Role of Tyrosine Kinase Syk in Thrombus Stabilisation at High Shear

Understanding the pathways involved in the formation and stability of the core and shell regions of a platelet-rich arterial thrombus may result in new ways to treat arterial thrombosis. The distinguishing feature between these two regions is the absence of fibrin in the shell which indicates that in vitro flow-based assays over thrombogenic surfaces, in the absence of coagulation, can be used to resemble this region. In this study, we have investigated the contribution of Syk tyrosine kinase in the stability of platelet aggregates (or thrombi) formed on collagen or atherosclerotic plaque homogenate at arterial shear (1000 s⁻¹). We show that post-perfusion of the Syk inhibitor PRT-060318 over preformed thrombi on both surfaces enhances thrombus breakdown and platelet detachment. The resulting loss of thrombus stability led to a reduction in thrombus contractile score which could be detected as early as 3 min after perfusion of the Syk inhibitor. A similar loss of thrombus stability was observed with ticagrelor and indomethacin, inhibitors of platelet adenosine diphosphate (ADP) receptor and thromboxane A₂ (TxA₂), respectively, and in the presence of the Src inhibitor, dasatinib. In contrast, the Btk inhibitor, ibrutinib, causes only a minor decrease in thrombus contractile score. Weak thrombus breakdown is also seen with the blocking GPVI nanobody, Nb21, which indicates, at best, a minor contribution of collagen to the stability of the platelet aggregate. These results show that Syk regulates thrombus stability in the absence of fibrin in human platelets under flow and provide evidence that this involves pathways additional to activation of GPVI by collagen.     

Hippocampal Expression of Cytochrome P450 1B1 in Penetrating Traumatic Brain Injury

Hippocampal dysfunction contributes to multiple traumatic brain injury sequala. Female rodents’ outcome is superior to male which has been ascribed the neuroprotective sex hormones 17β-estradiol and progesterone. Cytochrome P450 1B1 (CYP1B1) is an oxidative enzyme influencing the neuroinflammatory response by creating inflammatory mediators and metabolizing neuroprotective 17β-estradiol and progesterone. In this study, we aimed to describe hippocampal CYP1B1 mRNA expression, protein presence of CYP1B1 and its key redox partner Cytochrome P450 reductase (CPR) in both sexes, as well as the effect of penetrating traumatic brain injury (pTBI). A total 64 adult Sprague Dawley rats divided by sex received pTBI or sham-surgery and were assigned survival times of 1-, 3-, 5- or 7 days. CYP1B1 mRNA was quantified using in-situ hybridization and immunohistochemistry performed to verify protein colocalization. CYP1B1 mRNA expression was present in all subregions but greatest in CA2 irrespective of sex, survival time or intervention. At 3-, 5- and 7 days post-injury, expression in CA2 was reduced in male rats subjected to pTBI compared to sham-surgery. Females subjected to pTBI instead exhibited increased expression in all CA subregions 3 days post-injury, the only time point expression in CA2 was greater in females than in males. Immunohistochemical analysis confirmed neuronal CYP1B1 protein in all hippocampal subregions, while CPR was limited to CA1 and CA2. CYP1B1 mRNA is constitutively expressed in both sexes. In response to pTBI, females displayed a more urgent but brief regulatory response than males. This indicates there may be sex-dependent differences in CYP1B1 activity, possibly influencing inflammation and neuroprotection in pTBI.     

Influence of Ignition Sources on Heat Release Rate in the Furniture Calorimeter

When measuring the production rates of heat and smoke of upholstered furniture the ignition source must not influence the test results. In this paper variations in the strength, application time and geometry of the ignition burner used in the furniture calorimeter are studied. Results from replicate furniture calorimeter chair test show that the burning behaviour is almost independent of the burner intensity in the applied regime. Tests on six different furniture items were performed in the furniture calorimeter. The test objects were exposed to three intensity levels of ignition source. The sources were: a large propane burner giving 30 kW during 120 s and a smaller propane burner used at two levels of heat output, 1.7 and 5.8 kW during 90 s. The results showed that the burning behaviour of the furniture was very similar regardless of which burner was used. This was especially evident when the time regime between 50 and 400 kW was studied. The length of this period is a measurement on how quick untenable conditions are developing in a single, well-ventilated compartment.     

Mucoadhesive Electrospun Nanofiber-Based Hybrid System with Controlled and Unidirectional Release of Desmopressin

The sublingual mucosa is an attractive route for drug delivery, although challenged by a continuous flow of saliva that leads to a loss of drug by swallowing. It is of great benefit that drugs absorbed across the sublingual mucosa avoid exposure to the harsh environment of the gastro-intestinal lumen; this is especially beneficial for drugs of low physicochemical stability such as therapeutic peptides. In this study, a two-layered hybrid drug delivery system was developed for the sublingual delivery of the therapeutic peptide desmopressin. It consisted of peptide-loaded mucoadhesive electrospun chitosan/polyethylene oxide-based nanofibers (mean diameter of 183 ± 20 nm) and a saliva-repelling backing film to promote unidirectional release towards the mucosa. Desmopressin was released from the nanofiber-based hybrid system (approximately 80% of the loaded peptide was released within 45 min) in a unidirectional manner in vitro. Importantly, the nanofiber–film hybrid system protected the peptide from wash-out, as demonstrated in an ex vivo flow retention model with porcine sublingual mucosal tissue. Approximately 90% of the loaded desmopressin was retained at the surface of the ex vivo porcine sublingual mucosa after 15 min of exposure to flow rates representing salivary flow.     

Model selection, identification and validation in anaerobic digestion: a review

Anaerobic digestion enables waste (water) treatment and energy production in the form of biogas. The successful implementation of this process has lead to an increasing interest worldwide. However, anaerobic digestion is a complex biological process, where hundreds of microbial populations are involved, and whose start-up and operation are delicate issues. In order to better understand the process dynamics and to optimize the operating conditions, the availability of dynamic models is of paramount importance. Such models have to be inferred from prior knowledge and experimental data collected from real plants. Modeling and parameter identification are vast subjects, offering a realm of approaches and methods, which can be difficult to fully understand by scientists and engineers dedicated to the plant operation and improvements. This review article discusses existing modeling frameworks and methodologies for parameter estimation and model validation in the field of anaerobic digestion processes. The point of view is pragmatic, intentionally focusing on simple but efficient methods.     

Multi-level Modeling of Silica–Template Interactions During Initial Stages of Zeolite Synthesis

Zeolite synthesis is driven by structure-directing agents, such as tetrapropyl ammonium ions (TPA⁺) for Silicalite-1 and ZSM-5. However, the guiding role of these organic templates in the complex assembly to highly ordered frameworks remains unclear, limiting the prospects for advanced material synthesis. In this work, both static ab initio and dynamic classical modeling techniques are employed to provide insight into the interactions between TPA⁺ and Silicalite-1 precursors. We find that as soon as the typical straight 10-ring channel of Silicalite-1 or ZSM-5 is formed from smaller oligomers, the TPA⁺ template is partially squeezed out of the resulting cavity. Partial retention of the template in the cavity is, however, indispensable to prevent collapse of the channel and subsequent hydrolysis.

Development and validation of a new LC–MS/MS method for the simultaneous determination of six major ergot alkaloids and their corresponding epimers. Application to some food and feed commodities

A liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated for the simultaneous determination of six major ergot alkaloids, ergometrine, ergosine, ergotamine, ergocornine, ergokryptine and ergocristine, as well as their corresponding epimers in food and feed samples. The method involves extraction under alkaline conditions and subsequent clean-up by applying a simple and rapid liquid–liquid partitioning procedure prior to LC–MS/MS analysis. Evaluation of the method revealed good linearity, accuracy and precision. The limits of quantification varied from 0.1 to 1 μg/kg depending on the analyte and matrix. The average extraction and clean-up recoveries in different matrices were between 45 (only for ergometrine in biscuit) and 90%. The uncertainty associated with the analytical method was not higher than 51% and 30%, at concentration levels of 2.5 and 150 μg/kg respectively. Analyte epimerization proved to be minimal during the analytical procedure. The method has been successfully applied to the determination of ergot alkaloids in some Belgian food and feed commodities. Ergot alkaloids were found in 104 out of 122 samples investigated. Ergosine was the most frequently occurring alkaloid, while the highest levels were observed for ergotamine, ergocristine or ergosine, depending on the product type. The total alkaloid content in positive samples varied from 1 to 1145 μg/kg.     

Model selection and optimal sampling in high-throughput experimentation

The practical difficulties encountered in analyzing the kinetics of new reactions are considered from the viewpoint of the capabilities of state-of-the-art high-throughput systems. There are three problems. The first problem is that of model selection, i.e., choosing the correct reaction rate law. The second problem is how to obtain good estimates of the reaction parameters using only a small number of samples once a kinetic model is selected. The third problem is how to perform both functions using just one small set of measurements. To solve the first problem, we present an optimal sampling protocol to choose the correct kinetic model for a given reaction, based on T-optimal design. This protocol is then tested for the case of second-order and pseudo-first-order reactions using both experiments and computer simulations. To solve the second problem, we derive the information function for second-order reactions and use this function to find the optimal sampling points for estimating the kinetic constants. The third problem is further complicated by the fact that the optimal measurement times for determining the correct kinetic model differ from those needed to obtain good estimates of the kinetic constants. To solve this problem, we propose a Pareto optimal approach that can be tuned to give the set of best possible solutions for the two criteria. One important advantage of this approach is that it enables the integration of a priori knowledge into the workflow.     

Patterning of Epitaxial Perovskites from Micro and Nano Molded Stencil Masks

A process is developed that combines soft lithographic molding with pulsed laser deposition (PLD) to make heteroepitaxial patterns of functional perovskite oxide materials. Micro‐ and nanostructures of sacrificial ZnO are made by micro molding in capillaries (MiMiC) and nano transfer molding, respectively, and used to screen the single crystalline substrates during subsequent PLD. ZnO is used because of its compatibility with the high temperatures reached during PLD and because of the ease of its removal after use by benefiting from its amphoteric nature. Sub‐micrometer sized lines of La_0.67Sr_0.33MnO₃ are made by the transfer molding approach, preserving the anisotropic features expected for a fully oriented thin film and taking account for the magnetostatic contribution from the line shapes. Different patterns of SrRuO₃ are made with lateral dimensions of a few micrometers having individual features for which electrical isolation is illustrated. The bottom‐up soft lithographic methods can be compliantly utilized for making epitaxial structures of various shapes and sizes in the μm down to the nm range, and offer unique opportunities for fundamental studies as well as for realizing technological applications.