Improving light microscopy training routines with evidence-based education.

The low reproducibility of scientific data published in articles has recently become a cause of concern in many scientific fields. Data involving light microscopy is no exception. The low awareness of researchers of the technologies they use in their research has been identified as one of the main causes of the problem. Potential solutions have hinted at the need to improve technological and methodological education within research. Despite the pivotal role of microscopy core facilities in the education of researchers being well documented, facility staff (FS) often learn their trade on the job, without receiving themselves any structured education about the technology they teach others to use. Additionally, despite endorsing an important role at the highest level of education, most FS never receive any training in pedagogy, the field of research on teaching and learning methods. In this article, we argue that the low level of awareness that researchers have of microscopy stems from a knowledge gap formed between them and microscopy FS during training routines. On the one hand, FS consider that their teaching task is to explain what is needed to produce reliable data. On the other, despite understanding what is being taught, researchers fail to learn the most challenging aspects of microscopy, those involving their judgement and reasoning. We suggest that the misunderstanding between FS and researchers is due to FS not being educated in pedagogy and thus often confusing understanding and learning. To bridge this knowledge gap and improve the quality of the microscopy education available to researchers, we propose a paradigm shift where training staff at technological core facilities be acknowledged as full-fledged teachers and offered structured education not only in the technology they teach but also in pedagogy. We then suggest that training routines at facilities be upgraded to follow the principles of the Constructive Alignment pedagogical method. We give an example of how this can be applied to existing microscopy training routines. We also describe a model to define where the responsibility of FS in training researchers begins and ends. This involves a major structural change where university staff involved in teaching research technologies themselves receive appropriate education. For this to be achieved, we advocate that funding agencies, universities, microscopy and core facility organisations mobilise resources of time and funding. Such changes may involve funding the creation and development of 'Train-the-trainer' type of courses and giving incentives for FS to upgrade their technological and pedagogical knowledge, for example by including them in career paths. We believe that this paradigm shift is necessary to improve the level of microscopy education and ultimately the reproducibility of published data.

Antioxidants stimulate BACH1-dependent tumor angiogenesis.

Lung cancer progression relies on angiogenesis, which is a response to hypoxia typically coordinated by hypoxia-inducible transcription factors (HIFs), but growing evidence indicates that transcriptional programs beyond HIFs control tumor angiogenesis. Here, we show that the redox-sensitive transcription factor BTB and CNC homology 1 (BACH1) controls the transcription of a broad range of angiogenesis genes. BACH1 is stabilized by lowering ROS levels; consequently, angiogenesis gene expression in lung cancer cells, tumor organoids, and xenograft tumors increased substantially following administration of vitamins C and E and N-acetylcysteine in a BACH1-dependent fashion under normoxia. Moreover, angiogenesis gene expression increased in endogenous BACH1-overexpressing cells and decreased in BACH1-knockout cells in the absence of antioxidants. BACH1 levels also increased upon hypoxia and following administration of prolyl hydroxylase inhibitors in both HIF1A-knockout and WT cells. BACH1 was found to be a transcriptional target of HIF1α, but BACH1's ability to stimulate angiogenesis gene expression was HIF1α independent. Antioxidants increased tumor vascularity in vivo in a BACH1-dependent fashion, and overexpressing BACH1 rendered tumors sensitive to antiangiogenesis therapy. BACH1 expression in tumor sections from patients with lung cancer correlated with angiogenesis gene and protein expression. We conclude that BACH1 is an oxygen- and redox-sensitive angiogenesis transcription factor.

Multisite assessment of reproducibility in high-content cell migration imaging data.

High-content image-based cell phenotyping provides fundamental insights into a broad variety of life science disciplines. Striving for accurate conclusions and meaningful impact demands high reproducibility standards, with particular relevance for high-quality open-access data sharing and meta-analysis. However, the sources and degree of biological and technical variability, and thus the reproducibility and usefulness of meta-analysis of results from live-cell microscopy, have not been systematically investigated. Here, using high-content data describing features of cell migration and morphology, we determine the sources of variability across different scales, including between laboratories, persons, experiments, technical repeats, cells, and time points. Significant technical variability occurred between laboratories and, to lesser extent, between persons, providing low value to direct meta-analysis on the data from different laboratories. However, batch effect removal markedly improved the possibility to combine image-based datasets of perturbation experiments. Thus, reproducible quantitative high-content cell image analysis of perturbation effects and meta-analysis depend on standardized procedures combined with batch correction.

Zn3V4(PO4)6: A New Rocking-Chair-Type Cathode Material with High Specific Capacity Derived from Zn2+/H+ Cointercalation for Aqueous Zn-Ion Batteries.

Phosphate cathode materials with a stable and open framework structure are expected to be one of the favorable cathode materials for aqueous zinc-ion batteries (AZIBs). However, the slow migration rate of Zn2+ and complex mechanism in aqueous electrolyte are serious problems that limit their application at the present. Here, a new rocking-chair-type cathode material Zn3V4(PO4)6@C (ZVP@C) for AZIBs is synthesized for the first time and evaluated using a composite carbon coating to improve the electronic conductivity. Benefiting from the two-electron reaction of vanadium and the cointercalation of Zn2+/H+, ZVP@C/30%BP delivers a specific capacity as high as 120 mAh·g-1 at 0.04 A·g-1. A good capacity retention of 80% after 400 cycles at 1 A·g-1 is also obtained, which is attributed to the stable crystal structure and the cointercalation reaction of Zn2+/H+. The reaction mechanism is investigated by in situ X-ray diffraction (XRD), ex situ XRD, ex situ X-ray photoelectron spectroscopy (XPS), and energy dispersive spectroscopy (EDS). This work not only provides a new phosphate cathode material for AZIBs but also gives a new strategy for improving the specific capacity of phosphate cathode material.

Local temporal Rac1-GTP nadirs and peaks restrict cell protrusions and retractions.

Cells probe their microenvironment using membrane protrusion-retraction cycles. Spatiotemporal coordination of Rac1 and RhoA GTP-binding activities initiates and reinforces protrusions and retractions, but the control of their finite lifetime remains unclear. We examined the relations of Rac1 and RhoA GTP-binding levels to key protrusion and retraction events, as well as to cell-ECM traction forces at physiologically relevant ECM stiffness. High RhoA-GTP preceded retractions and Rac1-GTP elevation before protrusions. Notable temporal Rac1-GTP nadirs and peaks occurred at the maximal edge velocity of local membrane protrusions and retractions, respectively, followed by declined edge velocity. Moreover, altered local Rac1-GTP consistently preceded similarly altered traction force. Local optogenetic Rac1-GTP perturbations defined a function of Rac1 in restricting protrusions and retractions and in promoting local traction force. Together, we show that Rac1 plays a fundamental role in restricting the size and durability of protrusions and retractions, plausibly in part through controlling traction forces.

A small-molecule ICMT inhibitor delays senescence of Hutchinson-Gilford progeria syndrome cells.

A farnesylated and methylated form of prelamin A called progerin causes Hutchinson-Gilford progeria syndrome (HGPS). Inhibiting progerin methylation by inactivating the isoprenylcysteine carboxylmethyltransferase (ICMT) gene stimulates proliferation of HGPS cells and improves survival of Zmpste24-deficient mice. However, we don't know whether Icmt inactivation improves phenotypes in an authentic HGPS mouse model. Moreover, it is unknown whether pharmacologic targeting of ICMT would be tolerated by cells and produce similar cellular effects as genetic inactivation. Here, we show that knockout of Icmt improves survival of HGPS mice and restores vascular smooth muscle cell numbers in the aorta. We also synthesized a potent ICMT inhibitor called C75 and found that it delays senescence and stimulates proliferation of late-passage HGPS cells and Zmpste24-deficient mouse fibroblasts. Importantly, C75 did not influence proliferation of wild-type human cells or Zmpste24-deficient mouse cells lacking Icmt, indicating drug specificity. These results raise hopes that ICMT inhibitors could be useful for treating children with HGPS.

Titanium Hydride Nanoplates Enable 5 wt% of Reversible Hydrogen Storage by Sodium Alanate below 80°C.

Sodium alanate (NaAlH4) with 5.6 wt% of hydrogen capacity suffers seriously from the sluggish kinetics for reversible hydrogen storage. Ti-based dopants such as TiCl4, TiCl3, TiF3, and TiO2 are prominent in enhancing the dehydrogenation kinetics and hence reducing the operation temperature. The tradeoff, however, is a considerable decrease of the reversible hydrogen capacity, which largely lowers the practical value of NaAlH4. Here, we successfully synthesized a new Ti-dopant, i.e., TiH2 as nanoplates with ~50 nm in lateral size and ~15 nm in thickness by an ultrasound-driven metathesis reaction between TiCl4 and LiH in THF with graphene as supports (denoted as NP-TiH2@G). Doping of 7 wt% NP-TiH2@G enables a full dehydrogenation of NaAlH4 at 80°C and rehydrogenation at 30°C under 100 atm H2 with a reversible hydrogen capacity of 5 wt%, superior to all literature results reported so far. This indicates that nanostructured TiH2 is much more effective than Ti-dopants in improving the hydrogen storage performance of NaAlH4. Our finding not only pushes the practical application of NaAlH4 forward greatly but also opens up new opportunities to tailor the kinetics with the minimal capacity loss.

Blood pressure and cognitive decline over the course of 2 years in elderly people: a community-based prospective cohort study.

BACKGROUND: Numerous studies have shown a significant association between blood pressure (BP) and cognition, but little is known about the effect of BP on the rate of cognitive decline. AIMS: To investigate the relationship between blood pressure and the subsequent rate of cognitive decline in elderly people. METHODS: Based on a prospective cohort that has been followed since 2014, we collected baseline blood pressures and other covariates in 7874 Chinese individuals aged 60 years or older, and followed their cognitive change using the Mini-Mental State Examination (MMSE) until Dec 31, 2016. Linear mixed-effects models were used to measure changes in MMSE scores over time in relation to blood pressure values, and in addition to the covariates, we included random effects for intercepts and slopes. RESULTS: In the non-hypertension group, we observed that faster cognitive decline was associated with higher systolic blood pressure, lower diastolic blood pressure, lower mean arterial pressure, and higher pulse pressure. In the hypertension group, lower diastolic blood pressure, lower mean arterial pressure, and higher pulse pressure were associated with faster cognitive decline, but not systolic blood pressure. CONCLUSION: Higher systolic blood pressure, lower diastolic blood pressure, lower mean arterial pressure, and higher pulse pressure accelerate the subsequent rate of cognitive decline in elderly people. The results of this study may help improve blood-pressure control strategies to prevent cognitive decline.

A Unique Double-Layered Carbon Nanobowl-Confined Lithium Borohydride for Highly Reversible Hydrogen Storage.

Poor reversibility and high desorption temperature restricts the practical use of lithium borohydride (LiBH4 ) as an advanced hydrogen store. Herein, a LiBH4 composite confined in unique double-layered carbon nanobowls prepared by a facile melt infiltration process is demonstrated, thanks to powerful capillary effect under 100 bar of H2 pressure. The gradual formation of double-layered carbon nanobowls is witnessed by transmission electron microscopy (TEM) observation. Benefiting from the nanoconfinement effect and catalytic function of carbon, this composite releases hydrogen from 225 °C and peaks at 353 °C, with a hydrogen release amount up to 10.9 wt%. The peak temperature of dehydriding is lowered by 112 °C compared with bulk LiBH4 . More importantly, the composite readily desorbs and absorbs ≈8.5 wt% of H2 at 300 °C and 100 bar H2 , showing a significant reversibility of hydrogen storage. Such a high reversible capacity has not ever been observed under the identical conditions. The usable volumetric energy density reaches as high as 82.4 g L-1 with considerable dehydriding kinetics. The findings provide insights in the design and development of nanosized complex hydrides for on-board applications.

Amorphous-Carbon-Supported Ultrasmall TiB2 Nanoparticles With High Catalytic Activity for Reversible Hydrogen Storage in NaAlH4.

In this paper, we report amorphous-carbon-supported TiB2 nanoparticles having sizes of 2-4 nm (nano-TiB2@C) as highly active catalysts for hydrogen storage in NaAlH4. Nano-TiB2@C was synthesized by a simple calcination at 550°C with Cp2TiCl2 and MgB2 (molar ratio of 1:1) as precursors. The addition of 7 wt% nano-TiB2@C reduced the onset dehydrogenation temperature of NaAlH4 by 100 to 75°C. A practically available hydrogen capacity of 5.04 wt% could be desorbed at 140°C within 60 min, and completely hydrogenated at 100°C within 25 min under a hydrogen pressure of 100 bar. Notably, the hydrogen capacity was almost unchanged after 20 cycles, which shows the stable cyclability, considerably higher than those of structures catalyzed by Ti halides or TiO2. The stable catalytic function was closely related to the in-situ-formed Ti-Al alloy, which considerably facilitated the dissociation and recombination of H-H and Al-H bondings.

Higher Than 90% Initial Coulombic Efficiency with Staghorn-Coral-Like 3D Porous LiFeO2- x as Anode Materials for Li-Ion Batteries.

Transition metal oxides represent a promising class of anode materials for high-capacity lithium-ion batteries. However, low initial coulombic efficiency (ICE, <80%) still remains a crucial challenge for practical applications. Herein, a unique 3D Fe(II)-rich porous LiFeO2- x comprising of staghorn-coral-like skeleton measuring ≈100 nm in diameter is demonstrated, which is readily prepared by reacting Fe2 O3 with LiH at 550 °C. When used as an anode material, the Fe(II)-rich LiFeO2- x delivers the presently known highest ICE value of 90.2% with 1170 mAh g-1 discharge capacity. The high ICE value can be ascribed to a fast conversion reaction of LiFeO2- x upon lithiation/delithiation facilitated by the presence of Fe(II), which generates oxygen vacancies and makes electron transportation much easier, based on the experimental results and density functional theory (DFT) calculations.

Nanoscaled Lithium Powders with Protection of Ionic Liquid for Highly Stable Rechargeable Lithium Metal Batteries.

To suppress the dendrite formation and alleviate volume expansion upon striping/platting is a key challenge for developing practical lithium metal anodes. Lithium metal in powder form possesses great potential to address this issue due to large specific surface area. However, the fabrication of powdery metallic lithium is largely restricted because of its unique softness, stickiness, and high reactivity. Here, a safe and readily accessible cryomilling process toward lithium powders is reported. Nanoscaled lithium powders (<500 nm) are successfully prepared from lithium foils with the assistance of a high-melting-point ionic liquid under cryogenic temperature. The prepared lithium powder anode exhibits superior electrochemical properties in symmetric cells, including extraordinarily low yet stable overpotential (≈50 mV), ultrahigh area capacity (30 mAh cm-2), and good long-term cyclability (1200 h) even cycling at high current density (10 mA cm-2). The powdery form of lithium also functions as a favorable prelithiation reagent for lithium-free anodes (e.g., Si, SiO, and SnO2). The findings open up a new avenue for the real-world application of lithium metal anodes for next-generation lithium batteries.

Identification of the PAK4 interactome reveals PAK4 phosphorylation of N-WASP and promotion of Arp2/3-dependent actin polymerization.

p21-activated kinase 4 (PAK4) regulates cell proliferation, apoptosis, cell motility and F-actin remodeling, but the PAK4 interactome has not been systematically analyzed. Here, we comprehensively characterized the human PAK4 interactome by iTRAQ quantitative mass spectrometry of PAK4-immunoprecipitations. Consistent with its multiple reported functions, the PAK4 interactome was enriched in diverse protein networks, including the 14-3-3, proteasome, replication fork, CCT and Arp2/3 complexes. Because PAK4 co-immunoprecipitated most subunits of the Arp2/3 complex, we hypothesized that PAK4 may play a role in Arp2/3 dependent actin regulation. Indeed, we found that PAK4 interacts with and phosphorylates the nucleation promoting factor N-WASP at Ser484/Ser485 and promotes Arp2/3-dependent actin polymerization in vitro. Also, PAK4 ablation in vivo reduced N-WASP Ser484/Ser485 phosphorylation and altered the cellular balance between G- and F-actin as well as the actin organization. By presenting the PAK4 interactome, we here provide a powerful resource for further investigations and as proof of principle, we also indicate a novel mechanism by which PAK4 regulates actin cytoskeleton remodeling.

Study on the anisotropic response of condensed-phase RDX under repeated stress wave loading via ReaxFF molecular dynamics simulation.

Anisotropic mechanical response and chemical reaction process of cyclotrimethylene trinitramine (RDX) along crystal orientations were studied with molecular dynamics simulations using ReaxFF potential under repeated stress wave loading. In the simulations, shocks were propagated along the [010], [001], [210], [100], [111], and [102] orientations of crystal RDX at initial particle velocity Up in the range of 1∼4 km/s. For shocks at Up ≤ 2 km/s, local stacking fault and molecular conformational change can only cause marginal temperature and pressure increase without molecular decomposition. As shocks increase to Up ≥ 2.5 km/s, rupture of N-NO2 bond accompanied by partial HONO elimination dominates the main chemical reactions at the initial stage. The ordering of the follow-up consumption of NO2 and ring-breaking rate is directly consistent with that of increasing rate in temperature and pressure. The (210) and (100) planes are more sensitive to shocks in temperature and pressure profiles than the (111) plane, which agrees well with experimental observations and theoretical results in the literature. Therefore, the repeated dynamic loading model in conjunction with MD simulation using ReaxFF potential for crystal RDX indicates that these methods can be applied to study the mechanical response and chemical reaction process of polymer bonded explosives that are commonly subjected to compressive and tensile stress waves observed in practice.

[Real-time detection of mast cell degranulation in anaphylactoid reaction].

OBJECTIVE: To establish a new, real time, dynamic and direct optical detection method for mast cell degranulation caused by anaphylactoid reaction. METHOD: A CD63-GFP plasmid was constructed and introduced steadily into rat basophilic leukemia (RBL-2H3) cells. The movements of CD63-GFP, which was located on both the granule membranes and the plasma membranes of RBL cells stimulated by Compound 48/80, were studied by confocal laser scanning microscope (CLSM) and total internal reflection fluorescence microscope (TIRFM) both inside and on the surface of living RBL-2H3 cells. RESULT: Before antigen stimulation, most granules with CD63-GFP hardly moved in RBL cells. However, after antigen stimulation, the granules moved dramatically. They reached the plasma membranes in a few minutes and fused with them instantaneously. The velocity of the granule movement toward the plasma membranes on antigen stimulation was calculated to be 0.05 micron x s(-1). CONCLUSION: Analysis of the movement of each granule provided a new insight into the elementary process of degranulation. The method is rapid, sensitive and reliable, which could be used as a new detection method for anaphylactoid reaction in vitro.

[Considerations about study on mechanisms of acupuncture underlying efficacy-enhancement and toxicity-attenuation of digitalis drugs for heart failure].

Digitalis glycosides, a group of cardiotonic agents for heart failure, have been used for a long time, but may often trigger arrhythmias as a result of digitalis intoxication. It is of great significance in finding a method to reduce their toxicity and improve clinical curative effects simultaneously in the application of digitalis glycosides. It has been well documented that acupuncture has good therapeutic effects in improving cardiac function and inhibiting arrhythmia induced by myocardial ischemia. Acupuncture combined with chemotherapeutics can reduce the adverse effects of chemotherapeutics. Accordingly, acupuncture combined with digitalis is likely to reduce the side effects of digitalis by regulating intracellular Ca2+, improving the function of Ca(2+)-ATPase in the sarcoplasmic reticulum, increasing calcium sensitivity of cardiac troponin, etc. These considerations may provide a novel clue for treatment of heart failure by joint administration of acupuncture and drugs.

Anti-biofilm activity of TanReQing, a Traditional Chinese Medicine used for the treatment of acute pneumonia.

AIM OF THE STUDY: TanReQing (TRQ) is a Traditional Chinese Medicine used to treat biofilm related upper respiratory infections. However, its anti-biofilm mechanism remains unknown. The aim of this study is to investigate the anti-biofilm activity of TRQ and to compare it with penicillin in vitro. MATERIALS AND METHODS: The effect of TRQ and penicillin on free state, biofilm formation and mature biofilms of Staphylococcus aureus was studied using the crystal violet and the XTT reduction assays. Confocal Laser Scanning Microscopy was used to generate the 3D-transmission-fluorescence images of drug treated Staphylococcus aureus biofilms. RESULTS: The in vitro data showed that TRQ is less effective than penicillin in eradicating the planktonic bacteria. However, the anti-biofilm activity of TRQ is different from that of penicillin. TRQ not only does inhibit the formation of the Staphylococcus aureus biofilm, but also kills the viable cells embed in Staphylococcus aureus biofilm matrix. CONCLUSIONS: This study shows for the first time that TRQ possesses an antibiotic activity against biofilm bacteria. This activity is different from that of penicillin. The evaluation system applied in this study can be utilized for identifying new anti-biofilm products from Traditional Chinese Medicine.

[In vitro study on gastrointestinal absorption of FITC labeled pilose antler protein extraction].

An in vitro detection method of the gastrointestinal absorption of Pilose Antler protein was established for mixed protein activity. Five bands of protein with molecular weight of 17.8-160 kD derived from the Pilose Antler were extracted and sufficiently labeled with FITC (FITC-PE). The stability and variation of FITC-PE in gastrointestinal circumstances were detected by native polyacrylamide gel electrophoresis and confocal laser scanning microscope. Results showed that the main component of FITC-PE kept invariant after being reacted with artificial gastric fluid and artificial intestinal fluid. The fluorescence signal was detected 20 min after administration in the valgus intestinal purse experiment, and three kinds of protein, with molecular weight of 45, 25, and 17.8 kD, were detected in the mixture of absorbent protein. The research laid the foundation for the further in vivo study of Pilose Antler protein. Meanwhile, it would be an in vitro screening method for the absorption, distribution and metabolism of mixed protein from traditional Chinese medicine.

Altered thermodynamic and kinetic properties of MgH(2) infiltrated in microporous scaffold.

MgH(2) nanoparticles with a size of <3 nm were formed by direct hydrogenation of Bu(2)Mg inside the pores of a carbon scaffold. The activation energy for the dehydrogenation was lowered by 52 kJ mol(-1) compared to the bulk material, and a significantly reduced reaction enthalpy of 63.8 ± 0.5 kJ mol(-1) and entropy (117.2 ± 0.8 J mol(-1)) was found for the nanoconfined system.

Functions of LiBH4 in the hydrogen sorption reactions of the 2LiH-Mg(NH2)2 system.

LiBH(4) is an effective additive in enhancing hydrogen desorption and absorption properties for the 2LiH-Mg(NH(2))(2) hydrogen storage system. The improving factors brought by LiBH(4) were studied experimentally. Although a minimum in activation energy was observed in the middle stage of hydrogen desorption for the samples with and without LiBH(4) doping, a reduction in the activation energy was manifest in the overall conversion range due to the LiBH(4) introduction. In contrast to the pure 2LiH-Mg(NH(2))(2), the system component Mg(NH(2))(2) crystallized in the presence of LiBH(4) at about 150 °C in situ from the post-milled amorphous mixture prior to hydrogen desorption. We believe the formation of the N-atom matrix by the crystalline Mg(NH(2))(2) is relevant to the subsequent dehydrogenation as it shares a similar sublattice structure with the desorption product. The formation and disappearance of Li(4)(BH(4))(NH(2))(3) in the dehydrogenation step witnessed the capturing and releasing of LiNH(2) that is generated from the dehydrogenation of 2LiH-Mg(NH(2))(2), which alters not only the kinetics but also thermodynamics in favor of an improved hydrogen sorption performance. The reaction mechanism could be further clarified by the refinement of powder neutron diffraction data of the intermediate compound Li(4)(BD(4))(NH(2))(3).