A Suite of Tutorials for the WESTPA 2.0 Rare-Events Sampling Software [Article v2.0].

The weighted ensemble (WE) strategy has been demonstrated to be highly efficient in generating pathways and rate constants for rare events such as protein folding and protein binding using atomistic molecular dynamics simulations. Here we present two sets of tutorials instructing users in the best practices for preparing, carrying out, and analyzing WE simulations for various applications using the WESTPA software. The first set of more basic tutorials describes a range of simulation types, from a molecular association process in explicit solvent to more complex processes such as host-guest association, peptide conformational sampling, and protein folding. The second set ecompasses six advanced tutorials instructing users in the best practices of using key new features and plugins/extensions of the WESTPA 2.0 software package, which consists of major upgrades for larger systems and/or slower processes. The advanced tutorials demonstrate the use of the following key features: (i) a generalized resampler module for the creation of "binless" schemes, (ii) a minimal adaptive binning scheme for more efficient surmounting of free energy barriers, (iii) streamlined handling of large simulation datasets using an HDF5 framework, (iv) two different schemes for more efficient rate-constant estimation, (v) a Python API for simplified analysis of WE simulations, and (vi) plugins/extensions for Markovian Weighted Ensemble Milestoning and WE rule-based modeling for systems biology models. Applications of the advanced tutorials include atomistic and non-spatial models, and consist of complex processes such as protein folding and the membrane permeability of a drug-like molecule. Users are expected to already have significant experience with running conventional molecular dynamics or systems biology simulations.

pH-dependent water permeability switching and its memory in MoS2 membranes.

Intelligent transport of molecular species across different barriers is critical for various biological functions and is achieved through the unique properties of biological membranes1-4. Two essential features of intelligent transport are the ability to (1) adapt to different external and internal conditions and (2) memorize the previous state5. In biological systems, the most common form of such intelligence is expressed as hysteresis6. Despite numerous advances made over previous decades on smart membranes, it remains a challenge to create a synthetic membrane with stable hysteretic behaviour for molecular transport7-11. Here we demonstrate the memory effects and stimuli-regulated transport of molecules through an intelligent, phase-changing MoS2 membrane in response to external pH. We show that water and ion permeation through 1T' MoS2 membranes follows a pH-dependent hysteresis with a permeation rate that switches by a few orders of magnitude. We establish that this phenomenon is unique to the 1T' phase of MoS2, due to the presence of surface charge and exchangeable ions on the surface. We further demonstrate the potential application of this phenomenon in autonomous wound infection monitoring and pH-dependent nanofiltration. Our work deepens understanding of the mechanism of water transport at the nanoscale and opens an avenue for the development of intelligent membranes.

WESTPA 2.0: High-Performance Upgrades for Weighted Ensemble Simulations and Analysis of Longer-Timescale Applications.

The weighted ensemble (WE) family of methods is one of several statistical mechanics-based path sampling strategies that can provide estimates of key observables (rate constants and pathways) using a fraction of the time required by direct simulation methods such as molecular dynamics or discrete-state stochastic algorithms. WE methods oversee numerous parallel trajectories using intermittent overhead operations at fixed time intervals, enabling facile interoperability with any dynamics engine. Here, we report on the major upgrades to the WESTPA software package, an open-source, high-performance framework that implements both basic and recently developed WE methods. These upgrades offer substantial improvements over traditional WE methods. The key features of the new WESTPA 2.0 software enhance the efficiency and ease of use: an adaptive binning scheme for more efficient surmounting of large free energy barriers, streamlined handling of large simulation data sets, exponentially improved analysis of kinetics, and developer-friendly tools for creating new WE methods, including a Python API and resampler module for implementing both binned and "binless" WE strategies.

Establishing an SEIR-based framework for local modelling of COVID-19 infections, hospitalisations and deaths.

Without timely assessments of the number of COVID-19 cases requiring hospitalisation, healthcare providers will struggle to ensure an appropriate number of beds are made available. Too few could cause excess deaths while too many could result in additional waits for elective treatment. As well as supporting capacity considerations, reliably projecting future "waves" is important to inform the nature, timing and magnitude of any localised restrictions to reduce transmission. In making the case for locally owned and locally configurable models, this paper details the approach taken by one major healthcare system in founding a multi-disciplinary "Scenario Review Working Group", comprising commissioners, public health officials and academic epidemiologists. The role of this group, which met weekly during the pandemic, was to define and maintain an evolving library of plausible scenarios to underpin projections obtained through an SEIR-based compartmental model. Outputs have informed decision-making at the system's major incident Bronze, Silver and Gold Commands. This paper presents illustrated examples of use and offers practical considerations for other healthcare systems that may benefit from such a framework.

TRAFIC: statistical design and analysis plan for a pragmatic early phase 1/2 Bayesian adaptive dose escalation trial in rheumatoid arthritis.

BACKGROUND: Adaptive model-based dose-finding designs have demonstrated advantages over traditional rule-based designs but have increased statistical complexity but uptake has been slow especially outside of cancer trials. TRAFIC is a multi-centre, early phase trial in rheumatoid arthritis incorporating a model-based design. METHODS: A Bayesian adaptive dose-finding phase I trial rolling into a single-arm, single-stage phase II trial. Model parameters for phase I were chosen via Monte Carlo simulation evaluating objective performance measures under clinically relevant scenarios and incorporated stopping rules for early termination. Potential designs were further calibrated utilising dose transition pathways. DISCUSSION: TRAFIC is an MRC-funded trial of a re-purposed treatment demonstrating that it is possible to design, fund and implement a model-based phase I trial in a non-cancer population within conventional research funding tracks and regulatory constraints. The phase I design allows borrowing of information from previous trials, all accumulated data to be utilised in decision-making, verification of operating characteristics through simulation, improved understanding for management and oversight teams through dose transition pathways. The rolling phase II design brings efficiencies in trial conduct including site and monitoring activities and cost. TRAFIC is the first funded model-based dose-finding trial in inflammatory disease demonstrating that small phase I/II trials can have an underlying statistical basis for decision-making and interpretation. TRIAL REGISTRATION: Trials Registration: ISRCTN, ISRCTN36667085 . Registered on September 26, 2014.

Cutaneous immune-related adverse events in patients with melanoma treated with checkpoint inhibitors.

Checkpoint inhibitor (CPI) therapy has vastly improved long-term outcomes in metastatic malignant melanoma (MMM). Therapy takes the form of monoclonal antibody infusions that target immune cell checkpoint proteins, such as cytotoxic T-lymphocyte-associated protein 4 (CTLA4) and programmed death 1/programmed death ligand 1 (PD1/PDL1). Cutaneous immune-related adverse effects (IrAEs) are frequent in patients with MMM treated with CPIs. Our aim was to review the clinical presentations of cutaneous IrAEs associated with CPI therapy in adult patients with MMM. We carried out a literature review of clinical trials, case series and case reports of patients with melanoma and those with other cancers treated with anti-CTLA4, anti-PD1/PDL1, or a combination of these therapies. Diverse clinical presentations of cutaneous IrAEs are recognized. Anti-CTLA4 therapy has a higher associated rate of cutaneous IrAEs than anti-PD1/PDL1 therapies. Low-grade cutaneous IrAEs are common and are usually managed supportively while continuing CPI therapy. Delayed presentations arising after established use of CPIs can make therapy-associated cutaneous IrAEs difficult to distinguish from coincidental dermatological disease. Vitiligo-like depigmentation is a good prognostic indicator of outcome in patients with melanoma. Life-threatening adverse events including toxic epidermal necrolysis are rare. The identification of predictive biomarkers that highlight patients at risk of life-threatening IrAEs remains an unmet need. The involvement of dermatologists in the multidisciplinary assessment of cutaneous IrAEs is increasingly pertinent in the management and care of CPI-treated patients with melanoma.

Next generation secondary electron detector with energy analysis capability for SEM.

We report the working of a novel detector design based on a Bessel Box (BB) electron energy analyser in a scanning electron microscope (SEM). We demonstrate the application of our detector for elemental identification through Auger electron detection in an SEM environment and its potential as a complementary technique to energy dispersive X-ray (EDX) spectroscopy. We also demonstrate energy-filtered secondary electron imaging of a copper-on-silicon sample using an electron pass energy of 12 eV. LAY DESCRIPTION: Advancements in the field of the Scanning Electron Microscopy have been one of the major nanotechnology enablers. A Scanning Electron Microscope (SEM) generates a magnified image of the sample by bombarding it with an electron beam and detecting the electrons that scatter off the surface along with the electrons that are generated in the sample. Conventional detectors such as the Everhart-Thornley detector (ET) or through-the-lens (TTL) detectors, either offer little to no energy analysis (ET) or limited energy filtering capability (e.g the low-pass energy filter in TTL). This information is crucial to interpret the image of the sample under study. What is needed is a smart and compact detector that can detect electrons and furnish energy inside the SEM chamber. Here, we report a novel secondary electron (SE) detector design with energy analysis capability for use in scanning electron microscopes. The detector is based on the design of a Bessel Box (BB) energy analyser. We have designed and experimentally tested it in an SEM environment. The band-pass filter action of the detector enables the BB to be operated at a selected energy and allows a narrow window of energies to be detected for generating energy-filtered images.

Targeted endodontic microsurgery and endodontic microsurgery: a surgical simulation comparison.

AIM: To compare surgical time, bevel angle and site volumetric profiles of osteotomy and resection accomplished by targeted endodontic microsurgery (TEMS) and traditional endodontic microsurgery (EMS) in a surgical simulation model. METHODOLOGY: An 80x80-mm cone beam computed tomography (CBCT) file was imported into Mimics software where artificial periapical lesions were created encompassing twelve root apices. Maxillary and mandibular models were 3D-printed. TEMS surgical guides were designed and 3D-printed for each surgical site. Three board-certified endodontists used the original CBCT to plan and perform EMS on models of six maxillary and six mandibular teeth. Next, the same endodontists performed TEMS on duplicate 3D-printed models for the same teeth. All surgeries were timed. Postoperative CBCT images of experimental models were made and imported into Amira software for measurement of bevel angle and site volumetric profiles. Paired t-tests compared the mean differences between EMS and TEMS groups. A Bonferroni correction determined data to be significant at P < 0.004. RESULTS: TEMS significantly reduced surgical time (P < 0.00001), had bevel angles more closely approaching zero degrees (P < 0.01) and had significantly less volume of over-resection (P < 0.001) and length of root resection (P < 0.01). CONCLUSIONS: In this surgical simulation scenario, TEMS provided more efficient completion of osteotomy and resection, with a more appropriate root-end resection volume and bevel angle.

A Suite of Tutorials for the WESTPA Rare-Events Sampling Software [Article v1.0].

The weighted ensemble (WE) strategy has been demonstrated to be highly efficient in generating pathways and rate constants for rare events such as protein folding and protein binding using atomistic molecular dynamics simulations. Here we present five tutorials instructing users in the best practices for preparing, carrying out, and analyzing WE simulations for various applications using the WESTPA software. Users are expected to already have significant experience with running standard molecular dynamics simulations using the underlying dynamics engine of interest (e.g. Amber, Gromacs, OpenMM). The tutorials range from a molecular association process in explicit solvent to more complex processes such as host-guest association, peptide conformational sampling, and protein folding.

Electrically controlled water permeation through graphene oxide membranes.

Controlled transport of water molecules through membranes and capillaries is important in areas as diverse as water purification and healthcare technologies1-7. Previous attempts to control water permeation through membranes (mainly polymeric ones) have concentrated on modulating the structure of the membrane and the physicochemical properties of its surface by varying the pH, temperature or ionic strength3,8. Electrical control over water transport is an attractive alternative; however, theory and simulations9-14 have often yielded conflicting results, from freezing of water molecules to melting of ice14-16 under an applied electric field. Here we report electrically controlled water permeation through micrometre-thick graphene oxide membranes17-21. Such membranes have previously been shown to exhibit ultrafast permeation of water17,22 and molecular sieving properties18,21, with the potential for industrial-scale production. To achieve electrical control over water permeation, we create conductive filaments in the graphene oxide membranes via controllable electrical breakdown. The electric field that concentrates around these current-carrying filaments ionizes water molecules inside graphene capillaries within the graphene oxide membranes, which impedes water transport. We thus demonstrate precise control of water permeation, from ultrafast permeation to complete blocking. Our work opens up an avenue for developing smart membrane technologies for artificial biological systems, tissue engineering and filtration.

Enhancement of the spin polarization of an Fe3O4(100) surface by nitric oxide adsorption.

The geometric, electronic and magnetic properties of a nitric oxide (NO) adsorbed Fe3O4(100) surface have been investigated using density functional theory (DFT) calculations. NO molecules preferentially bond with surface Fe(B) atoms via their N atoms. The generalized gradient approximation (GGA) is not recommended to be used in such a strongly correlated system since it provides not only an overestimation of the adsorption energy and an underestimation of the Fe(B)-N bond length, but also magnetic quenching of the adsorbate and the bonded Fe(B) atoms. In contrast, a tilted geometry and magnetization of the adsorbate and the bonded Fe(B) atom are obtained after including the strong on-site Coulomb interactions through a Hubbard term (GGA+U). The spin-down 2π* states of the NO molecule are filled and broadened due to the adsorbate-substrate interaction and the molecule-molecule interaction. The surface spin polarization close to the Fermi level is expected to be greatly enhanced by the NO adsorption which has significance for interface design in spintronic devices.

Reversible switching of the spin state in a manganese phthalocyanine molecule by atomic nitrogen.

Reversible control of the spin state of an organic molecule is significant for the development of molecular spintronic devices. Here, density functional theory calculations have been performed to study the adsorption of atomic nitrogen on a single manganese phthalocyanine (MnPc) molecule, three-layered MnPc, and MnPc on an Fe(100) surface. For all three cases, the N atom strongly adsorbs on top of the Mn atom and induces a significant variation of the geometric, electronic and magnetic properties. After N adsorption, an energy gap appears and the electronic states become unpolarized. Different functionals including three hybrid functionals are used in these calculations, and all yield a switchable spin state.

Ultrathin graphene-based membrane with precise molecular sieving and ultrafast solvent permeation.

Graphene oxide (GO) membranes continue to attract intense interest due to their unique molecular sieving properties combined with fast permeation. However, their use is limited to aqueous solutions because GO membranes appear impermeable to organic solvents, a phenomenon not yet fully understood. Here, we report efficient and fast filtration of organic solutions through GO laminates containing smooth two-dimensional (2D) capillaries made from large (10-20 µm) flakes. Without modification of sieving characteristics, these membranes can be made exceptionally thin, down to ∼10 nm, which translates into fast water and organic solvent permeation. We attribute organic solvent permeation and sieving properties to randomly distributed pinholes interconnected by short graphene channels with a width of 1 nm. With increasing membrane thickness, organic solvent permeation rates decay exponentially but water continues to permeate quickly, in agreement with previous reports. The potential of ultrathin GO laminates for organic solvent nanofiltration is demonstrated by showing >99.9% rejection of small molecular weight organic dyes dissolved in methanol. Our work significantly expands possibilities for the use of GO membranes in purification and filtration technologies.

Tolerogenic dendritic cells generated with dexamethasone and vitamin D3 regulate rheumatoid arthritis CD4+ T cells partly via transforming growth factor-ß1.

Tolerogenic dendritic cells (tolDC) are a new immunotherapeutic tool for the treatment of rheumatoid arthritis (RA) and other autoimmune disorders. We have established a method to generate stable tolDC by pharmacological modulation of human monocyte-derived DC. These tolDC exert potent pro-tolerogenic actions on CD4+ T cells. Lack of interleukin (IL)-12p70 production is a key immunoregulatory attribute of tolDC but does not explain their action fully. Here we show that tolDC express transforming growth factor (TGF)-ß1 at both mRNA and protein levels, and that expression of this immunoregulatory cytokine is significantly higher in tolDC than in mature monocyte-derived DC. By inhibiting TGF-ß1 signalling we demonstrate that tolDC regulate CD4+ T cell responses in a manner that is at least partly dependent upon this cytokine. Crucially, we also show that while there is no significant difference in expression of TGF-ßRII on CD4+ T cells from RA patients and healthy controls, RA patient CD4+ T cells are measurably less responsive to TGF-ß1 than healthy control CD4+ T cells [reduced TGF-ß-induced mothers against decapentaplegic homologue (Smad)2/3 phosphorylation, forkhead box protein 3 (FoxP3) expression and suppression of (IFN)-γ secretion]. However, CD4+ T cells from RA patients can, nonetheless, be regulated efficiently by tolDC in a TGF-ß1-dependent manner. This work is important for the design and development of future studies investigating the potential use of tolDC as a novel immunotherapy for the treatment of RA.

Synovial CD4+ T-cell-derived GM-CSF supports the differentiation of an inflammatory dendritic cell population in rheumatoid arthritis.

OBJECTIVE: A population of synovial inflammatory dendritic cells (infDCs) has recently been identified in rheumatoid arthritis (RA) and is thought to be monocyte-derived. Here, we investigated the role and source of granulocyte macrophage-colony-stimulating factor (GM-CSF) in the differentiation of synovial infDC in RA. METHODS: Production of GM-CSF by peripheral blood (PB) and synovial fluid (SF) CD4+ T cells was assessed by ELISA and flow cytometry. In vitro CD4+ T-cell polarisation experiments were performed with T-cell activating CD2/CD3/CD28-coated beads in the absence or presence of pro-Th1 or pro-Th17 cytokines. CD1c+ DC and CD16+ macrophage subsets were flow-sorted and analysed morphologically and functionally (T-cell stimulatory/polarising capacity). RESULTS: RA-SF CD4+ T cells produced abundant GM-CSF upon stimulation and significantly more than RA-SF mononuclear cells depleted of CD4+ T cells. GM-CSF-producing T cells were significantly increased in RA-SF compared with non-RA inflammatory arthritis SF, active RA PB and healthy donor PB. GM-CSF-producing CD4+ T cells were expanded by Th1-promoting but not Th17-promoting conditions. Following coculture with RA-SF CD4+ T cells, but not healthy donor PB CD4+ T cells, a subpopulation of monocytes differentiated into CD1c+ infDC; a process dependent on GM-CSF. These infDC displayed potent alloproliferative capacity and enhanced GM-CSF, interleukin-17 and interferon-γ production by CD4+ T cells. InfDC with an identical phenotype to in vitro generated cells were significantly enriched in RA-SF compared with non-RA-SF/tissue/PB. CONCLUSIONS: We demonstrate a therapeutically tractable feedback loop of GM-CSF secreted by RA synovial CD4+ T cells promoting the differentiation of infDC with potent capacity to induce GM-CSF-producing CD4+ T cells.

Half-metallicity induced by boron adsorption on an Fe3O4(100) surface.

The spin polarization of magnetite Fe3O4 is significantly reduced at its surfaces, which is unfavorable for the development of spintronic devices based on this material. In order to enhance the surface spin polarization, the Fe3O4(100) surface is modified here through the adsorption of boron (B) atoms and investigated using density functional theory (DFT) calculations. We find for the bulk-terminated and cation-redistributed surfaces that a band gap is opened in the spin-up electronic states due to the formation of a strong bond between the B atom and a surface oxygen atom, i.e., B adsorption induces half-metallicity at the Fe3O4(100) surface. Besides the surface Fe and O atoms, the adsorbed B atoms have a considerable density of -100% spin-polarized electronic states at the Fermi level, which might provide a means of improving the efficiency of spin injection into spintronic devices.

Ultrahigh vacuum and low-temperature cleaning of oxide surfaces using a low-concentration ozone beam.

We present a novel method of delivering a low-concentration (<15%) ozone beam to an ultra-high vacuum environment for the purpose of cleaning and dosing experimental samples through oxidation processing. The system described is safe, low-cost, and practical and overcomes the limitations of ozone transport in the molecular flow environment of high or ultrahigh vacuum whilst circumventing the use of pure ozone gas which is potentially highly explosive. The effectiveness of this method in removing surface contamination is demonstrated through comparison of high-temperature annealing of a simple oxide (MgO) in ozone and oxygen environments as monitored using quadrupole mass spectroscopy and Auger electron spectroscopy. Additionally, we demonstrate the potential of ozone for obtaining clean complex oxide surfaces without the need for high-temperature annealing which may significantly alter surface structure.

Magnetic moment enhancement and spin polarization switch of the manganese phthalocyanine molecule on an IrMn(100) surface.

The geometric, electronic, and magnetic structures of a manganese phthalocyanine (MnPc) molecule on an antiferromagnetic IrMn(100) surface are studied by density functional theory calculations. Two kinds of orientation of the adsorbed MnPc molecule are predicted to coexist due to molecular self-assembly on the surface-a top-site geometry with the Mn-N bonds aligned along the ⟨100⟩ direction, and a hollow-site orientation in which the Mn-N bonds are parallel to the ⟨110⟩ direction. The MnPc molecule is antiferromagnetically coupled to the substrate at the top site with a slight reduction in the magnetic moment of the Mn atom of the MnPc molecule (Mnmol). In contrast, the magnetic moment of the Mnmol is enhanced to 4.28 µB at the hollow site, a value larger than that in the free MnPc molecule (3.51 µB). Molecular distortion induced by adsorption is revealed to be responsible for the enhancement of the magnetic moment. Furthermore, the spin polarization of the Mnmol atom at around the Fermi level is found to change from negative to positive through an elongation of the Mn-N bonds of the MnPc. We propose that a reversible switch of the low/high magnetic moment and negative/positive spin polarization might be realized through some mechanical engineering methods.

Significant variation of surface spin polarization through group IV atom (C, Si, Ge, Sn) adsorption on Fe3O4(100).

The adsorption of group IV atoms (C, Si, Ge, Sn) on the magnetite Fe3O4(100) surface is investigated by density functional theory calculations. All these atoms prefer to bond to the surface oxygen atom which has no tetrahedral Fe(A) neighbor. The spin-up surface states of clean Fe3O4(100) are completely removed and half-metallicity is recovered by C adsorption. The spin-up band gap of the C-adsorbed Fe3O4(100) surface is wider than that of the H-adsorbed one and closer to the value of bulk Fe3O4. For the adsorption of other group IV atoms, the adsorbate-substrate interaction decreases and the adsorbate-adsorbate interaction increases with the increase of atomic number Z. As a consequence, the spin polarization varies from -99.4% (C adsorption) to +44.2% (Sn adsorption) for the electronic states of the adsorbed atom integrated from -0.5 eV to the Fermi level. The ability to tune the surface spin polarization by the choice of adsorbate is of significance for magnetite-based spintronic devices.

Building HIA approaches into strategies for green space use: an example from Plymouth's (UK) Stepping Stones to Nature project.

The health and well-being benefits of access to green space are well documented. Research suggests positive findings regardless of social group, however barriers exist that limit access to green space, including proximity, geography and differing social conditions. Current public health policy aims to broaden the range of environmental public health interventions through effective partnership working, providing opportunities to work across agencies to promote the use of green space. Health Impact Assessment (HIA) is a combination of methods and procedures to assess the potential health and well-being impacts of policies, developments and projects. It provides a means by which negative impacts can be mitigated and positive impacts can be enhanced, and has potential application for assessing green space use. This paper describes the application of a HIA approach to a multi-agency project (Stepping Stones to Nature--SS2N) in the UK designed to improve local green spaces and facilitate green space use in areas classified as having high levels of deprivation. The findings suggest that the SS2N project had the potential to provide significant positive benefits in the areas of physical activity, mental and social well-being. Specific findings for one locality identified a range of actions that could be taken to enhance benefits, and mitigate negative factors such as anti-social behaviour. The HIA approach proved to be a valuable process through which impacts of a community development/public health project could be enhanced and negative impacts prevented at an early stage; it illustrates how a HIA approach could enhance multi-agency working to promote health and well-being in communities.