Protocol for generation and assessment of head-like structure in zebrafish.

In vitro embryonic analogue models, such as gastruloids, trunk-like structures and embryoids, have been developed to understand principles of early development and morphogenesis. However, models that can fully mimic head formation are still missing. Here, we present a protocol for generating the head-like structure (HLS) in zebrafish embryonic explants. We describe steps for dissection and constructing cell and patterning landscapes. We then detail assessment of this structure through axis induction. For complete details on the use and execution of this protocol, please refer to Cheng et al. (2023).1.

Endoscopy training in Australia during COVID-19: Efficacy and knowledge assessment of gastroenterology and general surgery trainees.

Background and Aim: During COVID-19, restrictions to elective endoscopy were introduced worldwide. A reduction in procedures may impact trainees' endoscopy learning. This study aims to assess Australian advanced gastroenterology and general surgery trainees' self-perceived efficacy and knowledge in endoscopy during the pandemic. Methods: All Australian gastroenterology and general surgery trainees in their last 2 years of accredited training were invited to participate through email (2020-2021 and 2021-2022 training cycles). The primary outcome was to assess trainees' self-efficacy and knowledge regarding gastrointestinal endoscopy. Secondary outcomes included subgroup analysis between gastroenterology and general surgery trainees. Self-perceived efficacy was assessed with Likert-scale questions on 20 endoscopy procedures and knowledge was assessed through 21 endoscopy-related multiple choice questions. Results: Eighty-one trainees responded to a self-efficacy questionnaire and 77 responded to the knowledge questionnaire. Over 90% of the trainees were confident or extremely confident in diagnostic endoscopy, but only half demonstrated similar efficacy for therapeutic endoscopy. The efficacy for basic endoscopy procedures was higher for gastroenterology trainees (64.0% vs 51.1%, P < 0.001). Last-year trainee achievement of conjoint committee requirements for upper gastrointestinal endoscopy was achieved in 95.8% of gastroenterology trainees versus 22.2% of surgical trainees (P < 0.001). The median score on the knowledge questionnaire was also higher for the gastroenterology subset (90.5% vs 71.4%, P < 0.001). Conclusion: During COVID-19, endoscopy trainees' self-efficacy in endoscopic diagnostic procedures was achieved for most trainees. The differences in self-perceived efficacy and knowledge between gastroenterology and surgical trainees may be reflective of the different opportunities for learning between the two groups.

Benefits of phone consultation for endoscopy-related clinics in the COVID-19 pandemic.

BACKGROUND AND AIM: During COVID-19 outbreak, restrictions to in-person consultations were introduced with a rise in telehealth. An indirect benefit of telehealth could be better attendance. This study aimed to assess "failure-to-attend" (FTA) rate and satisfaction for two endoscopy-related compulsory telehealth clinics during the COVID-19 outbreak. METHODS: Consecutive patients booked for endoscopy-related telehealth clinics at a tertiary hospital were prospectively assessed. In-person clinic control data were assessed retrospectively. Sample size was calculated to detect an anticipated increase in attendance of 8%. Secondary outcomes included FTA differences between clinics and evaluation of patients and doctors satisfaction. Satisfaction was assessed based on six Likert scale questions used in previous telehealth research and asked to both patients and doctors (6Q_score). This study was exempt from IRB review after institutional IRB review. RESULTS: There were 691 patients booked for appointments in our endoscopy clinics during the study periods (373 in 2020). FTA rates were lowered by half during the compulsory telehealth clinics (12.6% to 6.4%, P < 0.01). The patient 6Q_score was higher for the advanced endoscopy clinic (84.6% vs 73.8%, P < 0.01), while the doctor 6Q_score was similar between both advanced clinics and post endoscopy clinics (91.1% vs 92.5% respectively, P = 0.80). An in-person follow-up consultation was suggested for 3.5% of the appointments, while the necessity of physical examination was flagged in 5.1%. CONCLUSIONS: The use of phone consultations in endoscopy-related clinics during the COVID-19 outbreak has improved FTA rates while demonstrating high satisfaction rates. The need for in-person follow-up consultations and physical examination were low.

Narrow-band imaging with magnification and the water immersion technique: a case-finding, cost-effective approach to diagnose villous atrophy.

INTRODUCTION: Narrow-band imaging with magnification endoscopy (NBI-ME) allows real-time visual assessment of the mucosal surface and vasculature of the gastrointestinal tract. This study aimed to determine the performance of NBI-ME combined with the water immersion technique (NBI-ME-WIT) in detecting villous atrophy. METHODS: All patients who underwent gastroscopy were included. The duodenum was further examined with NBI-ME-WIT only after examination with white light endoscopy did not reveal a cause of anaemia or dyspepsia. Targeted biopsies were taken of visualised areas. NBI-ME-WIT findings were compared with the final histopathological analysis. We calculated the sensitivity (Sn), specificity (Sp), positive predictive value (PPV) and negative predictive value (NPV) of NBI-ME-WIT in detecting villous atrophy and the hypothetical cost saved by using a biopsy-avoiding approach. RESULTS: 124 patients (83 female) with a mean age of 46 (range 18-82) years were included. The most common indication for gastroscopy was abdominal pain (39%), followed by anaemia (35%), chronic diarrhoea/altered bowel habits (19%) and dyspepsia (6%). NBI-ME-WIT was able to detect all nine patients with villous atrophy - eight patchy and one total villous atrophy. The Sn, Sp, PPV and NPV of NBI-ME-WIT in detecting villous atrophy were 100.0%, 99.1%, 90.0% and 100.0%, respectively. Taking into account the cost of biopsy forceps (AUD 17) and pathology (AUD 140), this biopsy-avoidance strategy could have saved AUD 18,055 in these patients. CONCLUSION: NBI-ME-WIT is a specific and sensitive tool to recognise and accurately diagnose villous atrophy. Biopsies can be avoided in patients with normal-sized villi, which may decrease the overall cost of the procedure.

The safe and risk assessment of perioperative antiplatelet and anticoagulation therapy in inguinal hernia repair, a systematic review.

BACKGROUND: There is a lack of consensus on the continuation or cessation of perioperative antiplatelet and anticoagulation therapies in inguinal hernia repair, therefore, the local hospital protocols dealing with these agents vary. The aim of this study is to investigate the risk and safety of perioperative antiplatelet and anticoagulation therapies in patients undergoing elective inguinal hernia repair. METHODS: The major databases (PubMed, Embase, Springer, and Cochrane Library) were searched, and all studies published through January 2019 were identified, using the keywords Aspirin, Clopidogrel, Warfarin, antiplatelet, anticoagulation, inguinal hernia repair, bleeding, hematoma, complications. All relevant articles and reference lists in these original studies were also obtained from the above databases. RESULTS: Thirteen articles were identified, ten studies reported the results of perioperative application of antiplatelet therapy during inguinal hernia repair, nine trials reported the outcomes perioperative use of anticoagulation therapy in inguinal hernia repair, and six articles reported the results of both agents. One study did not indicate the perioperative cessation or continuation of the corresponding agents. Cessation of antiplatelets was reported in three studies and continuation of antiplatelets was used in six studies, they all demonstrated similar incidence of the bleeding complications, as compared with controls. Continued perioperative anticoagulation with Warfarin was reported in six studies and Warfarin discontinuation 3 days prior to operation was examined in two studies, both reported similar postoperative bleeding-related complication rates providing international normalized ratio (INR) < 3, and the postoperative hematoma incidence increased in the condition of INR > 3. CONCLUSION: Our results indicated that, for both open and laparoscopic inguinal hernia repair, there is no need to stop the antiplatelet therapy (Aspirin or Clopidogrel), and due to the limited evidence and the complexity of each patient' condition, the continuation or cessation of anticoagulation with Warfarin should be tailored on a case-by-case basis.

Improved estimation of aboveground biomass in wheat from RGB imagery and point cloud data acquired with a low-cost unmanned aerial vehicle system.

BACKGROUND: Aboveground biomass (AGB) is a widely used agronomic parameter for characterizing crop growth status and predicting grain yield. The rapid and accurate estimation of AGB in a non-destructive way is useful for making informed decisions on precision crop management. Previous studies have investigated vegetation indices (VIs) and canopy height metrics derived from Unmanned Aerial Vehicle (UAV) data to estimate the AGB of various crops. However, the input variables were derived either from one type of data or from different sensors on board UAVs. Whether the combination of VIs and canopy height metrics derived from a single low-cost UAV system can improve the AGB estimation accuracy remains unclear. This study used a low-cost UAV system to acquire imagery at 30 m flight altitude at critical growth stages of wheat in Rugao of eastern China. The experiments were conducted in 2016 and 2017 and involved 36 field plots representing variations in cultivar, nitrogen fertilization level and sowing density. We evaluated the performance of VIs, canopy height metrics and their combination for AGB estimation in wheat with the stepwise multiple linear regression (SMLR) and three types of machine learning algorithms (support vector regression, SVR; extreme learning machine, ELM; random forest, RF). RESULTS: Our results demonstrated that the combination of VIs and canopy height metrics improved the estimation accuracy for AGB of wheat over the use of VIs or canopy height metrics alone. Specifically, RF performed the best among the SMLR and three machine learning algorithms regardless of using all the original variables or selected variables by the SMLR. The best accuracy (R 2 = 0.78, RMSE = 1.34 t/ha, rRMSE = 28.98%) was obtained when applying RF to the combination of VIs and canopy height metrics. CONCLUSIONS: Our findings implied that an inexpensive approach consisting of the RF algorithm and the combination of RGB imagery and point cloud data derived from a low-cost UAV system at the consumer-grade level can be used to improve the accuracy of AGB estimation and have potential in the practical applications in the rapid estimation of other growth parameters.

An efficient and cost-effective method for primer-induced nucleotide labeling for massive sequencing on next-generation sequencing platforms.

Next generation sequencing (NGS) technologies play a powerful role in the preparation of large DNA databases such as DNA barcoding since it can produce a large number of sequence reads. Here we demonstrate a primer-induced sample labeling method aiming at sequencing a large number of samples simultaneously on NGS platforms. The strategy is to label samples with a unique oligo attached to the 5'-ends of primers. As a case study, 894 unique pentanucleotide oligoes were attached to the 5'-ends of three pairs of primers (for amplifying ITS, matK and rbcL) to label 894 samples. All PCR products of three barcodes of 894 samples were mixed together and sequenced on a high throughput sequencing platform. The results showed that 87.02%, 89.15% and 95.53% of the samples were successfully sequenced for rbcL, matK and ITS, respectively. The mean ratio of label mismatches for the three barcodes was 5.68%, and a sequencing depth of 30 ×to 40× was enough to obtain reliable sequences. It is flexible to label any number of samples simply by adjusting the length of oligoes. This easy, reliable and cost efficient method is useful in sequencing a large number of samples for construction of reference libraries for DNA barcoding, population biology and community phylogenetics.

Mechanism and kinetics of the electrocatalytic reaction responsible for the high cost of hydrogen fuel cells.

The sluggish oxygen reduction reaction (ORR) is a major impediment to the economic use of hydrogen fuel cells in transportation. In this work, we report the full ORR reaction mechanism for Pt(111) based on Quantum Mechanics (QM) based Reactive metadynamics (RµD) simulations including explicit water to obtain free energy reaction barriers at 298 K. The lowest energy pathway for 4 e- water formation is: first, *OOH formation; second, *OOH reduction to H2O and O*; third, O* hydrolysis using surface water to produce two *OH and finally *OH hydration to water. Water formation is the rate-determining step (RDS) for potentials above 0.87 Volt, the normal operating range. Considering the Eley-Rideal (ER) mechanism involving protons from the solvent, we predict the free energy reaction barrier at 298 K for water formation to be 0.25 eV for an external potential below U = 0.87 V and 0.41 eV at U = 1.23 V, in good agreement with experimental values of 0.22 eV and 0.44 eV, respectively. With the mechanism now fully understood, we can use this now validated methodology to examine the changes upon alloying and surface modifications to increase the rate by reducing the barrier for water formation.

Enhanced Generation of Integration-free iPSCs from Human Adult Peripheral Blood Mononuclear Cells with an Optimal Combination of Episomal Vectors.

We previously reported the generation of integration-free induced pluripotent stem cells from adult peripheral blood (PB) with an improved episomal vector (EV) system, which uses the spleen focus-forming virus U3 promoter and an extra factor BCL-XL (B). Here we show an ∼100-fold increase in efficiency by optimizing the vector combination. The two most critical factors are: (1) equimolar expression of OCT4 (O) and SOX2 (S), by using a 2A linker; (2) a higher and gradual increase in the MYC (M) to KLF4 (K) ratio during the course of reprogramming, by using two individual vectors to express M and K instead of one. The combination of EV plasmids (OS + M + K + B) is comparable with Sendai virus in reprogramming efficiency but at a fraction of the cost. The generated iPSCs are indistinguishable from those from our previous approach in pluripotency and phenotype. This improvement lays the foundation for broad applications of episomal vectors in PB reprogramming.

Quality assessment of traditional Chinese medicine herb couple by high-performance liquid chromatography and mass spectrometry combined with chemometrics.

This study was designed to develop a simple, specific and reliable method to overall analyze the chemical constituents in clematidis radix et rhizome/notopterygii rhizome et radix herb couple using high-performance liquid chromatography coupled with tandem mass spectrometry and multiple chemometric analysis. First, the separation and qualitative analysis of herb couple was achieved on an Agilent Zorbax Eclipse Plus C18 column (250 mm × 4.6 mm, 5 µm), and 69 compounds were unambiguously or tentatively identified. Moreover, in quantitative analysis, eight ingredients including six coumarins and two triterpenoid sapogenins were quantified by high-performance liquid chromatography coupled with tandem mass spectrometry. In terms of good linearity (r(2) ≥ 0.9995) with a relatively wide concentration range, recovery (85.40-102.50%) and repeatability (0.99-4.45%), the validation results suggested the proposed method was reliable, and successfully used to analyze ten batches of herb couple samples. Then, hierarchical cluster analysis and principal component analysis were used to classify samples and search significant ingredients. The results showed that ten batches of herb couple samples were classified into three groups, and six compounds were found for its better quality control.

Uniform manganese hexacyanoferrate hydrate nanocubes featuring superior performance for low-cost supercapacitors and nonenzymatic electrochemical sensors.

Uniform manganese hexacyanoferrate hydrate nanocubes are prepared via a simple chemical precipitation method at room temperature. Due to both micro/mesopores of the Prussian blue analogue and nanocubic structures, the manganese hexacyanoferrate hydrate nanocubes allow the efficient charge transfer and mass transport for electrolyte solution and chemical species. Thus, the manganese hexacyanoferrate hydrate nanocube electrode shows a good rate capability and cycling stability for electrochemical capacitors. Furthermore, electrodes modified with manganese hexacyanoferrate hydrate nanocubes demonstrate a sensitive electrochemical response to hydrogen peroxide (H2O2) in buffer solutions with a high selectivity.

Flexible supercapacitors based on paper substrates: a new paradigm for low-cost energy storage.

Paper-based supercapacitors (SCs), a novel and interesting group of flexible energy storage devices, are attracting more and more attention from both industry and academia. Cellulose papers with a unique porous bulk structure and rough and absorptive surface properties enable the construction of paper-based SCs with a reasonably good performance at a low price. The inexpensive and environmentally friendly nature of paper as well as simple fabrication techniques make paper-based SCs promising candidates for the future 'green' and 'once-use-and-throw-away' electronics. This review introduces the design, fabrication and applications of paper-based SCs, giving a comprehensive coverage of this interesting field. Challenges and future perspectives are also discussed.

Pathological features of localized prostate cancer in China: a contemporary analysis of radical prostatectomy specimens.

There has been a rapid increase in the incidence of prostate cancer in China, especially in areas with boosted economic development. In this study, we analyzed the pathological features of a contemporary series of radical prostatectomy cases. A total of 230 consecutive, whole-mounted radical prostatectomy specimens collected from 2012 to 2014 were reviewed. The median age of the patients was 68 years, and 64.3% of patients presented with prostate specific antigen alone. Pathological examination indicated that a high proportion (77.4%) of patients had intermediate- or high-risk disease according to the Cancer of the Prostate Risk Assessment Post-Surgical score. After surgery, only 28 patients met the criteria for active surveillance (organ-confined Gleason ≥6 disease). The Prostate Cancer Research International Active Surveillance criteria achieved a sensitivity of 57.1% and a specificity of 98.0% for identifying candidates. The probability of Gleason score upgrading was 24.8% in the entire group and 59.0% in biopsy-confirmed Gleason ≥6 disease. The predominant tumor was located in the transition zone in 14.8% of cases, while only three patients (1.3%) had a predominant tumor located in the anterior region. Patients with transition zone-predominant tumor were likely to have been referred with urinary symptoms and high prostate specific antigen levels. The results of this study highlight the contemporary pathological features of localized prostate cancer in urban China. There was an increased trend towards asymptomatic cases, though most patients had intermediate- or high-risk disease and were suitable for definitive treatment. The low prevalence of dominant cancer in the anterior region may reflect race-based pathological differences.

Assessment of hematopoietic failure due to Rpl11 deficiency in a zebrafish model of Diamond-Blackfan anemia by deep sequencing.

BACKGROUND: Diamond-Blackfan anemia is a rare congenital red blood cell dysplasia that develops soon after birth. RPL11 mutations account for approximately 4.8% of human DBA cases with defective hematopoietic phenotypes. However, the mechanisms by which RPL11 regulates hematopoiesis in DBA remain elusive. In this study, we analyzed the transcriptome using deep sequencing data from an Rpl11-deficient zebrafish model to identify Rpl11-mediated hematopoietic failure and investigate the underlying mechanisms. RESULTS: We characterized hematological defects in Rpl11-deficient zebrafish embryos by identifying affected hematological genes, hematopoiesis-associated pathways, and regulatory networks. We found that hemoglobin biosynthetic and hematological defects in Rpl11-deficient zebrafish were related to dysregulation of iron metabolism-related genes, including tfa, tfr1b, alas2 and slc25a37, which are involved in heme and hemoglobin biosynthesis. In addition, we found reduced expression of the hematopoietic stem cells (HSC) marker cmyb and HSC transcription factors tal1 and hoxb4a in Rpl11-deficient zebrafish embryos, indicating that the hematopoietic defects may be related to impaired HSC formation, differentiation, and proliferation. However, Rpl11 deficiency did not affect the development of other blood cell lineages such as granulocytes and myelocytes. CONCLUSION: We identified hematopoietic failure of Rpl11-deficient zebrafish embryos using transcriptome deep sequencing and elucidated potential underlying mechanisms. The present analyses demonstrate that Rpl11-deficient zebrafish may serve as a model of DBA and may provide insights into the pathogenesis of mutant RPL11-mediated human DBA disease.

Toxicological effects involved in risk assessment of rare earth lanthanum on roots of Vicia faba L. seedlings.

Combined chemical analyses and biological measurements were utilized to investigate potential toxicological effects and possible mechanisms involved in risk assessment of rare earth elements (REEs) on Vicia faba L. seedlings, which were hydroponically cultivated and exposed to various concentrations of lanthanum (La) for 15 days. The results showed that La contents in both the solution and roots increased with the increase of extraneous La, contributing to hormetic dose responses of superoxide dismutase (SOD), catalase (CAT), guaiacol peroxidase (GPX), ascorbate peroxidase (APX) and endoprotease (EP) isozymes activities, and HSP 70 production enhanced at low doses but suppressed at higher doses of La. These physiological responses constituted antioxidant and detoxification systems against La-induced oxidative stress. The elevated La levels also contributed to oxidatively modified proteins, which were most responsible for subsequent cell death and growth retardation of the roots. By combination of hormetic and traditional threshold dose levels, the threshold dose range was deduced to be 108-195 microg La/g dry weight in the roots, corresponding to 0.90-3.12 mg/L of soluble La in the culture solution. It suggests that persistent applications of REEs may lead to potential ecological risk in the environment.