Endothelial Cyp26b1 restrains murine heart valve growth during development.

Endothelial cells (ECs) are critical to proper heart valve development, directly contributing to the mesenchyme of the cardiac cushions, which progressively transform into mature valves. To date, investigators have lacked sufficient markers of valve ECs to evaluate their contributions during valve morphogenesis fully. As a result, it has been unclear whether the well-characterized regional differentiation of valves correlates with any endothelial domains in the heart. Furthermore, it has been difficult to ascertain whether endothelial heterogeneity in the heart influences underlying mesenchymal zones in an angiocrine manner. To identify regionally expressed EC genes in the heart valves, we screened publicly available databases and assembled a toolkit of endothelial-enriched genes. We identified Cyp26b1 as one of many endothelial enriched genes found to be expressed in the endocardium of the developing cushions and valves. Here, we show that Cyp26b1 is required for normal heart valve development. Genetic ablation of Cyp26b1 in mouse embryos leads to abnormally thickened aortic valve leaflets, which is due in part to increased endothelial and mesenchymal cell proliferation in the remodeling valves. In addition, Cyp26b1 mutant hearts display ventricular septal defects (VSDs) in a portion of null embryos. We show that loss of Cyp26b1 results in upregulation of retinoic acid (RA) target genes, supporting the observation that Cyp26b1 has RA-dependent roles. Together, this work identifies a novel role for Cyp26b1 in heart valve morphogenesis and points to a role of RA in this process. Understanding the spatiotemporal expression dynamics of cardiac EC genes will pave the way for investigation of both normal and dysfunctional heart valve development.

Cyp26b1 is an essential regulator of distal airway epithelial differentiation during lung development.

Proper organ development depends on coordinated communication between multiple cell types. Retinoic acid (RA) is an autocrine and paracrine signaling molecule essential for the development of most organs, including the lung. Despite extensive work detailing effects of RA deficiency in early lung morphogenesis, little is known about how RA regulates late gestational lung maturation. Here, we investigate the role of the RA catabolizing protein Cyp26b1 in the lung. Cyp26b1 is highly enriched in lung endothelial cells (ECs) throughout development. We find that loss of Cyp26b1 leads to reduction of alveolar type 1 cells, failure of alveolar inflation and early postnatal lethality in mouse. Furthermore, we observe expansion of distal epithelial progenitors, but no appreciable changes in proximal airways, ECs or stromal populations. Exogenous administration of RA during late gestation partially mimics these defects; however, transcriptional analyses comparing Cyp26b1-/- with RA-treated lungs reveal overlapping, but distinct, responses. These data suggest that defects observed in Cyp26b1-/- lungs are caused by both RA-dependent and RA-independent mechanisms. This work reports crucial cellular crosstalk during lung development involving Cyp26b1-expressing endothelium and identifies a novel RA modulator in lung development.

Vascular deficiencies in renal organoids and ex vivo kidney organogenesis.

Chronic kidney disease (CKD) and end stage renal disease (ESRD) are increasingly frequent and devastating conditions that have driven a surge in the need for kidney transplantation. A stark shortage of organs has fueled interest in generating viable replacement tissues ex vivo for transplantation. One promising approach has been self-organizing organoids, which mimic developmental processes and yield multicellular, organ-specific tissues. However, a recognized roadblock to this approach is that many organoid cell types fail to acquire full maturity and function. Here, we comprehensively assess the vasculature in two distinct kidney organoid models as well as in explanted embryonic kidneys. Using a variety of methods, we show that while organoids can develop a wide range of kidney cell types, as previously shown, endothelial cells (ECs) initially arise but then rapidly regress over time in culture. Vasculature of cultured embryonic kidneys exhibit similar regression. By contrast, engraftment of kidney organoids under the kidney capsule results in the formation of a stable, perfused vasculature that integrates into the organoid. This work demonstrates that kidney organoids offer a promising model system to define the complexities of vascular-nephron interactions, but the establishment and maintenance of a vascular network present unique challenges when grown ex vivo.

An anatomical feasibility study using CTA reconstruction for modified percutaneous lumbar vertebroplasty.

BACKGROUND: Lumbar vertebroplasty via several different types of extrapedicular approach has been reported with acceptable clinical results yet the anatomical basis for its safety is not fully explored. Injury to the lumbar arteries (LAs) is one of the most important potential complications. However, anatomical research on the course and variability of this structure is lacking. To investigate the anatomical feasibility of percutaneous vertebroplasty for lumbar osteoporotic vertebral compression fractures via a unilateral Extrapedicular approach. METHODS: A total of 300 LAs of 30 patients with non-spinal disorders were retrospectively analyzed by computed tomographic angiography (CTA). The lateral aspect of the vertebral body was divided into 9 zones of approximately equal area. The anatomy and orientation of LAs were analyzed in detail. RESULTS: LAs were most commonly found in the middle third of the body (zones 4, 5, and 6); the upper 1/3 of the vertebral body had LAs distributed only anteriorly and laterally (zones 1 and 2). No arteries were observed in the postero-superior segment (zone 3). From L1 to L3 an arched pattern predominated. At L4 an inferior oblique pattern (antero-superior to postero-inferior) predominated. Limited CTA visualization at L4 and particularly L5 as well as greater anatomical variation means that there is more uncertainty at these levels. CONCLUSION: From L1 to L3, the posterior superior segment (zone 1) of the vertebral body appears to be a safe area with low risk of arterial injury. This has relevance for design of a safe lumbar vertebral extrapedicular approach.

Baseline frailty status and outcomes important for shared decision-making in older adults receiving transcatheter aortic valve implantation, a prospective observational study.

AIMS: The objective of this study was to examine baseline frailty status (including cognitive deficits) and important clinical outcomes, to inform shared decision-making in older adults receiving transcatheter aortic valve implantation (TAVI). METHODS AND RESULTS: We conducted a prospective, observational study of 82 TAVI patients, recruited 2013 to 2015, with 2-year follow-up. Mean age was 83 years (standard deviation (SD) 4.7). Eighteen percent of the patients were frail, as assessed with an 8-item frailty scale. Fifteen patients (18%) had a Mini-Mental Status Examination (MMSE) score below 24 points at baseline, indicating cognitive impairment or dementia and five patients had an MMSE below 20 points. Mean New York Heart Association (NYHA) class at baseline and 6 months was 2.5 (SD 0.6) and 1.4 (SD 0.6), (p < 0.001). There was no change in mean Nottingham Extended Activities of Daily Living (NEADL) scale between baseline and 6 months, 54.2 (SD 11.5) and 54.5 (SD 10.3) points, respectively, mean difference 0.3 (p = 0.7). At 2 years, six patients (7%) had died, four (5%, n = 79) lived in a nursing home, four (5%) suffered from disabling stroke, and six (7%) contracted infective endocarditis. CONCLUSIONS: TAVI patients had improvement in symptoms and maintenance of activity of daily living at 6 months. They had low mortality and most patients lived in their own home 2 years after TAVI. Complications like death, stroke, and endocarditis occurred. Some patients had cognitive impairment before the procedure which might influence decision-making. Our findings may be used to develop pre-TAVI decision aids.

Exploring the impact of Patient Reported Outcome Measures (PROMs) among orthopaedic surgeons in mainland China: systematic review and survey-based study on hip and knee instruments.

BACKGROUND: Patient Reported Outcome Measures (PROMs) are widely used in Europe and North America in a variety of areas including research, clinical governance, clinical registries and insurance ascertainment. The aim of this study was to assess commonly used knee and hip PROMs among Chinese surgeons and to gain an insight into their impact on evaluation of clinical outcomes. METHODS: 1. A systematic literature search of databases Medline, EMBASE, CINAHL and CNKI was performed from the earliest records to 22/07/2020 for knee instruments and 22/08/2020 for hip instruments, to retrieve Chinese Mandarin cross culturally adapted and validated knee and hip PROMs. 2. An 11-item electronic questionnaire was then designed under four domain categories. The survey was distributed via a ubiquitous online social media platform to orthopaedic surgeons. Responses were collected and analyzed. Output from 1. was used to populate parts of the survey questionnaire. RESULTS: The systematic online search yielded a total of 41 evaluation instruments, (10 hip and 31 knee); all of which were incorporated as response options. 234 viable questionnaires were retrieved with the largest group representing attending surgeons. 59.0% were familiar with the concept of PROMs among which 78.4% reported to have used PROMs themselves. In order of frequency of use, PROMs were purposed for clinical assessment (55.6%), research (40.7%), health regulation policies (18.6%) and insurance service requirements (10.6%). Implementation was prompted by both departmental (43.4%) and institutional policy (34.5%). 89.4% of PROMs users reported difficulties in the use of PROMs, with major barriers including license fees, limited access, inadequate training and burden of fill-out time (all > 40%). CONCLUSION: There is evidence of limited familiarity with knee and hip PROMs among orthopaedic surgeons. Barriers to their use are significant. Development of a Chinese language PROMs database would be helpful.

The ethical landscape of gene drive research.

Gene drive technology has immense potential. The ability to bypass the laws of Mendelian inheritance and almost ensure the transmission of specific genetic material to future generations creates boundless possibilities. But alongside these boundless possibilities are major social and ethical issues. This article aims to introduce gene drive technology, some of its potential applications, and some of the social and ethical issues that arise during research into the technology. For example, is investigation into gene drives hubristic? Would applications of gene drives count as technological fixes? Or does research into such a technology sit on a slippery slope or lock us in to its full-scale use? Are there perverse effects of engaging in research, and, most importantly, who ought to be included in the decision-making process regarding research and field trials? Understanding the basic ethical landscape of this technology will prove invaluable to the public, scientists, and policy-makers as research moves forward.

Spatiotemporal heterogeneity and patterning of developing renal blood vessels.

The kidney vasculature facilitates the excretion of wastes, the dissemination of hormones, and the regulation of blood chemistry. To carry out these diverse functions, the vasculature is regionalized within the kidney and along the nephron. However, when and how endothelial regionalization occurs remains unknown. Here, we examine the developing kidney vasculature to assess its 3-dimensional structure and transcriptional heterogeneity. First, we observe that endothelial cells (ECs) grow coordinately with the kidney bud as early as E10.5, and begin to show signs of specification by E13.5 when the first arteries can be identified. We then focus on how ECs pattern and remodel with respect to the developing nephron and collecting duct epithelia. ECs circumscribe nephron progenitor populations at the distal tips of the ureteric bud (UB) tree and form stereotyped cruciform structures around each tip. Beginning at the renal vesicle (RV) stage, ECs form a continuous plexus around developing nephrons. The endothelial plexus envelops and elaborates with the maturing nephron, becoming preferentially enriched along the early distal tubule. Lastly, we perform transcriptional and immunofluorescent screens to characterize spatiotemporal heterogeneity in the kidney vasculature and identify novel regionally enriched genes. A better understanding of development of the kidney vasculature will help instruct engineering of properly vascularized ex vivo kidneys and evaluate diseased kidneys.

Reprogramming of human fibroblasts toward a cardiac fate.

Reprogramming of mouse fibroblasts toward a myocardial cell fate by forced expression of cardiac transcription factors or microRNAs has recently been demonstrated. The potential clinical applicability of these findings is based on the minimal regenerative potential of the adult human heart and the limited availability of human heart tissue. An initial but mandatory step toward clinical application of this approach is to establish conditions for conversion of adult human fibroblasts to a cardiac phenotype. Toward this goal, we sought to determine the optimal combination of factors necessary and sufficient for direct myocardial reprogramming of human fibroblasts. Here we show that four human cardiac transcription factors, including GATA binding protein 4, Hand2, T-box5, and myocardin, and two microRNAs, miR-1 and miR-133, activated cardiac marker expression in neonatal and adult human fibroblasts. After maintenance in culture for 4-11 wk, human fibroblasts reprogrammed with these proteins and microRNAs displayed sarcomere-like structures and calcium transients, and a small subset of such cells exhibited spontaneous contractility. These phenotypic changes were accompanied by expression of a broad range of cardiac genes and suppression of nonmyocyte genes. These findings indicate that human fibroblasts can be reprogrammed to cardiac-like myocytes by forced expression of cardiac transcription factors with muscle-specific microRNAs and represent a step toward possible therapeutic application of this reprogramming approach.

ATGme: Open-source web application for rare codon identification and custom DNA sequence optimization.

BACKGROUND: Codon usage plays a crucial role when recombinant proteins are expressed in different organisms. This is especially the case if the codon usage frequency of the organism of origin and the target host organism differ significantly, for example when a human gene is expressed in E. coli. Therefore, to enable or enhance efficient gene expression it is of great importance to identify rare codons in any given DNA sequence and subsequently mutate these to codons which are more frequently used in the expression host. RESULTS: We describe an open-source web-based application, ATGme, which can in a first step identify rare and highly rare codons from most organisms, and secondly gives the user the possibility to optimize the sequence. CONCLUSIONS: This application provides a simple user-friendly interface utilizing three optimization strategies: 1. one-click optimization, 2. bulk optimization (by codon-type), 3. individualized custom (codon-by-codon) optimization. ATGme is an open-source application which is freely available at: http://atgme.org.

Managing macromolecular crystallographic data with a laboratory information management system.

Protein crystallography is an established method to study the atomic structures of macromolecules and their complexes. A prerequisite for successful structure determination is diffraction-quality crystals, which may require extensive optimization of both the protein and the conditions, and hence projects can stretch over an extended period, with multiple users being involved. The workflow from crystallization and crystal treatment to deposition and publication is well defined, and therefore an electronic laboratory information management system (LIMS) is well suited to management of the data. Completion of the project requires key information on all the steps being available and this information should also be made available according to the FAIR principles. As crystallized samples are typically shipped between facilities, a key feature to be captured in the LIMS is the exchange of metadata between the crystallization facility of the home laboratory and, for example, synchrotron facilities. On completion, structures are deposited in the Protein Data Bank (PDB) and the LIMS can include the PDB code in its database, completing the chain of custody from crystallization to structure deposition and publication. A LIMS designed for macromolecular crystallography, IceBear, is available as a standalone installation and as a hosted service, and the implementation of key features for the capture of metadata in IceBear is discussed as an example.

Metastatic spinal cord compression: the Spinal Instability Neoplastic Score and early surgical intervention.

OBJECTIVES: To evaluate the value of Spinal Instability Neoplastic Score (SINS) in patients with spine metastasis who subsequently developed or did not develop metastatic spinal cord compression (MSCC). METHODS: In this single institutional retrospective descriptive observational study, of 589 patients with MSCC who were referred for radiotherapy, 34 patients (with 41 compression sites) met the inclusion criteria: availability of diagnostic MRI spine pre-development of MSCC (MRI-1) and at the time of MSCC development (MRI-2) (CordGroup).For comparison, NoCordGroup consisted of 152 patients (160 sites) treated with radiotherapy to spinal metastases. SINS was compared between the two groups. RESULTS: In CordGroup, the median interval between MRI-1 and MRI-2 was 11 weeks. The median SINS was 8 (range: 4-14) and 9 (range: 7-14) on MRI-1 and MRI-2, respectively. In NoCordGroup, the median SINS was 6 (range: 4-10). CONCLUSIONS: Our study showed a trend in difference in SINS value between the two groups. This difference should be a subject of future prospective research in this patient population with poor survival.

Work Patterns and Intensity of Chinese Surgical Residents- A Multicenter Time-and-Motion Study.

OBJECTIVE: This study aimed to record and analyze surgical resident trainee time allocation among junior doctors in China in order to understand the training environment and optimize realistic training and patient care objectives. DESIGN: Multicenter observational time and motion study. SETTING: Multicenter, carried out in 5 tier 3 public hospitals in 5 provinces across China. PARTICIPANTS: Surgical resident trainees at various stages of training were eligible to enter the study, total n = 44. Registered nurses were eligible to be observers, n = 4 from each hospital.  An expert team comprising 4 chief surgeons and 10 surgical residents participated in establishing the clinical activity list. RESULTS: Participants were observed during working hours (08.00-17.00) for 10 consecutive working days and time spent on different activities were recorded. Work patterns between hospitals were often dissimilar. Most time was spent on direct patient care (34.1%; 95% CI, 28.0%-40.1%) followed by indirect patient care (24.4%; 95% CI, 15.5%-33.2%), scholarly activity (21.1%; 95% CI, 13.7%-28.5%) and other (20.4%; 95% CI, 14.1%-26.8%). Subcategory analysis showed that the amount of time spent each day performing certain tasks was 137 minutes for operating theatre tasks, 103 minutes for medical record-keeping, 25 minutes for direct patient contact, 20 minutes being taught, 12 minutes teaching others, 12 minutes hand-over time, and 0 minutes of outpatient clinic attendance. Inter-observer reliability of 96.5% was obtained prior to recordings. CONCLUSIONS: Chinese surgical resident work patterns fall within the range found in other international studies albeit with some exceptions. The training environment appears broadly suitable for competence-based surgical training in China. Inadequate outpatient activity has led to changes in trainee work rosters and trainer requirements. Both strengths and deficiencies were confirmed and addressed. Further audit is required.

COVID-19 trends at the University of Tennessee: predictive insights from raw sewage SARS-CoV-2 detection and evaluation and PMMoV as an indicator for human waste.

Wastewater-based epidemiology (WBE) has become a valuable tool for monitoring the prevalence of SARS-CoV-2 on university campuses. However, concerns about effectiveness of raw sewage as a COVID-19 early warning system still exist, and it's not clear how useful normalization by simultaneous comparison of Pepper Mild Mottle Virus (PMMoV) is in addressing variations resulting from fecal discharge dilution. This study aims to contribute insights into these aspects by conducting an academic-year field trial at the student residences on the University of Tennessee, Knoxville campus, raw sewage. This was done to investigate the correlations between SARS-CoV-2 RNA load, both with and without PMMoV normalization, and various parameters, including active COVID-19 cases, self-isolations, and their combination among all student residents. Significant positive correlations between SARS-CoV-2 RNA load a week prior, during the monitoring week, and the subsequent week with active cases. Despite these correlations, normalization by PMMoV does not enhance these associations. These findings suggest the potential utility of SARS-CoV-2 RNA load as an early warning indicator and provide valuable insights into the application and limitations of WBE for COVID-19 surveillance specifically within the context of raw sewage on university campuses.

Evaluation of venous thromboembolism risk assessment models for hospital inpatients: the VTEAM evidence synthesis.

Background: Pharmacological prophylaxis during hospital admission can reduce the risk of acquired blood clots (venous thromboembolism) but may cause complications, such as bleeding. Using a risk assessment model to predict the risk of blood clots could facilitate selection of patients for prophylaxis and optimise the balance of benefits, risks and costs. Objectives: We aimed to identify validated risk assessment models and estimate their prognostic accuracy, evaluate the cost-effectiveness of different strategies for selecting hospitalised patients for prophylaxis, assess the feasibility of using efficient research methods and estimate key parameters for future research. Design: We undertook a systematic review, decision-analytic modelling and observational cohort study conducted in accordance with Enhancing the QUAlity and Transparency Of health Research (EQUATOR) guidelines. Setting: NHS hospitals, with primary data collection at four sites. Participants: Medical and surgical hospital inpatients, excluding paediatric, critical care and pregnancy-related admissions. Interventions: Prophylaxis for all patients, none and according to selected risk assessment models. Main outcome measures: Model accuracy for predicting blood clots, lifetime costs and quality-adjusted life-years associated with alternative strategies, accuracy of efficient methods for identifying key outcomes and proportion of inpatients recommended prophylaxis using different models. Results: We identified 24 validated risk assessment models, but low-quality heterogeneous data suggested weak accuracy for prediction of blood clots and generally high risk of bias in all studies. Decision-analytic modelling showed that pharmacological prophylaxis for all eligible is generally more cost-effective than model-based strategies for both medical and surgical inpatients, when valuing a quality-adjusted life-year at £20,000. The findings were more sensitive to uncertainties in the surgical population; strategies using risk assessment models were more cost-effective if the model was assumed to have a very high sensitivity, or the long-term risks of post-thrombotic complications were lower. Efficient methods using routine data did not accurately identify blood clots or bleeding events and several pre-specified feasibility criteria were not met. Theoretical prophylaxis rates across an inpatient cohort based on existing risk assessment models ranged from 13% to 91%. Limitations: Existing studies may underestimate the accuracy of risk assessment models, leading to underestimation of their cost-effectiveness. The cost-effectiveness findings do not apply to patients with an increased risk of bleeding. Mechanical thromboprophylaxis options were excluded from the modelling. Primary data collection was predominately retrospective, risking case ascertainment bias. Conclusions: Thromboprophylaxis for all patients appears to be generally more cost-effective than using a risk assessment model, in hospitalised patients at low risk of bleeding. To be cost-effective, any risk assessment model would need to be highly sensitive. Current evidence on risk assessment models is at high risk of bias and our findings should be interpreted in this context. We were unable to demonstrate the feasibility of using efficient methods to accurately detect relevant outcomes for future research. Future work: Further research should evaluate routine prophylaxis strategies for all eligible hospitalised patients. Models that could accurately identify individuals at very low risk of blood clots (who could discontinue prophylaxis) warrant further evaluation. Study registration: This study is registered as PROSPERO CRD42020165778 and Researchregistry5216. Funding: This award was funded by the National Institute for Health and Care Research (NIHR) Health Technology Assessment programme (NIHR award ref: NIHR127454) and will be published in full in Health Technology Assessment; Vol. 28, No. 20. See the NIHR Funding and Awards website for further award information.

People who are admitted to hospital are at risk of blood clots that can cause serious illness or death. Patients are often given low doses of blood-thinning drugs to reduce this risk. However, these drugs can cause side effects, such as bleeding. Hospitals currently use complex risk assessment models (risk scores, which usually include patient, disease, mobility and intervention factors) to determine the individual risk of blood clots and identify people most likely to benefit from blood-thinning drugs. There are a lot of different risk scores and we do not know which one is best. We also do not know how these scores compare to each other or whether using scores to decide who should get blood-thinning drugs provides good value for money to the NHS. We reviewed all previous studies of risk scores. We found that they did not predict blood clots very well and we could not recommend one score over another. We then created a mathematical model to simulate the use of blood-thinning drugs in people admitted to hospital. The model suggested that giving blood-thinning drugs to everyone who could have them would probably provide the best value for money, in medical patients. Our findings were the same, but less certain, for surgical patients. We also collected information from four NHS hospitals to explore possibilities for future research. Our work showed that routinely collected electronic data on blood clots and bleeding events is not very accurate and that using different scores could result in variable use of blood-thinning medications. Our findings suggest that it may be better value to the NHS and better for patients if we were to offer blood-thinning medications to everyone on admission to hospital, without using any risk score. However, this approach needs further research to ensure it is safe and effective. Such research would not be able to rely on routine electronic data to identify blood clots or bleeding events, in isolation.

Undisclosed Placebo Trials in Clinical Practice: Undercover Beneficence or Unwarranted Deception?

A placebo is an intervention that is believed to lack specific pharmacological or physiological efficacy for a patient's condition. While placebo-controlled trials are considered the gold standard when it comes to researching and testing new pharmacological treatments, the use of placebos in clinical practice is more controversial. The focus of this case study is an undisclosed placebo trial used as an attempt to diagnose a patient's complex and unusual symptomology. In this case, the placebo was used not just as a treatment, but as a diagnostic intervention in order to determine the best course of treatment for a patient. Could the deceptive use of a placebo be justified in clinical practice on the grounds of beneficence?

Exploiting Field Dependencies for Learning on Categorical Data.

Traditional approaches for learning on categorical data underexploit the dependencies between columns (a.k.a. fields) in a dataset because they rely on the embedding of data points driven alone by the classification/regression loss. In contrast, we propose a novel method for learning on categorical data with the goal of exploiting dependencies between fields. Instead of modelling statistics of features globally (i.e., by the covariance matrix of features), we learn a global field dependency matrix that captures dependencies between fields and then we refine the global field dependency matrix at the instance-wise level with different weights (so-called local dependency modelling) w.r.t. each field to improve the modelling of the field dependencies. Our algorithm exploits the meta-learning paradigm, i.e., the dependency matrices are refined in the inner loop of the meta-learning algorithm without the use of labels, whereas the outer loop intertwines the updates of the embedding matrix (the matrix performing projection) and global dependency matrix in a supervised fashion (with the use of labels). Our method is simple yet it outperforms several state-of-the-art methods on six popular dataset benchmarks. Detailed ablation studies provide additional insights into our method.