects of platelet behavior, 2) require a small amount of blood, 3) examine platelet function under shear flow conditions, 4) be relatively simple to use, and 5) be relatively easy to manufacture. Modern micromanufacturing methods are readily available for the construction of the microchannels that can be filled with volumes of blood on the order of ?L. However, a technique is still needed to coat these channels with the appropriate protein substrates. Recently, a process called layer-by-layer assembly (Lvov et al., 2000) has been developed that allows well-controlled protein coatings to be adsorbed onto charged surfaces. The process takes advantage of the charge already on the surface to lay down, in alternating layers, positively and negatively charged ions. The combination of layer-bylayer assembly and micromanufacturing can thus form a basis for the device being sought. Key questions to be answered are whether it is possible to assemble the needed substrates and whether the platelet adhesion patterns behave as expected. For example, the amount of adhesion should be dependent on the substrate and the fluid shear stress. Because fibrinogen is a key protein in both platelet adhesion and platelet aggregation, this was selected as the first protein to be examined. Methods The microchannels were first created as ridges on a silicon wafer. The photopolymer SU-8 was spun onto the wafer and exposed through a mask under ultraviolet light to allow the polymer to cure. Afterwards, the wafer was deve loped in SU-8 developer, leaving SU-8 ridges only where the surface was exposed to light. The cured wafer was then used as a mold for polydimethylsiloxane (PDMS), an optically clear polymer that cures at room temperature in three days. The raised portions on the silicon then become microchannels in the PDMS. The PDMS microchannels were coated by the layer-by-layer selfassembly technique to provide controlled nanometer-thick layers of fibrinogen. A plexiglass plate was created to cover the channels, and inlet and outlet ports were incorporated into the cover to allow the injection of blood. A syringe injector (Cole Parmer) was used to inject platelet rich plasma at a controlled flow rate. Anticoagulated platelet rich plasma labeled with both acridine orange and a fluorescin isothiocynate-tagged anti-GpIIb/IIIa-antibody was passed through the microchannels. Several experimental runs for different shear rates were carried out. To estimate shear, Couette flow was assumed in the microchannels. Control experiments were performed on bare PDMS surfaces. Images were recorded with a fluorescent microscope. For each image, background subtraction was applied, followed by a thresholding technique to distinguish dark areas (platelets) from light areas (substrate with no adhesion). From these steps, the extent o

Cohort profile: seek, test, treat and retain United States criminal justice cohort

Abstract Background The STTR treatment cascade provides a framework for research aimed at improving the delivery of services, care and outcomes of PLWH. The development of effective approaches to increase HIV diagnoses and engage PLWH in subsequent steps of the treatment cascade could lead to earlier and sustained ART treatment resulting in viral suppression. There is an unmet need for research applying the treatment cascade to improve outcomes for those with criminal justice involvement. Methods The Seek, Test, Treat, and Retain (STTR) criminal justice (CJ) cohort combines data from 11 studies across the HIV treatment cascade that focused on persons involved in the criminal justice system, often but not exclusively for reasons related to substance use. The studies were conducted in a variety of CJ settings and collected information across 11 pre-selected domains: demographic characteristics, CJ involvement, HIV risk behaviors, HIV and/or Hepatitis C infections, laboratory measures of CD4 T-cell count (CD4) and HIV RNA viral load (VL), mental illness, health related quality of life (QoL), socioeconomic status, health care access, substance use, and social support. Results The STTR CJ cohort includes data on 11,070 individuals with and without HIV infection who range in age from 18 to 77 years, with a median age at baseline of 37 years. The cohort reflects racial, ethnic and gender distributions in the U.S. CJ system, and 64% of participants are African-American, 12% are Hispanic and 83% are men. Cohort members reported a wide range of HIV risk behaviors including history of injection drug use and, among those who reported on pre-incarceration sexual behaviors, the prevalence of unprotected sexual intercourse ranged across studies from 4% to 79%. Across all studies, 53% percent of the STTR CJ cohort reported recent polysubstance use. Conclusions The STTR CJ cohort is comprised of participants from a wide range of CJ settings including jail, prison, and community supervision who report considerable diversity in their characteristics and behavioral practices. We have developed harmonized measures, where feasible, to improve the integration of these studies together to answer questions that cannot otherwise be addressed

2015 ESC Guidelines for the management of patients with ventricular arrhythmias and the prevention of sudden cardiac death the Task Force for the Management of Patients with Ventricular Arrhythmias and the Prevention of Sudden Cardiac Death of the European Society of Cardiology (ESC) Endorsed by: Association for European Paediatric and Congenital Cardiology (AEPC)

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The Staphylococcus aureus superantigen SElX is a bifunctional toxin that inhibits neutrophil function:SElX Inhibits Neutrophil Function

Bacterial superantigens (SAgs) cause Vβ-dependent T-cell proliferation leading to immune dysregulation associated with the pathogenesis of life-threatening infections such as toxic shock syndrome, and necrotizing pneumonia. Previously, we demonstrated that staphylococcal enterotoxin-like toxin X (SElX) from Staphylococcus aureus is a classical superantigen that exhibits T-cell activation in a Vβ-specific manner, and contributes to the pathogenesis of necrotizing pneumonia. Here, we discovered that SElX can also bind to neutrophils from human and other mammalian species and disrupt IgG-mediated phagocytosis. Site-directed mutagenesis of the conserved sialic acid-binding motif of SElX abolished neutrophil binding and phagocytic killing, and revealed multiple glycosylated neutrophil receptors for SElX binding. Furthermore, the neutrophil binding-deficient mutant of SElX retained its capacity for T-cell activation demonstrating that SElX exhibits mechanistically independent activities on distinct cell populations associated with acquired and innate immunity, respectively. Finally, we demonstrated that the neutrophil-binding activity rather than superantigenicity is responsible for the SElX-dependent virulence observed in a necrotizing pneumonia rabbit model of infection. Taken together, we report the first example of a SAg, that can manipulate both the innate and adaptive arms of the human immune system during S. aureus pathogenesis

Attitudes Concerning Wind Energy in Central Illinois

Energy consumption in the United Sates has been increasing and the cost of fossil fuels has been unstable in recent years. Expanding investment in renewable energy is one way to reduce the nation’s dependence on fossil fuels. However, the Not-In-My-Back- Yard (NIMBY) opposition and various concerns raised at public hearings may inhibit the expansion of wind energy in Illinois. This study aims to characterize the public beliefs and opinions toward wind energy in central Illinois

Severity of Demyelinating and Axonal Neuropathy Mouse Models Is Modified by Genes Affecting Structure and Function of Peripheral Nodes

Charcot-Marie-Tooth (CMT) disease is a clinically and genetically heterogeneous group of inherited polyneuropathies. Mutations in 80 genetic loci can cause forms of CMT, resulting in demyelination and axonal dysfunction. The clinical presentation, including sensory deficits, distal muscle weakness, and atrophy, can vary greatly in severity and progression. Here, we used mouse models of CMT to demonstrate genetic interactions that result in a more severe neuropathy phenotype. The cell adhesion molecule Nrcam and the Na+ channel Scn8a (NaV1.6) are important components of nodes. Homozygous Nrcam and heterozygous Scn8a mutations synergized with both an Sh3tc2 mutation, modeling recessive demyelinating Charcot-Marie-Tooth type 4C, and mutations in Gars, modeling dominant axonal Charcot-Marie-Tooth type 2D. We conclude that genetic variants perturbing the structure and function of nodes interact with mutations affecting the cable properties of axons by thinning myelin or reducing axon diameter. Therefore, genes integral to peripheral nodes are candidate modifiers of peripheral neuropathy

Preclinical Models of Pancreatic Ductal Adenocarcinoma and Their Utility in Immunotherapy Studies

The advent of immunotherapy has transformed the treatment landscape for several human malignancies. Antibodies against immune checkpoints, such as anti-PD-1/PD-L1 and anti-CTLA-4, demonstrate durable clinical benefits in several cancer types. However, checkpoint blockade has failed to elicit effective anti-tumor responses in pancreatic ductal adenocarcinoma (PDAC), which remains one of the most lethal malignancies with a dismal prognosis. As a result, there are significant efforts to identify novel immune-based combination regimens for PDAC, which are typically first tested in preclinical models. Here, we discuss the utility and limitations of syngeneic and genetically-engineered mouse models that are currently available for testing immunotherapy regimens. We also discuss patient-derived xenograft mouse models, human PDAC organoids, and ex vivo slice cultures of human PDAC tumors that can complement murine models for a more comprehensive approach to predict response and resistance to immunotherapy regimens