Accelerated flushing of contaminants from MSW landfill at the field scale using a fill-and-draw method.

A fill-and-draw flushing test on a landfill cell containing MSW waste was carried out to examine the operational viability of this method for accelerating the flushing of contaminants and landfill stabilisation. During the fill cycle, 800 m3 of water containing the tracer bromide was pumped into the base of a 0.44 ha landfill cell, resulting in the estimated saturation of 9,400 m3 of waste. Abstraction took place in two phases, during which 1,100 m3 of tracer/leachate was recovered. Samples of leachate were analysed for the tracer, electrical conductivity and indigenous solutes chloride and ammonia. Tracer recovery was between 63 and 72 % for bromide. An estimated 227 kg of ammonia and 575 kg of chloride were removed. Test data was used to calibrate a 1D, dual-porosity model involving advection in a mobile zone, and diffusion into 'blocks' of a less mobile zone. The model fitted well to the early time data, whereas later data appears to have been affected by recharge. The results of this trial demonstrate the possibilities of the 'fill-and-draw' concept using the basal leachate drainage system of landfills as a potential accelerated landfill remediation technique. However, modelling results suggest low contaminant removal efficiency. Including a pause between the fill and the draw cycles improves mass removal.

Antivesiculation and Complete Unbinding of Tail-Tethered Lipids.

We report the effect of tail-tethering on vesiculation and complete unbinding of bilayered membranes. Amphiphilic molecules of a bolalipid, resembling the tail-tethered molecular structure of archaeal lipids, with two identical zwitterionic phosphatidylcholine headgroups self-assemble into a large flat lamellar membrane, in contrast to the multilamellar vesicles (MLVs) observed in its counterpart, monopolar nontethered zwitterionic lipids. The antivesiculation is confirmed by small-angle X-ray scattering (SAXS) and cryogenic transmission electron microscopy (cyro-TEM). With the net charge of zero and higher bending rigidity of the membrane (confirmed by neutron spin echo (NSE) spectroscopy), the current membrane theory would predict that membranes should stack with each other (aka "bind") due to dominant van der Waals attraction, while the outcome of the nonstacking ("unbinding") membrane suggests that the theory needs to include entropic contribution for the nonvesicular structures. This report pioneers an understanding of how the tail-tethering of amphiphiles affects the structure, enabling better control over the final nanoscale morphology.

Pretreatment of Mesenchymal Stem Cells with Electrical Stimulation as a Strategy to Improve Bone Tissue Engineering Outcomes.

Electrical stimulation (EStim), whether used alone or in combination with bone tissue engineering (BTE) approaches, has been shown to promote bone healing. In our previous in vitro studies, mesenchymal stem cells (MSCs) were exposed to EStim and a sustained, long-lasting increase in osteogenic activity was observed. Based on these findings, we hypothesized that pretreating MSC with EStim, in 2D or 3D cultures, before using them to treat large bone defects would improve BTE treatments. Critical size femur defects were created in 120 Sprague-Dawley rats and treated with scaffold granules seeded with MSCs that were pre-exposed or not (control group) to EStim 1 h/day for 7 days in 2D (MSCs alone) or 3D culture (MSCs + scaffolds). Bone healing was assessed at 1, 4, and 8 weeks post-surgery. In all groups, the percentage of new bone increased, while fibrous tissue and CD68+ cell count decreased over time. However, these and other healing features, like mineral density, bending stiffness, the amount of new bone and cartilage, and the gene expression of osteogenic markers, did not significantly differ between groups. Based on these findings, it appears that the bone healing environment could counteract the long-term, pro-osteogenic effects of EStim seen in our in vitro studies. Thus, EStim seems to be more effective when administered directly and continuously at the defect site during bone healing, as indicated by our previous studies.

Discovery of IACS-52825, a Potent and Selective DLK Inhibitor for Treatment of Chemotherapy-Induced Peripheral Neuropathy.

Chemotherapy-induced peripheral neuropathy (CIPN) is a major unmet medical need with limited treatment options. Despite different mechanisms of action, diverse chemotherapeutics can cause CIPN through a converged pathway─an active axon degeneration program that engages the dual leucine zipper kinase (DLK). DLK is a neuronally enriched kinase upstream in the MAPK-JNK cascade, and while it is dormant under physiological conditions, DLK mediates a core mechanism for neuronal injury response under stress conditions, making it an attractive target for treatment of neuronal injury and neurodegenerative diseases. We have developed potent, selective, brain penetrant DLK inhibitors with excellent PK and activity in mouse models of CIPN. Lead compound IACS-52825 (22) showed strongly effective reversal of mechanical allodynia in a mouse model of CIPN and was advanced into preclinical development.

Structural insights into selective small molecule activation of PKG1α.

cGMP-dependent protein kinase I-α (PKG1α) is a target for pulmonary arterial hypertension due to its role in the regulation of smooth muscle function. While most work has focused on regulation of cGMP turnover, we recently described several small molecule tool compounds which were capable of activating PKG1α via a cGMP independent pathway. Selected molecules were crystallized in the presence of PKG1α and were found to bind to an allosteric site proximal to the low-affinity nucleotide binding domain. These molecules act to displace the switch helix and cause activation of PKG1α representing a new mechanism for the activation and control of this critical therapeutic path. The described structures are vital to understanding the function and control of this key regulatory pathway.

Intensive hands-on microsurgery course provides a solid foundation for performing clinical microvascular surgery.

PURPOSE: Microvascular surgery requires highly specialized and individualized training; most surgical residency training programs are not equipped with microsurgery teaching expertise and/or facilities. The aim of this manuscript was to describe the methodology and clinical effectiveness of an international microsurgery course, currently taught year-round in eight countries. METHODS: In the 5-day microsurgery course trainees perform arterial and venous end-to-end, end-to-side, one-way-up, and continuous suture anastomoses and vein graft techniques in live animals, supported by video demonstrations and hands-on guidance by a full-time instructor. To assess and monitor each trainee's progress, the course's effectiveness is evaluated using "in-course" evaluations, and participant satisfaction and clinical relevance are assessed using a "post-course" survey. RESULTS: Between 2007 and 2017, more than 600 trainees participated in the microsurgery course. "In-course" evaluations of patency rates revealed 80.3% (arterial) and 39% (venous) performed in end-to-end, 82.7% in end-to-side, 72.6% in continuous suture, and 89.5% (arterial) and 62.5% (venous) one-way-up anastomoses, and 58.1% in vein graft technique. "Post-course" survey results indicated that participants considered the most important components of the microcourse to be "practicing on live animals", followed by "the presence of a full-time instructor". In addition, almost all respondents indicated that they were more confident performing clinical microsurgery cases after completing the course. CONCLUSIONS: Microvascular surgery requires highly specialized and individualized training to achieve the competences required to perform and master the delicate fine motor skills necessary to successfully handle and anastomose very small and delicate microvascular structures. The ever-expanding clinical applications of microvascular procedures has led to an increased demand for training opportunities. By teaching time-tested basic motor skills that form the foundation of microsurgical technique this international microsurgery-teaching course is helping to meet this demand.

Facile polymerization in a bicellar template to produce polymer nano-rings.

HYPOTHESIS: A well-defined discoidal bicelle composed of three lipids, specifically zwitterionic long-chain 1,2­dipalmitoyl phosphocholine (DPPC) and short-chain 1,2­dihexanoyl phosphocholine (DHPC) doped with anionic 1,2­dipalmitoyl phosphoglycerol (DPPG) provides a generalized template for the synthesis of hydrophobic polymer nano-rings. The lipid molar ratio of DPPC/DHPC/DPPG is 0.71/0.25/0.04. The detailed investigation and discussion were based on styrene but tested on three other vinyl monomers. EXPERIMENTS: The structure of nano-rings is identified through the detailed analysis of small angle X-ray/neutron scattering (SAXS and SANS) data and transmission electron micrographs (TEM), supported by the differential scanning calorimetric (DSC) data before and after polymerization. The investigation covers samples with a styrene-to-lipid ratio ranged varied from 1:50 to 1:10. FINDINGS: The styrene monomers are initially located at both the discoidal planar (long-chain lipid rich) and rim (short-chain lipid rich) regions. During polymerization, they migrate to the more fluid rim regionsection. The formation mechanism involves the interplay of hydrophobic interaction, mismatched miscibility of polystyrene between the ordered and disordered phases, and crystallinity of the long lipid acyl chains. This facile synthesis is proven applicable for several hydrophobic monomers. The well-defined nano-rings greatly enhance the interfacial area and have the potential to be the building blocks for functional materials, if monomers are incorporated with desirable functions, for future applications.

Optimization and Mechanistic Investigations of Novel Allosteric Activators of PKG1α.

Activation of PKG1α is a compelling strategy for the treatment of cardiovascular diseases. As the main effector of cyclic guanosine monophosphate (cGMP), activation of PKG1α induces smooth muscle relaxation in blood vessels, lowers pulmonary blood pressure, prevents platelet aggregation, and protects against cardiac stress. The development of activators has been mostly limited to cGMP mimetics and synthetic peptides. Described herein is the optimization of a piperidine series of small molecules to yield activators that demonstrate in vitro phosphorylation of vasodilator-stimulated phosphoprotein as well as antiproliferative effects in human pulmonary arterial smooth muscle cells. Hydrogen/deuterium exchange mass spectrometry experiments with the small molecule activators revealed a mechanism of action consistent with cGMP-induced activation, and an X-ray co-crystal structure with a construct encompassing the regulatory domains illustrated a binding mode in an allosteric pocket proximal to the low-affinity cyclic nucleotide-binding domain.

The very small angle neutron scattering instrument at the National Institute of Standards and Technology.

A description and the performance of the very small angle neutron scattering diffractometer at the National Institute of Standards and Technology are presented. The measurement range of the instrument extends over three decades of momentum transfer q from 2 × 10-4 to 0.7 Å-1. The entire scattering angle range from 8 × 10-5 to π/6 rad (30°) can be measured simultaneously using three separate detector carriages on rails holding nine 2D detector arrays. Versatile choices of collimation options and neutron wavelength selection allow the q resolution and beam intensity to be optimized for the needs of the experiment. High q resolution is achieved using multiple converging-beam collimation with circular pinholes combined with refractive lenses and prisms. Relaxed vertical resolution with much higher beam intensity can be achieved with narrow slit collimation and a broad wavelength range chosen by truncating the moderator source distribution below 4 Šwith a Be crystalline filter and above 8 Šwith a supermirror deflector. Polarized beam measurements with full polarization analysis are also provided by a high-performance supermirror polarizer and spin flipper, capable of producing flipping ratios of over 100, along with a high-efficiency 3He polarization analyzer.

CH2 + O2: reaction mechanism, biradical and zwitterionic character, and formation of CH2OO, the simplest Criegee intermediate.

The singlet and triplet potential surfaces for the title reaction were investigated using the CBS-QB3 level of theory. The wave functions for some species exhibited multireference character and required the CASPT2/6-31+G(d,p) and CASPT2/aug-cc-pVTZ levels of theory to obtain accurate relative energies. A Natural Bond Orbital Analysis showed that triplet 3CH2OO (the simplest Criegee intermediate) and 3CH2O2 (dioxirane) have mostly polar biradical character, while singlet 1CH2OO has some zwitterionic character and a planar structure. Canonical variational transition state theory (CVTST) and master equation simulations were used to analyze the reaction system. CVTST predicts that the rate constant for reaction of 1CH2 + 3O2 is more than ten times as fast as the reaction of 3CH2 (X3B1) + 3O2 and the ratio remains almost independent of temperature from 900 K to 3000 K. The master equation simulations predict that at low pressures the 1CH2O + 3O product set is dominant at all temperatures and the primary yield of OH radicals is negligible below 600 K, due to competition with other primary reactions in this complex system.

Discovery of Novel UDP-N-Acetylglucosamine Acyltransferase (LpxA) Inhibitors with Activity against Pseudomonas aeruginosa.

This study describes a novel series of UDP-N-acetylglucosamine acyltransferase (LpxA) inhibitors that was identified through affinity-mediated selection from a DNA-encoded compound library. The original hit was a selective inhibitor of Pseudomonas aeruginosa LpxA with no activity against Escherichia coli LpxA. The biochemical potency of the series was optimized through an X-ray crystallography-supported medicinal chemistry program, resulting in compounds with nanomolar activity against P. aeruginosa LpxA (best half-maximal inhibitory concentration (IC50) <5 nM) and cellular activity against P. aeruginosa (best minimal inhibitory concentration (MIC) of 4 µg/mL). Lack of activity against E. coli was maintained (IC50 > 20 µM and MIC > 128 µg/mL). The mode of action of analogues was confirmed through genetic analyses. As expected, compounds were active against multidrug-resistant isolates. Further optimization of pharmacokinetics is needed before efficacy studies in mouse infection models can be attempted. To our knowledge, this is the first reported LpxA inhibitor series with selective activity against P. aeruginosa.

Mitigating background caused by extraneous scattering in small-angle neutron scattering instrument design.

Measurements, calculations and design ideas to mitigate background caused by extraneous scattering in small-angle neutron scattering (SANS) instruments are presented. Scattering includes processes such as incoherent scattering, inelastic scattering and Bragg diffraction. Three primary sources of this type of background are investigated: the beam stop located in front of the detector, the inside lining of the detector vessel and the environment surrounding the sample. SANS measurements were made where materials with different albedos were placed in all three locations. Additional measurements of the angle-dependent scattering over the angular range of 0.7π-0.95π rad were completed on 16 different shielding materials at five wavelengths. The data were extrapolated to cover scattering angles from π/2 to π rad in order to estimate the materials' albedos. Modifications to existing SANS instruments and sample environments to mitigate extraneous scattering from surfaces are discussed.

Development of a robust crystallization platform for immune receptor TREM2 using a crystallization chaperone strategy.

TREM2 has been identified by genomic analysis as a potential and novel target for the treatment of Alzheimer's disease. To enable structure-based screening of potential small molecule therapeutics, we sought to develop a robust crystallization platform for the TREM2 Ig-like domain. A systematic set of constructs containing the structural chaperone, maltose binding protein (MBP), fused to the Ig domain of TREM2, were evaluated in parallel expression and purification, followed by crystallization studies. Using protein crystallization and high-resolution diffraction as a readout, a MBP-TREM2 Ig fusion construct was identified that generates reproducible protein crystals diffracting at 2.0 Å, which makes it suitable for soaking of potential ligands. Importantly, analysis of crystal packing interfaces indicates that most of the surface of the TREM2 Ig domain is available for small molecule binding. A proof of concept co-crystallization study with a small library of fragments validated potential utility of this system for the discovery of new TREM2 therapeutics.

Electrical Stimulation Decreases Dental Pulp Stem Cell Osteo-/Odontogenic Differentiation.

Dental pulp stem cells (DPSCs) have great potential for use in tissue engineering (TE)-based dental treatments. Electrical stimulation (EStim) has been shown to influence cellular functions that could play an important role in the success of TE treatments. Despite many recent studies focused on DPSCs, few have investigated the effect EStim has on these cells. The aim of this research was to investigate the effects of direct current (DC) EStim on osteo-/odontogenic differentiation of DPSCs. To do so cells were isolated from male Sprague Dawley rats (7-8 weeks old), and phenotype characterization and multilineage differentiation analysis were conducted to verify their "stemness." Different voltages of DC EStim were administrated 1 h/day for 7 days, and the effect of EStim on DPSC osteo-/odontogenic differentiation was assessed by measuring calcium and collagen deposition, alkaline phosphatase (ALP) activity, and expression of osteo- and odontogenic marker genes at days 7 and 14 of culture. We found that while 10 and 50 mV/mm of EStim had no effect on cell number or metabolic activity, 100 mV/mm caused a significant reduction in cell number, and 150 mV/mm resulted in cell death. Despite increased gene expression of osteo-/odontogenic gene markers, Osteocalcin, RunX2, BSP, and DMP1, at day 7 in EStim treated cells, 50 mV/mm of EStim decreased collagen deposition and ALP activity at both time points, and calcium deposition was found to be lower at day 14. In conclusion, under the conditions tested, EStim appears to impair DPSC osteo-/odontogenic differentiation. Additional studies are needed to further characterize and understand the mechanisms involved in DPSC response to EStim, with an eye toward its potential use in TE-based dental treatments.

Quantum Transport in a Silicon Nanowire FET Transistor: Hot Electrons and Local Power Dissipation.

A review and perspective is presented of the classical, semiclassical and fully quantum routes to the simulation of electrothermal phenomena in ultrascaled silicon nanowire fieldeffect transistors. It is shown that the physics of ultrascaled devices requires at least a coupled electron quantum transport semiclassical heat equation model outlined here. The importance of the local density of states (LDOS) is discussed from classical to fully quantum versions. It is shown that the minimal quantum approach requires selfconsistency with the Poisson equation and that the electronic LDOS must be determined within at least the selfconsistent Born approximation. To bring in this description and to provide the energy resolved local carrier distributions it is necessary to adopt the nonequilibrium Green function (NEGF) formalism, briefly surveyed here. The NEGF approach describes quantum coherent and dissipative transport, Pauli exclusion and nonequilibrium conditions inside the device. There are two extremes of NEGF used in the community. The most fundamental is based on coupled equations for the Green functions electrons and phonons that are computed at the atomically resolved level within the nanowire channel and into the surrounding device structure using a tight binding Hamiltonian. It has the advantage of treating both the nonequilibrium heat flow within the electron and phonon systems even when the phonon energy distributions are not described by a temperature model. The disadvantage is the grand challenge level of computational complexity. The second approach, that we focus on here, is more useful for fast multiple simulations of devices important for TCAD (Technology Computer Aided Design). It retains the fundamental quantum transport model for the electrons but subsumes the description of the energy distribution of the local phonon subsystem statistics into a semiclassical Fourier heat equation that is sourced by the local heat dissipation from the electron system. It is shown that this selfconsistent approach retains the salient features of the fullscale approach. For focus, we outline our electrothermal simulations for a typical narrow Si nanowire gate allaround fieldeffect transistor. The selfconsistent Born approximation is used to describe electronphonon scattering as the source of heat dissipation to the lattice. We calculated the effect of the device selfheating on the current voltage characteristics. Our fast and simpler methodology closely reproduces the results of a more fundamental computeintensive calculations in which the phonon system is treated on the same footing as the electron system. We computed the local power dissipation and "local lattice temperature" profiles. We compared the selfheating using hot electron heating and the Joule heating, i.e., assuming the electron system was in local equilibrium with the potential. Our simulations show that at low bias the source region of the device has a tendency to cool down for the case of the hot electron heating but not for the case of Joule heating. Our methodology opens the possibility of studying thermoelectricity at nanoscales in an accurate and computationally efficient way. At nanoscales, coherence and hot electrons play a major role. It was found that the overall behaviour of the electron system is dominated by the local density of states and the scattering rate. Electrons leaving the simulated drain region were found to be far from equilibrium.

Role of Bioelectricity During Cell Proliferation in Different Cell Types.

Most living organisms possess varying degrees of regenerative capabilities but how these regenerative processes are controlled is still poorly understood. Naturally occurring bioelectric voltages (like Vmem) are thought to be playing instructive role in tissue regeneration, as well as embryonic development. The different distribution of ions on the either side of the cell membrane results in intra- and extra-cellular voltage differences, known as membrane potential or Vmem. The relationship between Vmem and cell physiology is conserved in a wide range of cell types and suggests that Vmem regulation is a fundamental control mechanism for regeneration related processes e.g., proliferation and differentiation. In the present study we measured Vmem in three different cell types (human osteogenic sarcoma cell line (OSC), rat bone marrow derived mesenchymal stem cells (BM-MSC), and rat dermal fibroblasts) and characterized the relationship between their Vmem and proliferation. In order to find out if Vmem controls proliferation, or visa-versa, we blocked and then unblocked Na+/K+-exchanging ATPase using ouabain and measured the proliferation. Our results demonstrate that Vmem can be pharmacologically manipulated to control proliferation in certain cell types like BM-MSC. Taken together, it is clear that control of bioelectrical properties in non-excitable cells could prove to be potentially a useful tool in regenerative medicine efforts.

Electrical stimulation in bone tissue engineering treatments.

Electrical stimulation (EStim) has been shown to promote bone healing and regeneration both in animal experiments and clinical treatments. Therefore, incorporating EStim into promising new bone tissue engineering (BTE) therapies is a logical next step. The goal of current BTE research is to develop combinations of cells, scaffolds, and chemical and physical stimuli that optimize treatment outcomes. Recent studies demonstrating EStim's positive osteogenic effects at the cellular and molecular level provide intriguing clues to the underlying mechanisms by which it promotes bone healing. In this review, we discuss results of recent in vitro and in vivo research focused on using EStim to promote bone healing and regeneration and consider possible strategies for its application to improve outcomes in BTE treatments. Technical aspects of exposing cells and tissues to EStim in in vitro and in vivo model systems are also discussed.

Effects of Electrical Stimulation on Stem Cells.

Recent interest in developing new regenerative medicine- and tissue engineering-based treatments has motivated researchers to develop strategies for manipulating stem cells to optimize outcomes in these potentially, game-changing treatments. Cells communicate with each other, and with their surrounding tissues and organs via electrochemical signals. These signals originate from ions passing back and forth through cell membranes and play a key role in regulating cell function during embryonic development, healing, and regeneration. To study the effects of electrical signals on cell function, investigators have exposed cells to exogenous electrical stimulation and have been able to increase, decrease and entirely block cell proliferation, differentiation, migration, alignment, and adherence to scaffold materials. In this review, we discuss research focused on the use of electrical stimulation to manipulate stem cell function with a focus on its incorporation in tissue engineering-based treatments.