The COVID-19 paradox of online collaborative education: when you cannot physically meet, you need more social interactions.

Collaborative learning is a teaching method that brings together students to discuss a topic important for a given course or curriculum and solve a related problem or create a product. By doing this, learners create knowledge together and gain 21st -century skills such as communication, critical thinking, decision making, leadership and conflict management. Universities had to close their campuses and turn their education fully online in 2020 due to the COVID-19 pandemic, which created a forced step in the evolution of the digitalisation of collaborative teaching. How did TU Delft face this challenge? How did the students experience the online version of collaborative learning? How did distant learning affect their motivation? This article presents four student team projects investigating these questions from the collaborative learning perspective. One of the significant findings of these projects is the lack of socio-emotional interactions during online collaborative work. We present a few guidelines on how to enable these interactions when designing online or blended collaborative education.

Dosage uniformity problems which occur due to technological errors in extemporaneously prepared suppositories in hospitals and pharmacies.

The availability of suppositories in Hungary, especially in clinical pharmacy practice, is usually provided by extemporaneous preparations. Due to the known advantages of rectal drug administration, its benefits are frequently utilized in pediatrics. However, errors during the extemporaneous manufacturing process can lead to non-homogenous drug distribution within the dosage units. To determine the root cause of these errors and provide corrective actions, we studied suppository samples prepared with exactly known errors using both cerimetric titration and HPLC technique. Our results show that the most frequent technological error occurs when the pharmacist fails to use the correct displacement factor in the calculations which could lead to a 4.6% increase/decrease in the assay in individual dosage units. The second most important source of error can occur when the molding excess is calculated solely for the suppository base. This can further dilute the final suppository drug concentration causing the assay to be as low as 80%. As a conclusion we emphasize that the application of predetermined displacement factors in calculations for the formulation of suppositories is highly important, which enables the pharmacist to produce a final product containing exactly the determined dose of an active substance despite the different densities of the components.

Simultaneous quantification of paracetamol, acetylsalicylic acid and papaverine with a validated HPLC method.

Combined drug products have the advantages of better patient compliance and possible synergic effects. The simultaneous application of several active ingredients at a time is therefore frequently chosen. However, the quantitative analysis of such medicines can be challenging. The aim of this study is to provide a validated method for the investigation of a multidose packed oral powder that contained acetylsalicylic acid, paracetamol and papaverine-HCl. Reversed-phase high-pressure liquid chromatography was used. The Agilent Zorbax SB-C18 column was found to be the most suitable of the three different stationary phases tested for the separation of the components of this sample. The key parameters in the method development (apart from the nature of the column) were the pH of the aqueous phase (set to 3.4) and the ratio of the organic (acetonitrile) and the aqueous (25 mM phosphate buffer) phases, which was varied from 7:93 (v/v) to 25:75 (v/v) in a linear gradient, preceded by an initial hold. The method was validated: linearity, precision (repeatability and intermediate precision), accuracy, specificity and robustness were all tested, and the results met the ICH guidelines.

Novel sample preparation method for surfactant containing suppositories: effect of micelle formation on drug recovery.

Rectal drug delivery is currently at the focus of attention. Surfactants promote drug release from the suppository bases and enhance the formulation properties. The aim of our work was to develop a sample preparation method for HPLC analysis for a suppository base containing 95% hard fat, 2.5% Tween 20 and 2.5% Tween 60. A conventional sample preparation method did not provide successful results as the recovery of the drug failed to fulfil the validation criterion 95-105%. This was caused by the non-ionic surfactants in the suppository base incorporating some of the drug, preventing its release. As guidance for the formulation from an analytical aspect, we suggest a well defined surfactant content based on the turbidimetric determination of the CMC (critical micelle formation concentration) in the applied methanol-water solvent. Our CMC data correlate well with the results of previous studies. As regards the sample preparation procedure, a study was performed of the effects of ionic strength and pH on the drug recovery with the avoidance of degradation of the drug during the procedure. Aminophenazone and paracetamol were used as model drugs. The optimum conditions for drug release from the molten suppository base were found to be 100 mM NaCl, 20-40 mM NaOH and a 30 min ultrasonic treatment of the final sample solution. As these conditions could cause the degradation of the drugs in the solution, this was followed by NMR spectroscopy, and the results indicated that degradation did not take place. The determined CMCs were 0.08 mM for Tween 20, 0.06 mM for Tween 60 and 0.04 mM for a combined Tween 20, Tween 60 system.

Automated high-throughput mapping of promoter-enhancer interactions in zebrafish embryos.

Zebrafish embryos offer a unique combination of high-throughput capabilities and the complexity of the vertebrate animal for a variety of phenotypic screening applications. However, there is a need for automation of imaging technologies to exploit the potential of the transparent embryo. Here we report a high-throughput pipeline for registering domain-specific reporter expression in zebrafish embryos with the aim of mapping the interactions between cis-regulatory modules and core promoters. Automated microscopy coupled with custom-built embryo detection and segmentation software allowed the spatial registration of reporter activity for 202 enhancer-promoter combinations, based on images of thousands of embryos. The diversity of promoter-enhancer interaction specificities underscores the importance of the core promoter sequence in cis-regulatory interactions and provides a promoter resource for transgenic reporter studies. The technology described here is also suitable for the spatial analysis of fluorescence readouts in genetic, pharmaceutical or toxicological screens.

Chaperone activity of ERD10 and ERD14, two disordered stress-related plant proteins.

ERD10 and ERD14 (for early response to dehydration) proteins are members of the dehydrin family that accumulate in response to abiotic environmental stresses, such as high salinity, drought, and low temperature, in Arabidopsis (Arabidopsis thaliana). Whereas these proteins protect cells against the consequences of dehydration, the exact mode(s) of their action remains poorly understood. Here, detailed evidence is provided that ERD10 and ERD14 belong to the family of intrinsically disordered proteins, and it is shown in various assays that they act as chaperones in vitro. ERD10 and ERD14 are able to prevent the heat-induced aggregation and/or inactivation of various substrates, such as lysozyme, alcohol dehydrogenase, firefly luciferase, and citrate synthase. It is also demonstrated that ERD10 and ERD14 bind to acidic phospholipid vesicles without significantly affecting membrane fluidity. Membrane binding is strongly influenced by ionic strength. Our results show that these intrinsically disordered proteins have chaperone activity of rather wide substrate specificity and that they interact with phospholipid vesicles through electrostatic forces. We suggest that these findings provide the rationale for the mechanism of how these proteins avert the adverse effects of dehydration stresses.

A multicassette Gateway vector set for high throughput and comparative analyses in ciona and vertebrate embryos.

BACKGROUND: The past few years have seen a vast increase in the amount of genomic data available for a growing number of taxa, including sets of full length cDNA clones and cis-regulatory sequences. Large scale cross-species comparisons of protein function and cis-regulatory sequences may help to understand the emergence of specific traits during evolution. PRINCIPAL FINDINGS: To facilitate such comparisons, we developed a Gateway compatible vector set, which can be used to systematically dissect cis-regulatory sequences, and overexpress wild type or tagged proteins in a variety of chordate systems. It was developed and first characterised in the embryos of the ascidian Ciona intestinalis, in which large scale analyses are easier to perform than in vertebrates, owing to the very efficient embryo electroporation protocol available in this organism. Its use was then extended to fish embryos and cultured mammalian cells. CONCLUSION: This versatile vector set opens the way to the mid- to large-scale comparative analyses of protein function and cis-regulatory sequences across chordate evolution. A complete user manual is provided as supplemental material.

Shuffling of cis-regulatory elements is a pervasive feature of the vertebrate lineage.

BACKGROUND: All vertebrates share a remarkable degree of similarity in their development as well as in the basic functions of their cells. Despite this, attempts at unearthing genome-wide regulatory elements conserved throughout the vertebrate lineage using BLAST-like approaches have thus far detected noncoding conservation in only a few hundred genes, mostly associated with regulation of transcription and development. RESULTS: We used a unique combination of tools to obtain regional global-local alignments of orthologous loci. This approach takes into account shuffling of regulatory regions that are likely to occur over evolutionary distances greater than those separating mammalian genomes. This approach revealed one order of magnitude more vertebrate conserved elements than was previously reported in over 2,000 genes, including a high number of genes found in the membrane and extracellular regions. Our analysis revealed that 72% of the elements identified have undergone shuffling. We tested the ability of the elements identified to enhance transcription in zebrafish embryos and compared their activity with a set of control fragments. We found that more than 80% of the elements tested were able to enhance transcription significantly, prevalently in a tissue-restricted manner corresponding to the expression domain of the neighboring gene. CONCLUSION: Our work elucidates the importance of shuffling in the detection of cis-regulatory elements. It also elucidates how similarities across the vertebrate lineage, which go well beyond development, can be explained not only within the realm of coding genes but also in that of the sequences that ultimately govern their expression.

Hsp90 isoforms: functions, expression and clinical importance.

The 90 kDa heat shock protein, Hsp90, is a main functional component of an important cytoplasmic chaperone complex, and it is involved in various cellular processes, such as cell proliferation, differentiation and apoptosis. Identification of Hsp90 as a molecular target of various anticancer drugs highlighted its importance from the clinical point of view. Here we summarize the current knowledge of various Hsp90 isoforms regarding their genomic location, molecular evolution, functional differences, differential induction after various environmental stresses and in pathological conditions as well as the growing importance of discriminating between Hsp90 isoforms in clinical practice.