Additional Rounds Research Articles

Doubled Haploid Technology: Generation of Doubled Haploid Maize Lines Using Haploid Inducers.

Doubled haploid (DH) technology allows for the development of completely homozygous lines from heterozygous plants in only two generations. This approach has been widely adopted in maize breeding programs, as it expedites the generation of inbred lines compared to traditional methods. The DH approach is based on the use of maize genotypes that have the ability to induce haploid seeds when used as the pollen parent. The most common method for producing maize haploid plants for the generation of DH lines is in vivo maternal haploid induction. The process involves pollination with a haploid inducer maize line to generate haploid seeds. Then, haploids are screened for and identified (typically via the expression of a particular marker gene), germinated, treated with an exogenous doubling agent to induce genome duplication, and transplanted to the field. Following successful self-pollination, seeds harvested from the ear represent fully homozygous lines. The seed set at this stage, however, is often low, necessitating one or two additional rounds of self-pollination to increase the number of fully homozygous inbred lines. Here, we describe a protocol for the generation of maize DH lines using maternal haploid-inducing maize lines. We outline the steps for setting up the donor material, performing induction crosses, selecting haploids based on two different marker alleles, treating seedlings with colchicine to double the genome, transplanting the treated seedlings to the field, and self-pollinating the treated plants.

Botulinum Toxin A for the management of temporomandibular myofascial pain - a cohort study

Myofascial pain represents the largest subgroup of TMD (Temporomandibular Disorders), which accounts for a common cause of non-dental orofacial pain. The management of TMD is complex, due to the chronic nature of the condition, alongside acute episodes presenting to the clinician. A fundamental part of TMD management is consideration of the bio-psycho-social element in the aetiology of TMD. First-line treatment of myofascial TMD includes; early diagnosis, explanation and education and conservative self-care measures. More recently, Botulinum Toxin A (BTX-A) is being used increasingly as an adjunct to conservative management of muskulo-skeletal pain disorders, due to its muscle-relaxant and analgesic properties. However, the scientific evidence regarding this is conflicting and it has been suggested that there is insufficient evidence to support the efficacy of BTX-A. A mixed method analysis of a TMD-myofascial pain cohort who underwent BTX-A injections to assess the effectiveness of masseteric BTX-A was carried out. 149 patients completed one round of masseteric BTX-A treatment, of whom 61 completed an additional round of BTX-A. In total, 398 masseter muscles were injected. The average VAS pain score pre-operatively was 8/10, compared with a postoperative mean score of 3/10 at 6 weeks after treatment. The mean percentage reduction in pain was 50%. Pain scores were classified as mild (≤4) , moderate (≤7) or severe (≥8). Pain scores and quality of life scores were found to improve considerably more in the severe pain group in comparison to the mild group. Complete resolution of symptoms was reported in 21% of patients (n=31). The treatment significantly improved patients’ reported pain and quality of life scores, highlighting key beneficial effects for the myofascial pain subgroup of TMD.

Make it STING: nanotechnological approaches for activating cGAS/STING as an immunomodulatory node in osteosarcoma.

Osteosarcoma is a highly aggressive bone cancer primarily affecting children, adolescents, and young adults. The current gold standard for treatment of osteosarcoma patients consists of two to three rounds of chemotherapy, followed by extensive surgical intervention from total limb reconstruction to amputation, followed by additional rounds of chemotherapy. Although chemotherapy has advanced the treatment of osteosarcoma significantly, the overall 5-year survival rate in resistant forms of osteosarcoma is still below 20%. The interaction between cancer and the immune system has long been recognized as a critical aspect of tumour growth. Tumour cells within the tumour microenvironment (TME) suppress antitumour immunity, and immunosuppressive cells and cytokines provide the extrinsic factors of tumour drug resistance. Emerging research demonstrates an immunostimulatory role for the cGAS/STING pathway in osteosarcoma, typically considered an immune-cold or immunosuppressed cancer type. cGAS/STING signalling appears to drive an innate immune response against tumours and potentiates the efficacy of other common therapies including chemo and radiotherapy. Nanotechnological delivery systems for improved therapy delivery for osteosarcoma have also been under investigation in recent years. This review provides an overview of cGAS/STING signalling, its divergent roles in the context of cancer, and collates current research which activates cGAS/STING as an adjuvant immunomodulatory target for the treatment of osteosarcoma. It will also discuss current nanotechnological delivery approaches that have been developed to stimulate cGAS/STING. Finally, it will highlight the future directions that we believe will be central to the development of this transformative field.

Towards the automatic risk of bias assessment on randomized controlled trials: A comparison of RobotReviewer and humans.

RobotReviewer is a tool for automatically assessing the risk of bias in randomized controlled trials, but there is limited evidence of its reliability. We evaluated the agreement between RobotReviewer and humans regarding the risk of bias assessment based on 1955 randomized controlled trials. The risk of bias in these trials was assessed via two different approaches: (1) manually by human reviewers, and (2) automatically by the RobotReviewer. The manual assessment was based on two groups independently, with two additional rounds of verification. The agreement between RobotReviewer and humans was measured via the concordance rate and Cohen's kappa statistics, based on the comparison of binary classification of the risk of bias (low vs. high/unclear) as restricted by RobotReviewer. The concordance rates varied by domain, ranging from 63.07% to 83.32%. Cohen's kappa statistics showed a poor agreement between humans and RobotReviewer for allocation concealment (κ = 0.25, 95% CI: 0.21-0.30), blinding of outcome assessors (κ = 0.27, 95% CI: 0.23-0.31); While moderate for random sequence generation (κ = 0.46, 95% CI: 0.41-0.50) and blinding of participants and personnel (κ = 0.59, 95% CI: 0.55-0.64). The findings demonstrate that there were domain-specific differences in the level of agreement between RobotReviewer and humans. We suggest that it might be a useful auxiliary tool, but the specific manner of its integration as a complementary tool requires further discussion.

A new patient-reported outcome measure for the evaluation of ankle instability: description of the development process and validation protocol

BackgroundAcute ankle sprains represent one of the most common traumatic injuries to the musculoskeletal system. Many individuals with these injuries experience unresolved symptoms such as instability and recurrent sprains, leading to chronic ankle instability (CAI), which affects their ability to maintain an active lifestyle. While rehabilitation programs focusing on sensorimotor, neuromuscular, strength and balance training are primary treatments, some patients require surgery when rehabilitation fails. A critical analysis of the patient-reported outcome tools (PROs) used to assess CAI surgical outcomes raises some concerns about their measurement properties in CAI patients, which may ultimately affect the quality of evidence supporting current surgical practice. The aim of this research is to develop and validate a new PRO for the assessment of ankle instability and CAI treatment outcomes, following recent methodological guidelines, with the implicit aim of contributing to the generation of scientifically meaningful evidence for clinical practice in patients with ankle instability.MethodsFollowing the COnsensus-based Standards for the selection of Health Measurement Instruments (COSMIN), an Ankle Instability Treatment Index (AITI) will be developed and validated. The process begins with qualitative research based on face‒to‒face interviews with CAI individuals to explore the subjective experience of living with ankle instability. The data from the interviews will be coded following an inductive approach and used to develop the AITI content. The preliminary version of the scale will be refined through an additional round of face‒to‒face interviews with a new set of CAI subjects to define the AITI content coverage, relevance and clarity. Once content validity has been examined, the AITI will be subjected to quantitative analysis of different measurement properties: construct validity, reliability and responsiveness.DiscussionThe development of AITI aims to address the limitations of existing instruments for evaluating surgical outcomes in patients with CAI. By incorporating patient input and adhering to contemporary standards for validity and reliability, this tool seeks to provide a reliable and meaningful assessment of treatment effects.Trial registrationNot applicable.

Application of Machine Learning in a Rodent Malaria Model for Rapid, Accurate, and Consistent Parasite Counts.

Rodent malaria models serve as important preclinical antimalarial and vaccine testing tools. Evaluating treatment outcomes in these models often requires manually counting parasite-infected red blood cells (iRBCs), a time-consuming process, which can be inconsistent between individuals and laboratories. We have developed an easy-to-use machine learning (ML)-based software, Malaria Screener R, to expedite and standardize such studies by automating the counting of Plasmodium iRBCs in rodents. This software can process Giemsa-stained blood smear images captured by any camera-equipped microscope. It features an intuitive graphical user interface that facilitates image processing and visualization of the results. The software has been developed as a desktop application that processes images on standard Windows and MacOS computers. A previous ML model created by the authors designed to count Plasmodium falciparum-infected human RBCs did not perform well counting Plasmodium-infected mouse RBCs. We leveraged that model by loading the pretrained weights and training the algorithm with newly collected data to target Plasmodium yoelii- and Plasmodium berghei-infected mouse RBCs. This new model reliably measured both P. yoelii and P. berghei parasitemia (R2 = 0.9916). Additional rounds of training data to incorporate variances due to length of Giemsa staining and type of microscopes, etc., have produced a generalizable model, meeting WHO competency level 1 for the subcategory of parasite counting using independent microscopes. Reliable, automated analyses of blood-stage parasitemia will facilitate rapid and consistent evaluation of novel vaccines and antimalarials across laboratories in an easily accessible invivo malaria model.

A Framework for Measuring Neighborhood Walkability for Older Adults-A Delphi Consensus Study.

While mobility in older age is of crucial importance for health and well-being, it is worth noting that currently, there is no German language framework for measuring walkability for older adults that also considers the functional status of a person. Therefore, we combined the results of an expert workshop, a literature review, and a Delphi consensus survey. Through this, we identified and rated indicators relevant for walkability for older adults, additionally focusing on their functional status. The expert workshop and the review led to an extensive list of potential indicators, which we hope will be useful in future research. Those indicators were then adapted and rated in a three-stage Delphi expert survey. A fourth additional Delphi round was conducted to assess the relevance of each indicator for the different frailty levels, namely "robust," "pre-frail," and "frail." Between 20 and 28 experts participated in each round of the Delphi survey. The Delphi process resulted in a list of 72 indicators deemed relevant for walkability in older age groups, grouped into three main categories: "Built environment and transport infrastructure," "Accessibility and meeting places," and "Attractiveness and sense of security." For 35 of those indicators, it was suggested that functional status should be additionally considered. This framework represents a significant step forward in comprehensively covering indicators for subjective and objective walkability in older age, while also incorporating aspects of functioning relevant to older adults. It would be beneficial to test and apply the indicator set in a community setting.

Into the depths: Unveiling ELAIS-N1 with LOFAR’s deepest sub-arcsecond wide-field images

We present the deepest wide-field 115–166 MHz image at sub-arcsecond resolution spanning an area of 2.5° × 2.5° centred at the ELAIS-N1 deep field. To achieve this, we improved the direction-independent (DI) and direction-dependent (DD) calibrations for the International LOw Frequency ARray (LOFAR) Telescope. This enhancement enabled us to efficiently process 32 h of data from four different 8-h observations using the high-band antennas (HBAs) of all 52 stations, covering baselines up to approximately 2000 km across Europe. The DI calibration was improved by using an accurate sky model and refining the series of calibration steps on the in-field calibrator, while the DD calibration was improved by adopting a more automated approach for selecting the DD calibrators and inspecting the self-calibration on these sources. For our brightest calibrators, we also added an additional round of self-calibration for the Dutch core and remote stations in order to refine the solutions for shorter baselines. To complement our highest resolution at 0.3″, we also made intermediate resolution wide-field images at 0.6″ and 1.2″. Our resulting wide-field images achieve a central noise level of 14 μJy beam−1 at 0.3″, doubling the depth and uncovering four times more objects than the Lockman Hole deep field image at comparable resolution but with only 8 h of data. Compared to LOFAR imaging without the international stations, we note that due to the increased collecting area and the absence of confusion noise, we reached a point-source sensitivity comparable to a 500-h ELAIS-N1 6″ image with 16 times less observing time. Importantly, we have found that the computing costs for the same amount of data are almost halved (to about 139 000 CPU h per 8 h of data) compared to previous efforts, though they remain high. Our work underscores the value and feasibility of exploiting all Dutch and international LOFAR stations to make deep wide-field images at sub-arcsecond resolution.

Force and the α-C-terminal domains bias RNA polymerase recycling

After an RNA polymerase reaches a terminator, instead of dissociating from the template, it may diffuse along the DNA and recommence RNA synthesis from the previous or a different promoter. Magnetic tweezers were used to monitor such secondary transcription and determine the effects of low forces assisting or opposing translocation, protein roadblocks, and transcription factors. Remarkably, up to 50% of Escherichia coli (E. coli) RNA polymerases diffused along the DNA after termination. Force biased the direction of diffusion (sliding) and the velocity increased rapidly with force up to 0.7 pN and much more slowly thereafter. Sigma factor 70 (σ70) likely remained associated with the DNA promoting sliding and enabling re-initiation from promoters in either orientation. However, deletions of the α-C-terminal domains severely limited the ability of RNAP to turn around between successive rounds of transcription. The addition of elongation factor NusG, which competes with σ70 for binding to RNAP, limited additional rounds of transcription. Surprisingly, sliding RNA polymerases blocked by a DNA-bound lac repressor could slowly re-initiate transcription and were not affected by NusG, suggesting a σ-independent pathway. Low forces effectively biased promoter selection suggesting a prominent role for topological entanglements that affect RNA polymerase translocation.

Creation of a Systematic Interdental Brush Set Based on the Passage Hole Diameter (PHD) - An In-Vitro Study.

This study aimed to develop a systematic interdental brush set with size distribution based on the passage hole diameter (PHD), addressing existing gaps in size selection criteria for effective interdental cleaning. In the first step, an interdental brush set that ascends stepwise according to the PHD value was envisioned. The study was divided into three phases: (i) in-vitro determination of PHD values of a currently existing assortment on the market by 13 calibrated dental professionals, (ii) in-vitro assessment of forces during insertion, and (iii) creation and evaluation of new prototypes for missing or non-matching PHD sizes. Intra- and inter-rater reliability, assessed with the intraclass correlation coefficient (ICC), as well as insertion forces and PHD sizes at all stages were reported. In the existing range, three interdental brushes fitting the desired PHD sizes were initially identified. Mean insertion forces between 0.3 and 1.7 N were documented based on raters' PHD choices. Two additional rounds of measurements with prototypes adapted in diameter and shape were necessary, particularly for PHD values of 1.4, 2.3 and 2.6. High intra- and inter-rater reliability was observed throughout the study (ICC > 0.95), ensuring consistent evaluations. After three rounds of assessments, a prototype was successfully identified for each targeted PHD value in the systematised set, showcasing reliable sizing and insertion forces. Using a structured approach, a comprehensive interdental brush set was developed with reliable PHD sizing and moderate insertion forces. The verification of size reliability through measurements by dentists represents a novelty in development and underlines the importance of accurate brush size selection for optimal biofilm control. Whether a systematic set based on the PHD value offers added value for clinical practice, and at what intervals, must be demonstrated in further studies.

Building a performance measurement framework for telephone triage services in Finland: a consensus-making study based on nominal group technique

BackgroundThere has been a significant expansion in the measurement of healthcare system performance. However, there is a lack of a comprehensive performance measurement framework to assess the effects of telephone triage services on the urgent care system. The aim of our Delphi study was to construct and validate a performance measurement framework designed explicitly for telephone triage services.MethodsThis study was conducted in Finland with a group of eight experienced senior physicians from the country's 20 largest joint emergency departments, serving over 90% of the population for urgent care. The Nominal Group Technique (NGT) was utilised to achieve consensus on measuring telephone triage performance. Initially, performance indicators (PIs) were identified through Delphi method rounds from December 10th to December 27th, 2021, with eight experts participating, and from December 29th, 2021, to January 23rd, 2022, where five of these experts responded. NGT further deepened these themes and perspectives, aiding in the development of a comprehensive performance measurement framework. The final framework validation began with an initial round from February 13th to March 3rd, 2022, receiving five responses. Due to the limited number of responses, an additional validation round was conducted from October 29th to November 7th, 2023, resulting in two more responses, increasing the total number of respondents in the validation phase to seven.ResultsThe study identified a strong desire among professionals to implement a uniform framework for measuring telephone triage performance. The finalised framework evaluates telephone triage across five dimensions: service accessibility, patient experience, quality and safety, process outcome, and cost per case. Eight specific PIs were established, including call response metrics, service utility, follow-up care type and distribution, ICPC-2 classified encounter reasons, patient compliance with follow-up care, medical history review during assessment, and service cost per call.ConclusionsThis study validated a performance measurement framework for telephone triage services, utilising existing literature and the NGT method. The framework includes five key dimensions: patient experience, quality and safety, outcome of the telephone triage process, cost per case, and eight PIs. It offers a structured and comprehensive approach to measuring the overall performance of telephone triage services, enhancing our ability to evaluate these services effectively.

Bioactivity-driven fungal metabologenomics identifies antiproliferative stemphone analogs and their biosynthetic gene cluster

IntroductionFungi biosynthesize chemically diverse secondary metabolites with a wide range of biological activities. Natural product scientists have increasingly turned towards bioinformatics approaches, combining metabolomics and genomics to target secondary metabolites and their biosynthetic machinery. We recently applied an integrated metabologenomics workflow to 110 fungi and identified more than 230 high-confidence linkages between metabolites and their biosynthetic pathways.ObjectivesTo prioritize the discovery of bioactive natural products and their biosynthetic pathways from these hundreds of high-confidence linkages, we developed a bioactivity-driven metabologenomics workflow combining quantitative chemical information, antiproliferative bioactivity data, and genome sequences.MethodsThe 110 fungi from our metabologenomics study were tested against multiple cancer cell lines to identify which strains produced antiproliferative natural products. Three strains were selected for further study, fractionated using flash chromatography, and subjected to an additional round of bioactivity testing and mass spectral analysis. Data were overlaid using biochemometrics analysis to predict active constituents early in the fractionation process following which their biosynthetic pathways were identified using metabologenomics.ResultsWe isolated three new-to-nature stemphone analogs, 19-acetylstemphones G (1), B (2) and E (3), that demonstrated antiproliferative activity ranging from 3 to 5 µM against human melanoma (MDA-MB-435) and ovarian cancer (OVACR3) cells. We proposed a rational biosynthetic pathway for these compounds, highlighting the potential of using bioactivity as a filter for the analysis of integrated—Omics datasets.ConclusionsThis work demonstrates how the incorporation of biochemometrics as a third dimension into the metabologenomics workflow can identify bioactive metabolites and link them to their biosynthetic machinery.

Detection of Contralateral Malignancies on Breast MRI.

Women with unilateral breast cancer are at increased risk for having simultaneous cancer of the contralateral breast. Overall, earlier detection of contralateral breast cancer prevents the burden of additional surgery or chemotherapy rounds and is associated with higher overall survival. However, MRI screening for the contralateral breast is seldom done following an initial unilateral breast cancer diagnosis. The purpose of this study is to retrospectively evaluate patients with known, biopsy-proven malignancy who went on to obtain a breast MRI and were later found to have cancer of the contralateral breast. Methods: This was a retrospective study that reviewed the charts of women aged over 18 years who were determined to have synchronous bilateral breast cancer from January 2017 to January 2022 at the University of Florida, Gainesville, FL. The study extracted data from this institution's cancer registry database, which provided information on patients with breast cancer diagnoses. The study conducted a review of mammography (MAM) and MRI imaging reports to ascertain the presence or absence of contralateral breast cancer identified by each respective imaging modality. Surgical pathology reports from the biopsy of the contralateral breast were reviewed to obtain information on the histological type of cancer and TNM (tumor, node, metastasis) staging. Of the 17 cases in which MAM missed contralateral cancer, follow-up MRI detected contralateral malignancy in 12 cases (70.59%) and subsequently changed management, resulting in additional imaging, biopsy, and eventual diagnosis and treatment of contralateral breast cancer. Examining the number of contralateral breast cancers detected by patients who had undergone MAM followed by MRI and those who had only undergone MAM, the study found that the detection rate of contralateral breast cancer from MAM was 45.45% (15/33). The tumor stages of the missed cancers were all T1 or Tis stage with one T1mi, and there was no nodal involvement. Conclusion: In addition to its utility in staging breast cancers, MRI also has the superior ability to detect otherwise undetected contralateral breast malignancy. This retrospective study found that MRI imaging led to a considerable increase in the detection of contralateral cancer. The study found that these undetected contralateral breast cancers by MAM were often of lower staging with no nodal involvement, highlighting the opportunity for MRI to assist in early cancer detection while the patient's prognosis is still good. Its high cost should be balanced with staging and occult malignancy detection utility in future practice.

Use of federated learning to develop an artificial intelligence model predicting usable blastocyst formation from pre-ICSI oocyte images

Research questionCan Federated Learning be used to develop an artificial intelligence (AI) for evaluating oocyte competence using two-dimensional images of denuded oocytes in metaphase II prior to intracytoplasmic sperm injection (ICSI)? Design10,677 oocyte images with associated metadata were prospectively collected by 8 in vitro fertilization (IVF) clinics across 6 countries. AI training used Federated Learning, where data were retained on regional servers to comply with data privacy laws. The final AI required a single image as input to evaluate oocyte competence, which was defined by the formation of a usable blastocyst (≥expansion grade 3 by Day 5/6 post-ICSI). ResultsThe oocytes AI demonstrated area under the curve (AUC) up to 0.65 on two blind test datasets. A high sensitivity for predicting competent oocytes (83-88%) was offset by a lower specificity (26-36%). Exclusion of confounding biological variables (male factor infertility and maternal age ≥35 years) improved AUC up to 14%, primarily due to increased specificity. AI scores correlated with size of the zona pellucida and perivitelline space, and ooplasm appearance. AI scores also correlated with blastocyst expansion grade and morphological quality. The sum of AI scores from oocytes in group culture images predicted formation of ≥2 usable blastocysts (AUC=0.77). ConclusionAn AI for evaluating oocyte competence was developed using Federated Learning, representing an essential step in protecting patient data. The AI was significantly predictive of oocyte competence as defined by usable blastocyst formation, which is a critical factor for IVF success. Potential clinical utility ranges from selective oocyte fertilization, to guiding treatment decisions regarding additional rounds of oocyte retrieval.

Activin Signaling Pathway Specialization During Embryonic and Skeletal Muscle Development in Rainbow Trout (Oncorhynchus mykiss).

Activin signaling is essential for proper embryonic, skeletal muscle, and reproductive development. Duplication of the pathway in teleost fish has enabled diversification of gene function across the pathway but how gene duplication influences the function of activin signaling in non-mammalian species is poorly understood. Full characterization of activin receptor signaling pathway expression was performed across embryonic development and during early skeletal muscle growth in rainbow trout (RBT, Oncorhynchus mykiss). Rainbow trout are a model salmonid species that have undergone two additional rounds of whole genome duplication. A small number of genes were expressed early in development and most genes increased expression throughout development. There was limited expression of activin Ab in RBT embryos despite these genes exhibiting significantly elevated expression in post-hatch skeletal muscle. CRISPR editing of the activin Aa1 ohnolog and subsequent production of meiotic gynogenetic offspring revealed that biallelic disruption of activin Aa1 did not result in developmental defects, as occurs with knockout of activin A in mammals. The majority of gynogenetic offspring exhibited homozygous activin Aa1 genotypes (wild type, in-frame, or frameshift) derived from the mosaic founder female. The research identifies mechanisms of specialization among the duplicated activin ohnologs across embryonic development and during periods of high muscle growth in larval and juvenile fish. The knowledge gained provides insights into potential viable gene-targeting approaches for engineering the activin receptor signaling pathway and establishes the feasibility of employing meiotic gynogenesis as a tool for producing homozygous F1 genome-edited fish for species with long-generation times, such as salmonids.

Zebrafish: unraveling genetic complexity through duplicated genes.

The zebrafish is an invaluable model organism for genetic, developmental, and disease research. Although its high conservation with humans is often cited as justification for its use, the zebrafish harbors oft-ignored genetic characteristics that may provide unique insights into gene structure and function. Zebrafish, along with other teleost fish, underwent an additional round of whole genome duplication after their split from tetrapods-resulting in an abundance of duplicated genes when compared to other vertebrates. These duplicated genes have evolved in distinct ways over the ensuing 350 million years. Thus, each gene within a duplicated gene pair has nuanced differences that create a unique identity. By investigating both members of the gene pair together, we can elucidate the mechanisms that underly protein structure and function and drive the complex interplay within biological systems, such as signal transduction cascades, genetic regulatory networks, and evolution of tissue and organ function. It is crucial to leverage such studies to explore these molecular dynamics, which could have far-reaching implications for both basic science and therapeutic development. Here, we will review the role of gene duplications and the existing models for gene divergence and retention following these events. We will also highlight examples within each of these models where studies comparing duplicated genes in the zebrafish have yielded key insights into protein structure, function, and regulation.

Using Simulation-Based Mastery Learning to Achieve Excellent Learning Outcomes in Physical Therapist Education.

The 2 aims of this observational study are (a) to describe the implementation and feasibility of a bed mobility skills simulation-based mastery learning (SBML) curricular module for physical therapist students and (b) to measure learning outcomes and student perceptions of this module. Simulation-based mastery learning is an outcome-based educational approach that has been successful in other health professions but has not been explored in physical therapy education. Eighty-seven students in a single cohort of a Doctor of Physical Therapy program. The SBML module in this pretest-posttest study included a pretest, instruction, initial posttest, and additional rounds of instruction and assessment as needed for all learners to achieve the minimum passing standard (MPS) set using the Mastery Angoff and Patient Safety methods. Outcome measures were bed mobility assessment pass rates and scores, additional student and faculty time compared with a traditional approach, and student perceptions of their self-confidence and the module. All students achieved the MPS after 3 rounds of training and assessment beyond the initial posttest. Mean Total Scores improved from 67.6% (12.9%) at pretest to 91.4% (4.8%) at mastery posttest (P < .001, Cohen's d = 1.8, 95% CI [1.4-2.1]); mean Safety Scores improved from 75.2% (16.0%) at pretest to 100.0% (0.0%) at mastery posttest (P < .001, Cohen's d = 1.5, 95% CI [1.2-1.9]). Students who did not achieve the MPS at the initial posttest (n = 30) required a mean of 1.2 hours for additional instruction and assessment. Survey results revealed an increase in student confidence (P < .001) and positive student perceptions of the module. Implementation of this SBML module was feasible and resulted in uniformly high levels of bed mobility skill acquisition. Based on rigorous learning outcomes, feasible requirements for implementation, and increased student confidence, SBML offers a promising approach for wider implementation in physical therapy education.

A Comprehensive Investigation of Methanol Electrooxidation on Copper Anodes: Spectroelectrochemical Insights and Energy Conversion in Microfluidic Fuel Cells.

Here, we comprehensively investigated methanol electrooxidation on Cu-based catalysts, allowing us to build the first microfluidic fuel cell (μFC) equipped with a Cu anode and a metal-free cathode that converts energy from methanol. We applied a simple, fast, small-scale, and surfactant-free strategy for synthesizing Cu-based nanoparticles at room temperature in steady state (ST), under mechanical stirring (MS), or under ultrasonication (US). The morphology evaluation of the Cu-based samples reveals that they have the same nanoparticle (NP) needle-like form. The elemental mapping composition spectra revealed that pure Cu or Cu oxides were obtained for all synthesized materials. In addition to having more Cu2O on the surface, sample US had more Cu(OH)2 than the others, according to X-ray diffractograms and X-ray photoelectron spectroscopy. The sample US is less carbon-contaminated because of the local heating of the sonic bath, which also enhances the cleanliness of the Cu surface. The activity of the Cu NPs was investigated for methanol electrooxidation in an alkaline medium through electrochemical and spectroelectrochemical measurements. The potentiodynamic and potentiostatic experiments showed higher current densities for the NPs synthesized in the US. In situ FTIR experiments revealed that the three synthesized NP materials eletcrooxidize methanol completely to carbonate through formate. Most importantly, all pathways were led without detectable CO, a poisoning molecule not found at high overpotentials. The reaction path using the US electrode experienced an additional round of formate formation and conversion into carbonate (or CO2 in the thin layer) after 1.0 V (vs. Ag/Ag/Cl), suggesting improved catalysis. The high activity of NPs synthesized in the US is attributed to effective dissociative adsorption of the fuel due to the site's availability and the presence of hydroxyl groups that may fasten the oxidation of adsorbates from the surface. After understanding the surface reaction, we built a mixed-media μFC fed by methanol in alkaline medium and sodium persulfate in acidic medium. The μFC was equipped with Cu NPs synthesized in ultrasonic-bath-modified carbon paper as the anode and metal-free carbon paper as the cathode. Since the onset potential for methanol electrooxidation was 0.45 V and the reduction reaction revealed 0.90 V, the theoretical OCV is 0.45 V, which provides a spontaneous coupled redox reaction to produce power. The μFC displayed 0.56 mA cm-2 of maximum current density and 26 μW cm-2 of peak power density at 100 μL min-1. This membraneless system optimizes each half-cell individually, making it possible to build fuel cells with noble metal-free anodes and metal-free cathodes.

From Protein to a Bioactive Peptide: Potent IL‐6 Peptide Antagonist Generated by a Novel Method

ABSTRACTHere, we describe a new technology, designed to accelerate peptide discovery by quick identification and optimization of the residues critical for protein–protein interactions or ligand binding. We called it PepFusion. It is based on ligation of short DNA sequences generated from known ligand‐binding regions. We tested it by selecting peptide antagonists of interleukin‐6 (IL‐6), a key mediator of inflammatory diseases such as rheumatoid arthritis (RA), Crohn's disease, and Castleman disease (CD). The PepFusion library demonstrated superiority over a random library by yielding a peptide with low micromolar affinity for IL‐6, whereas the random library failed. The affinity of the peptide from the PepFusion library was further enhanced by additional rounds of mutagenesis leading to peptide variants with low nanomolar IL‐6 affinity. In addition to generating high‐affinity peptides, our method opens the way to solve the problem of the false positive sequences, which are common with all display technologies.

ExSeq: Expansion Sequencing for Single-Cell Spatial Transcriptomics

The innovation of spatial transcriptomics allows for the study of gene expression while preserving spatial information on the detected transcripts. Expansion Sequencing (ExSeq) is an advanced spatial transcriptomic method combining in situ sequencing with expansion microscopy, thus enablingthe sequencing of RNAs in a physically enlarged tissue. The experiment involves the steps of tissue expansion, RNA library preparation and multiplexed sequencing by rounds of fluorescent dye addition, imaging and fluorescent dye removal. ExSeq significantly enhances resolution capacity of RNA transcripts up to the nanoscale level, thus identifying subcellular location of RNAs. The method is applicable to exploring cellular RNA in an untargeted approach, as well as to specifically detecting certain RNA types in a targeted approach, thus making the method suitable for a variety of research questions. However, limitations to the method include referring to the readout length of transcripts obtainable during sequencing and the availability of the method. ExSeq has the potential to be used in multiple tissues to allow for new insights into cellular identity, heterogeneity, interactions and implications in diseases and could possibly be adapted to inquire other cellular molecules, such as DNA or proteins on a nanoscale level.