Echicetin, a GPIb-binding snake C-type lectin from Echis carinatus, also contains a binding site for IgMkappa responsible for platelet agglutination in plasma and inducing signal transduction

Echicetin, a heterodimeric snake C-type lectin from Echis carinatus, is known to bind specifically to platelet glycoprotein (GP)Ib. We now show that, in addition, it agglutinates platelets in plasma and induces platelet signal transduction. The agglutination is caused by binding to a specific protein in plasma. The protein was isolated from plasma and shown to cause platelet agglutination when added to washed platelets in the presence of echicetin. It was identified as immunoglobulin Mkappa (IgMkappa) by peptide sequencing and dot blotting with specific heavy and light chain anti-immunoglobulin reagents. Platelet agglutination by clustering echicetin with IgMkappa induced P-selectin expression and activation of GPIIb/IIIa as well as tyrosine phosphorylation of several signal transduction molecules, including p53/56(LYN), p64, p72(SYK), p70 to p90, and p120. However, neither ethylenediaminetetraacetic acid nor specific inhibition of GPIIb/IIIa affected platelet agglutination or activation by echicetin. Platelet agglutination and induction of signal transduction could also be produced by cross-linking biotinylated echicetin with avidin. These data indicate that clustering of GPIb alone is sufficient to activate platelets. In vivo, echicetin probably activates platelets rather than inhibits platelet activation, as previously proposed, accounting for the observed induction of thrombocytopenia.

Platelet activation and signal transduction by convulxin, a C-type lectin from Crotalus durissus terrificus (tropical rattlesnake) venom via the p62/GPVI collagen receptor

Convulxin, a powerful platelet activator, was isolated from Crotalus durissus terrificus venom, and 20 amino acid N-terminal sequences of both subunits were determined. These indicated that convulxin belongs to the heterodimeric C-type lectin family. Neither antibodies against GPIb nor echicetin had any effect on convulxin-induced platelet aggregation showing that, in contrast to other venom C-type lectins acting on platelets, GPIb is not involved in convulxin-induced platelet activation. In addition, partially reduced/denatured convulxin only affects collagen-induced platelet aggregation. The mechanism of convulxin-induced platelet activation was examined by platelet aggregation, detection of time-dependent tyrosine phosphorylation of platelet proteins, and binding studies with 125I-convulxin. Convulxin induces signal transduction in part like collagen, involving the time-dependent tyrosine phosphorylation of Fc receptor gamma chain, phospholipase Cgamma2, p72(SYK), c-Cbl, and p36-38. However, unlike collagen, pp125(FAK) and some other bands are not tyrosine-phosphorylated. Convulxin binds to a glycosylated 62-kDa membrane component in platelet lysate and to p62/GPVI immunoprecipitated by human anti-p62/GPVI antibodies. Convulxin subunits inhibit both aggregation and tyrosine phosphorylation in response to collagen. Piceatannol, a tyrosine kinase inhibitor with some specificity for p72(SYK), showed differential effects on collagen and convulxin-stimulated signaling. These results suggest that convulxin uses the p62/GPVI but not the alpha2beta1 part of the collagen signaling pathways to activate platelets. Occupation and clustering of p62/GPVI may activate Src family kinases phosphorylating Fc receptor gamma chain and, by a mechanism previously described in T- and B-cells, activate p72(SYK) that is critical for downstream activation of platelets.

Stable transduction of bovine TLR4 and bovine MD-2 into LPS-nonresponsive cells and soluble CD14 promote the ability to respond to LPS

The interaction of bovine cells with lipopolysaccharide (LPS) was explored using human embryo kidney (HEK) 293 cell line stably transduced with bovine toll-like receptor-4 (TLR4) alone or in combination with bovine MD-2. These lines and mock-transduced HEK293 cells were tested by flow cytometry for LPS-fluorescein isothiocyanate (LPS-FITC) binding, nuclear factor kappa B (NFkappaB) activation, interleukin-8 (IL-8) production and interferon-beta mRNA expression/interferon (IFN) type I production. Whereas bovine TLR4 was sufficient to promote binding of high concentrations of LPS-FITC, both bovine TLR4 and MD-2 were required for activation by LPS, as assessed by NFkappaB activation and IL-8 production. Induction of IFN bioactivity was not observed in doubly transduced HEK293 cells, and no evidence for IFN-beta mRNA induction in response to LPS was obtained, although cells responded by IFN-beta mRNA expression to stimulation by Sendai virus and poly-inosinic acid-poly-cytidylic acid (poly(I:C)). Cells stably transduced with both bovine TLR4 and bovine MD-2 responded to LPS by IL-8 production, in decreasing order, in the presence of fetal bovine serum (FCS), of human serum, and of human serum albumin (HSA). The reduced activity in the presence of HSA could be restored by the addition of soluble CD14 (sCD14) but not of LPS binding protein (LBP). This is in contrast to macrophages which show a superior response to LPS in the presence of HSA when compared with macrophages stimulated by LPS in the presence of FCS. This suggests that macrophages but not HEK293 cells express factors rendering LPS stimulation serum-independent. Stably double-transduced cells reacted, in decreasing order, to LPS from Rhodobacter sphaeroides, to LPS from Escherichia coli, to synthetic lipd-IVa (compound 406), to diphosphoryl-lipid-A (S. minnesota) and to monophosphoryl-lipid-A (S. minnesota). They failed to react to the murine MD-2/TLR4 ligand taxol. This resembles the reactivity of bovine macrophages with regard to sensitivity (ED(50)) and order of potency but is distinct from the reactivity pattern of other species. This formally establishes that in order to react to LPS, cattle cells require serum factors (e.g. sCD14) and cell-expressed factors such as MD-2 and TLR4. The cell lines described are the first of a series expressing defined pattern recognition receptors (PRR) of bovine origin. They will be useful in the study of the interaction of the bovine TLR4-MD-2 complex and Gram-negative bovine pathogens, e.g. the agents causing Gram-negative bovine mastitis.

Chlorhexidine and preservation of sound tooth structure in older adults. A placebo-controlled trial

The Trial to Enhance Elderly Teeth Health (TEETH) was designed to test the impact of regular rinsing with a 0.12% chlorhexidine (CHX) solution on tooth loss, and the causes of tooth loss (caries, periodontal disease and trauma) were also investigated. This paper reports on the effectiveness of a 0.12% CHX solution for controlling caries using a tooth surface (coronal and root) survival analysis. A total of 1,101 low income elders in Seattle (United States) and Vancouver (Canada), aged 60-75 years, were recruited for a double-blind clinical trial and assigned to either a CHX (n = 550) or a placebo (n = 551) mouth rinse. Subjects alternated between daily rinsing for 1 month, followed by weekly rinsing for 5 months. All sound coronal and root surfaces at baseline were followed annually for up to 5 years. At each follow-up examination, those tooth surfaces with caries, restored, or extracted were scored as 'carious'. The hazard ratio associated with CHX for a sound surface to become filled, decayed, or extracted was 0.87 for coronal surfaces (95% confidence interval: 0.71-1.14, p = 0.20) and 0.91 for root surfaces (95% confidence interval: 0.73-1.14, p = 0.41). These findings suggest that regular rinsing with CHX does not have a substantial effect on the preservation of sound tooth structure in older adults.

Sound As Artifact

A distinguishing feature of the discipline of archaeology is its reliance upon sensory dependant investigation. As perceived by all of the senses, the felt environment is a unique area of archaeological knowledge. It is generally accepted that the emergence of industrial processes in the recent past has been accompanied by unprecedented sonic extremes. The work of environmental historians has provided ample evidence that the introduction of much of this unwanted sound, or "noise" was an area of contestation. More recent research in the history of sound has called for more nuanced distinctions than the noisy/quiet dichotomy. Acoustic archaeology tends to focus upon a reconstruction of sound producing instruments and spaces with a primary goal of ascertaining intentionality. Most archaeoacoustic research is focused on learning more about the sonic world of people within prehistoric timeframes while some research has been done on historic sites. In this thesis, by way of a meditation on industrial sound and the physical remains of the Quincy Mining Company blacksmith shop (Hancock, MI) in particular, I argue for an acceptance and inclusion of sound as artifact in and of itself. I am introducing the concept of an individual sound-form, or sonifact, as a reproducible, repeatable, representable physical entity, created by tangible, perhaps even visible, host-artifacts. A sonifact is a sound that endures through time, with negligible variability. Through the piecing together of historical and archaeological evidence, in this thesis I present a plausible sonifactual assemblage at the blacksmith shop in April 1916 as it may have been experienced by an individual traversing the vicinity on foot: an 'historic soundwalk.' The sensory apprehension of abandoned industrial sites is multi-faceted. In this thesis I hope to make the case for an acceptance of sound as a primary heritage value when thinking about the industrial past, and also for an increased awareness and acceptance of sound and listening as a primary mode of perception.

Investigations into Velocity and Distance Perception Based on Different Types of Moving Sound Sources with Respect to Auditory Virtual Environments

The characteristics of moving sound sources have strong implications on the listener's distance perception and the estimation of velocity. Modifications of the typical sound emissions as they are currently occurring due to the tendency towards electromobility have an impact on the pedestrian's safety in road traffic. Thus, investigations of the relevant cues for velocity and distance perception of moving sound sources are not only of interest for the psychoacoustic community, but also for several applications, like e.g. virtual reality, noise pollution and safety aspects of road traffic. This article describes a series of psychoacoustic experiments in this field. Dichotic and diotic stimuli of a set of real-life recordings taken from a passing passenger car and a motorcycle were presented to test subjects who in turn were asked to determine the velocity of the object and its minimal distance from the listener. The results of these psychoacoustic experiments show that the estimated velocity is strongly linked to the object's distance. Furthermore, it could be shown that binaural cues contribute significantly to the perception of velocity. In a further experiment, it was shown that - independently of the type of the vehicle - the main parameter for distance determination is the maximum sound pressure level at the listener's position. The article suggests a system architecture for the adequate consideration of moving sound sources in virtual auditory environments. Virtual environments can thus be used to investigate the influence of new vehicle powertrain concepts and the related sound emissions of these vehicles on the pedestrians' ability to estimate the distance and velocity of moving objects.

EU Copyright Law, Lobbying and Transparency of Policy-Making: The cases of sound recordings term extension and orphan works provisions

The objective of this paper is to discuss EU lobbying in the area of copyright. Legislation needs to regulate the legal position of various different stakeholders in a balanced manner. However, a number of EU copyright provisions brought into effect over recent years were highly controversial and have led to suggestions that powerful lobbying forces may have had some influence. This article investigates the effects of lobbying on copyright law-making in Europe. A specific comparative and multi-faceted analysis is provided of the legislative process for two recently adopted directives: 2011/77/EU which extends the term of protection of sound recordings and 2012/28/EU which introduces certain permitted uses of orphan works (some references are also made to the ACTA case). Firstly, a short presentation is given of the legal bases for the EU consultation process and lobbying. Next, an analysis is provided of the two cases, taking into consideration the policy-making procedures (with special focus on how the consultation process was handled), the legal solutions proposed and adopted and the various stakeholders’ claims. Lastly, it asks why some interest groups were successful and some others failed (the analysis identifies two types of factor for the effectiveness of lobbying: those resulting from stakeholders’ actions and those connected with the consultation process).

Hearing performance with 2 different high-power sound processors for osseointegrated auditory implants

OBJECTIVE To compare speech understanding of the BAHA BP110 and BAHA Intenso sound processors. STUDY DESIGN Prospective experimental study. SETTING Tertiary referral center. PATIENTS Twenty experienced user of osseointegrated auditory implants with conductive or mixed hearing loss. INTERVENTIONS In a first session, half of the participants were fitted with an Intenso, the other half with a BP110. After 1 month of use, aided speech understanding in quiet and in noise was measured, and the other test processor was fitted. One month later, speech understanding with the second sound processor was assessed. MAIN OUTCOME MEASURES Speech understanding in quiet and in noise, with noise arriving either from the front, the rear, or the side of the user with the osseointegrated bone conductor. RESULTS Significant improvements were found for both processors for speech understanding in quiet (+9.6 to +34.8 percent points; p = 0.02 to 0.001) and in noise (+6.2 to +13.8 dB, p < 0.001). No significant differences were found between the 2 devices for speech in quiet. For noise from the rear, subjects were able to understand speech at signal-to-noise ratios which were lower (less favorable) by -5.1 dB (p < 0.001) when compared with the Intenso. CONCLUSION Speech understanding is substantially improved by both devices, with no significant differences between the sound processors in quiet. In noise, speech understanding is significantly better with the BP110 when compared to the Intenso for noise from the rear.

Nano-mechanical transduction of polymer micro-cantilevers to detect bio-molecular interactions

Using variothermal polymer micro-injection molding, disposable arrays of eight polymer micro-cantilevers each 500 μm long, 100 μm wide and 25 μm thick were fabricated. The present study took advantage of an easy flow grade polypropylene. After gold coating for optical read-out and asymmetrical sensitization, the arrays were introduced into the Cantisens(®) Research system to perform mechanical and functional testing. We demonstrate that polypropylene cantilevers can be used as biosensors for medical purposes in the same manner as the established silicon ones to detect single-stranded DNA sequences and metal ions in real-time. A differential signal of 7 nm was detected for the hybridization of 1 μM complementary DNA sequences. For 100 nM copper ions the differential signal was found to be (36 ± 5) nm. Nano-mechanical sensing of medically relevant, nanometer-size species is essential for fast and efficient diagnosis.