Piloting a Basic Life Support instructor course: A short report.

Aim: The aim was to describe a new shortened pilot of the European Resuscitation Council's standard Basic Instructor Course. Methods: The four-hour pilot followed a blended learning strategy (pre-course preparation, on-site small-group sessions). Each participant taught a short Basic Life Support competency to the group (micro-teaching) and received the group's feedback. A feedback "drill" session followed. Primary quantitative outcome was the proportion of Basic Instructor Course participants subsequently teaching Basic Life Support. Post-course teachings were recorded and compared to standard eight-hour Basic Instructor Courses. Participants' open feedback question answers were qualitatively analyzed and presented descriptively. Results: This pilot Basic Instructor Course taught 31 healthcare providers in 4 courses in 2019-2021 (aged 31.5 ± 12.9 years; 61 % women; 29 % physicians; 71 % medical students; 21 % no teaching experience). Participants reported that they gained most from micro-teaching (64 %), and advice on their teaching (50 %). Some judged the course as being too long (29 %). Twenty-seven pilot course participants (87 %) (including three instructor candidates) started teaching, whereas only nine of 37 participants of the 3 courses (24 %, including three instructor candidates) from the standard eight-hour course did. Conclusion: Participants of the pilot shortened Basic Instructor Course in a healthcare setting were successfully trained to teach European Resuscitation Council's Basic Life Support provider courses in a short four-hour format. The pilot course seems to enable future instructors to teach Basic Life Support provider courses. Higher motivation to teach resulted in four times as many instructors who taught courses after the pilot course compared to the standard course.

Debriefing interaction patterns and learning outcomes in simulation: an observational mixed-methods network study.

BACKGROUND: Debriefing is effective and inexpensive to increase learning benefits of participants in simulation-based medical education. However, suitable communication patterns during debriefings remain to be defined. This study aimed to explore interaction patterns during debriefings and to link these to participants' satisfaction, perceived usefulness, and self-reported learning outcomes. METHODS: We assessed interaction patterns during debriefings of simulation sessions for residents, specialists, and nurses from the local anaesthesia department at the Bern University Hospital, Bern, Switzerland. Network analysis was applied to establish distinctive interaction pattern categories based on recorded interaction links. We used multilevel modelling to assess relationships between interaction patterns and self-reported learning outcomes. RESULTS: Out of 57 debriefings that involved 111 participants, discriminatory analyses revealed three distinctive interaction patterns: 'fan', 'triangle', and 'net'. Participants reported significantly higher self-reported learning effects in debriefings with a net pattern, compared to debriefings with a fan pattern. No effects were observed for participant satisfaction, learning effects after 1 month, and perceived usefulness of simulation sessions. CONCLUSIONS: A learner-centred interaction pattern (i.e. net) was significantly associated with improved short-term self-reported individual learning and team learning. This supports good-practice debriefing guidelines, which stated that participants should have a high activity in debriefings, guided by debriefers, who facilitate discussions to maximize the development for the learners.

Simulation-based medical education for Ambulance Jet and Helicopter Emergency Medical Services: A program description and evaluation.

This article was migrated. The article was marked as recommended. Introduction: In aviation, crew resource management trainings are established methods to enhance safety, a method that also gained popularity in medicine. In 2015, the Swiss Air Rescue (Rega) Helicopter Emergency Medical Services decided to start a simulation-based medical education program for its helicopter and ambulance jet crews (emergency physicians, paramedics/flight nurses and pilots). The aim of this program was to improve technical skills and the application of human factors during rescue missions. This report shows a five-year summary of the participants' course evaluation. Methods: A 1-day high-fidelity simulation on crisis resource management with video-assisted debriefing took place at 3 centres, two in Switzerland; one in Germany. Crew members participated once per year. Simulation covered critical situations in the helicopter or jet, during handovers at an intensive care unit or in ambulances. Extra Corporeal Membrane Oxygenation and Intra-Aortic Balloon Pump use was simulated during helicopter transports. Additionally, four times per year flight crews rehearsed basic and advanced life support skills using low-fidelity equipment between missions. Participants answered an anonymized course evaluation survey. Answers were rated on a Numeric Rating Scale ranging from 1=no agreement to 5=total agreement. Results: 329 participated and answered the questionnaire; 50% were emergency physicians, 40% paramedics, 9% flight nurses, and 1% pilots. Participants agreed that the course taught competencies that were useful for their clinical practice. However, confidence to apply Extra Corporeal Membrane Oxygenation or Intra-Aortic Balloon Pump skills was significantly lower compared to other emergency competencies. Instructors were rated as experienced, engaged and motivated, as well as responsive to course participants. Conclusions: This simulation-based medical education program, with the goal to increase patient's safety and outcome, was launched successfully. Participants especially valued the time to reflect on clinical performance as well as on crew interaction and ways to apply human factors to improve their team performance and task management.

Recruiting Medical Students for a First Responder Project in the Social Age: Direct Contact Still Outperforms Social Media.

INTRODUCTION: Efficient recruitment of first responders (FRs) is crucial for long-term success of any FR project. FRs are laypersons who are trained in cardiopulmonary resuscitation (CPR), medical professionals, and firemen, police officers, and other professions with a duty of help. As social media are widely used for rapid communication, we carried out a prospective observational study to test the hypothesis that recruitment of FRs via social media is more efficient than recruitment via direct face-to-face contact. METHODS: Following ethics committee agreement, we informed 600 medical students about becoming FRs when they attended a didactic lecture about the FR project or during their mandatory CPR-course. Furthermore, recruitment was opened to medical students through Facebook, which accessed ∼1,000 medical students to see if they expressed interest in becoming FRs. All of the recruited students successfully completed the FR training. We then used an online questionnaire to ask these students how they had been recruited. RESULTS: Out of 63 registered student FRs, 59 responded to the online questionnaire. Overall, 15.3% of these FR students were recruited via social media. The majority (78.0%) were recruited through direct contact. CONCLUSIONS: Despite widespread use of social media, over three-quarters of these medical students were recruited to the FR project via direct personal contact. This suggests that the advantage of a larger reachable population using social media does not outweigh the impact of personal contact with experts.

Barley as a green factory for the production of functional Flt3 ligand.

Biologically active recombinant human Flt3 ligand was expressed and isolated from transgenic barley seeds. Its expression is controlled by a tissue specific promoter that confines accumulation of the recombinant protein to the endosperm tissue of the seed. The recombinant Flt3 ligand variant expressed in the seeds contains an HQ-tag for affinity purification on immobilized metal ion affinity chromatography (IMAC) resin. The tagged protein was purified from seed extracts to near homogeneity using sequential chromatography on IMAC affinity resin and cation exchange resin. We also show that the recombinant Flt3 ligand protein undergoes posttranslational modifications: it is a glycoprotein containing alpha-1,3-fucose and alpha-1,2-xylose. The HQ-tagged Flt3 ligand variant exhibits comparable biological activity to commercial Flt3 ligand. This is the first report showing expression and accumulation of recombinant human growth factor in barley seeds with a yield of active protein similar to a bacterial expression system. The present results demonstrate that plant molecular farming is a viable approach for the bioproduction of human-derived growth factors.

Selective redox regulation of cytokine receptor signaling by extracellular thioredoxin-1.

The thiol-disulfide oxidoreductase thioredoxin-1 (Trx1) is known to be secreted by leukocytes and to exhibit cytokine-like properties. Extracellular effects of Trx1 require a functional active site, suggesting a redox-based mechanism of action. However, specific cell surface proteins and pathways coupling extracellular Trx1 redox activity to cellular responses have not been identified so far. Using a mechanism-based kinetic trapping technique to identify disulfide exchange interactions on the intact surface of living lymphocytes, we found that Trx1 catalytically interacts with a single principal target protein. This target protein was identified as the tumor necrosis factor receptor superfamily member 8 (TNFRSF8/CD30). We demonstrate that the redox interaction is highly specific for both Trx1 and CD30 and that the redox state of CD30 determines its ability to engage the cognate ligand and transduce signals. Furthermore, we confirm that Trx1 affects CD30-dependent changes in lymphocyte effector function. Thus, we conclude that receptor-ligand signaling interactions can be selectively regulated by an extracellular redox catalyst.

Thioredoxin target proteins in chloroplast thylakoid membranes.

In recent years, impressive progress has been made in the identification of thioredoxin-linked proteins. However, due to technical difficulties inherent in working with hydrophobic proteins, identifications so far have been restricted to proteins in the soluble fraction. Thus, our knowledge of redox regulated membrane proteins is quite limited. To gain information in this area, the authors have applied an adaptation of the approach based on the fluorescent thiol probe monobromobimane (mBBr) to identify redox-linked proteins of chloroplast thylakoids. By application of this procedure, 14 potential membrane-bound thioredoxin target proteins were identified, including seven new candidates functional in processes associated with photosynthetic electron flow, ATP synthesis, and Photosystem II/Photosystem I state transitions.

Proteome of amyloplasts isolated from developing wheat endosperm presents evidence of broad metabolic capability.

By contrast to chloroplasts, our knowledge of amyloplasts--organelles that synthesize and store starch in heterotrophic plant tissues--is in a formative stage. While our understanding of what is considered their primary function, i.e. the biosynthesis and degradation of starch, has increased dramatically in recent years, relatively little is known about other biochemical processes taking place in these organelles. To help fill this gap, a proteomic analysis of amyloplasts isolated from the starchy endosperm of wheat seeds (10 d post-anthesis) has been conducted. The study has led to the identification of 289 proteins that function in a range of processes, including carbohydrate metabolism, cytoskeleton/plastid division, energetics, nitrogen and sulphur metabolism, nucleic acid-related reactions, synthesis of various building blocks, protein-related reactions, transport, signalling, stress, and a variety of other activities grouped under 'miscellaneous'. The function of 12% of the proteins was unknown. The results highlight the role of the amyloplast as a starch-storing organelle that fulfills a spectrum of biosynthetic needs of the parent tissue. When compared with a recent proteomic analysis of whole endosperm, the current study demonstrates the advantage of using isolated organelles in proteomic studies.

A complete ferredoxin/thioredoxin system regulates fundamental processes in amyloplasts.

A growing number of processes throughout biology are regulated by redox via thiol-disulfide exchange. This mechanism is particularly widespread in plants, where almost 200 proteins have been linked to thioredoxin (Trx), a widely distributed small regulatory disulfide protein. The current study extends regulation by Trx to amyloplasts, organelles prevalent in heterotrophic plant tissues that, among other biosynthetic activities, catalyze the synthesis and storage of copious amounts of starch. Using proteomics and immunological methods, we identified the components of the ferredoxin/Trx system (ferredoxin, ferredoxin-Trx reductase, and Trx), originally described for chloroplasts, in amyloplasts isolated from wheat starchy endosperm. Ferredoxin is reduced not by light, as in chloroplasts, but by metabolically generated NADPH via ferredoxin-NADP reductase. However, once reduced, ferredoxin appears to act as established for chloroplasts, i.e., via ferredoxin-Trx reductase and a Trx (m-type). A proteomics approach in combination with affinity chromatography and a fluorescent thiol probe led to the identification of 42 potential Trx target proteins, 13 not previously recognized, including a major membrane transporter (Brittle-1 or ADP-glucose transporter). The proteins function in a range of processes in addition to starch metabolism: biosynthesis of lipids, amino acids, and nucleotides; protein folding; and several miscellaneous reactions. The results suggest a mechanism whereby light is initially recognized as a thiol signal in chloroplasts, then as a sugar during transit to the sink, where it is converted again to a thiol signal. In this way, amyloplast reactions in the grain can be coordinated with photosynthesis taking place in leaves.

Redox regulation: a broadening horizon.

Initially discovered in the context of photosynthesis, regulation by change in the redox state of thiol groups (S-S <--> 2SH) is now known to occur throughout biology. Several systems, each linking a hydrogen donor to an intermediary disulfide protein, act to effect changes that alter the activity of target proteins: the ferredoxin/thioredoxin system, comprised of reduced ferredoxin, a thioredoxin, and the enzyme, ferredoxin-thioredoxin reductase; the NADP/thioredoxin system, including NADPH, a thioredoxin, and NADP-thioredoxin reductase; and the glutathione/glutaredoxin system, composed of reduced glutathione and a glutaredoxin. A related disulfide protein, protein disulfide isomerase (PDI) acts in protein assembly. Regulation linked to plastoquinone and signaling induced by reactive oxygen species (ROS) and other agents are also being actively investigated. Progress made on these systems has linked redox to the regulation of an increasing number of processes not only in plants, but in other types of organisms as well. Research in areas currently under exploration promises to provide a fuller understanding of the role redox plays in cellular processes, and to further the application of this knowledge to technology and medicine.

Structural analysis uncovers a role for redox in regulating FKBP13, an immunophilin of the chloroplast thylakoid lumen.

Change in redox status has long been known to link light to the posttranslational regulation of chloroplast enzymes. So far, studies have been conducted primarily with thioredoxin-linked members of the stroma that function in a broad array of biosynthetic and degradatory processes. Consequently, little is known about the role of redox in regulating the growing number of enzymes found to occur in the lumen, the site of oxygen evolution in thylakoid membranes. To help fill this gap, we have studied AtFKBP13, an FKBP-type immunophilin earlier shown to interact with a redox-active protein of the lumen, and found the enzyme to contain a pair of disulfide bonds in x-ray structural studies. These disulfides, which in protein mutagenesis experiments were shown to be essential for the associated peptidyl-prolyl isomerase activity, are unique to chloroplast FKBPs and are absent in animal and yeast counterparts. Both disulfide bonds were redox-active and were reduced by thioredoxin from either chloroplast or bacterial sources in a reaction that led to loss of enzyme activity. The results suggest a previously unrecognized paradigm for redox regulation in chloroplasts in which activation by light is achieved in concert with oxygen evolution by the oxidation of sulfhydryl groups (conversion of SH to S-S). Such a mechanism, occurring in the thylakoid lumen, is in direct contrast to regulation of enzymes in the stroma, where reduction of disulfides targeted by thioredoxin (S-S converted to SH) leads to an increase in activity in the light.

Thioredoxin targets of developing wheat seeds identified by complementary proteomic approaches.

The role of thioredoxin in wheat starchy endosperm was investigated utilizing two proteomic approaches. Thioredoxin targets were isolated from total KCl-soluble extracts of endosperm and flour and separated by 2-DE following (1) reduction of the extract by the NADP/thioredoxin system and labeling the newly generated sulfhydryl (SH) groups with monobromobimane (mBBr), and, in parallel, (2) trapping covalently interacting proteins on an affinity column prepared with mutant thioredoxin h in which one of the active site cysteines was replaced by serine. The two procedures were complementary: of the total targets, one-third were observed with both procedures and one-third were unique to each. Altogether 68 potential targets were identified; almost all containing conserved cysteines. In addition to confirming known interacting proteins, we identified 40 potential thioredoxin targets not previously described in seeds. A comparison of the results obtained with young endosperm (isolated 10 days after flowering) to those with mature endosperm (isolated 36 days after flowering) revealed a unique set of proteins functional in processes characteristic of each developmental stage. Flour contained 36 thioredoxin targets, most of which have been found in the isolated developing endosperm.

Thioredoxin links redox to the regulation of fundamental processes of plant mitochondria.

Mitochondria contain thioredoxin (Trx), a regulatory disulfide protein, and an associated flavoenzyme, NADP/Trx reductase, which provide a link to NADPH in the organelle. Unlike animal and yeast counterparts, the function of Trx in plant mitochondria is largely unknown. Accordingly, we have applied recently devised proteomic approaches to identify soluble Trx-linked proteins in mitochondria isolated from photosynthetic (pea and spinach leaves) and heterotrophic (potato tubers) sources. Application of the mitochondrial extracts to mutant Trx affinity columns in conjunction with proteomics led to the identification of 50 potential Trx-linked proteins functional in 12 processes: photorespiration, citric acid cycle and associated reactions, lipid metabolism, electron transport, ATP synthesis/transformation, membrane transport, translation, protein assembly/folding, nitrogen metabolism, sulfur metabolism, hormone synthesis, and stress-related reactions. Almost all of these targets were also identified by a fluorescent gel electrophoresis procedure in which reduction by Trx can be observed directly. In some cases, the processes targeted by Trx depended on the source of the mitochondria. The results support the view that Trx acts as a sensor and enables mitochondria to adjust key reactions in accord with prevailing redox state. These and earlier findings further suggest that, by sensing redox in chloroplasts and mitochondria, Trx enables the two organelles of photosynthetic tissues to communicate by means of a network of transportable metabolites such as dihydroxyacetone phosphate, malate, and glycolate. In this way, light absorbed and processed by means of chlorophyll can be perceived and function in regulating fundamental mitochondrial processes akin to its mode of action in chloroplasts.

Proteomics uncovers proteins interacting electrostatically with thioredoxin in chloroplasts.

The ability of thioredoxin f to form an electrostatic (non-covalent) complex, earlier found with fructose-1,6-bisphosphatase, was extended to include 27 previously unrecognized proteins functional in 11 processes of chloroplasts. The proteins were identified by combining thioredoxin f affinity chromatography with proteomic analysis using tandem mass spectrometry. The results provide evidence that an association with thioredoxin enables the interacting protein to achieve an optimal conformation, so as to facilitate: (i) the transfer of reducing equivalents from the ferredoxin/ferredoxin-thioredoxin reductase complex to a target protein; (ii) in some cases, to enable the channeling of metabolite substrates; (iii) to function as a subunit in the formation of multienzyme complexes.

Unraveling thioredoxin-linked metabolic processes of cereal starchy endosperm using proteomics.

Application of a thiol-specific probe, monobromobimane, with proteomics and enzyme assays led to the identification of 23 thioredoxin targets in the starchy endosperm of mature wheat seeds (Triticum aestivum cv. Butte), almost all containing at least two conserved cysteines. The identified targets, 12 not known to be thioredoxin-linked, function in a spectrum of processes: metabolism (12 targets), protein storage (three), oxidative stress (three), protein degradation (two), protein assembly/folding (one) and unknown reactions (two). In addition to formulating metabolic pathways functional in the endosperm, the results suggest that thioredoxin acts in redox regulation throughout the life cycle of the seed.

Proteomics gives insight into the regulatory function of chloroplast thioredoxins.

Thioredoxins are small multifunctional redox active proteins widely if not universally distributed among living organisms. In chloroplasts, two types of thioredoxins (f and m) coexist and play central roles in regulating enzyme activity. Reduction of thioredoxins in chloroplasts is catalyzed by an iron-sulfur disulfide enzyme, ferredoxin-thioredoxin reductase, that receives photosynthetic electrons from ferredoxin, thereby providing a link between light and enzyme activity. Chloroplast thioredoxins function in the regulation of the Calvin cycle and associated processes. However, the relatively small number of known thioredoxin-linked proteins (about 16) raised the possibility that others remain to be identified. To pursue this opportunity, we have mutated thioredoxins f and m, such that the buried cysteine of the active disulfide has been replaced by serine or alanine, and bound them to affinity columns to trap target proteins of chloroplast stroma. The covalently linked proteins were eluted with DTT, separated on gels, and identified by mass spectrometry. This approach led to the identification of 15 potential targets that function in 10 chloroplast processes not known to be thioredoxin linked. Included are proteins that seem to function in plastid-to-nucleus signaling and in a previously unrecognized type of oxidative regulation. Approximately two-thirds of these targets contained conserved cysteines. We also identified 11 previously unknown and 9 confirmed target proteins that are members of pathways known to be regulated by thioredoxin. In contrast to results with individual enzyme assays, specificity for thioredoxin f or m was not observed on affinity chromatography.

Yet another plant thioredoxin.

Thioredoxins are widely distributed proteins that function in a broad spectrum of cellular reactions. Plant cells have well characterized chloroplast and cytosolic thioredoxin systems, but, unlike animals and yeast, a mitochondrial counterpart has not been clearly defined. Recently, a complete thioredoxin system has been described in plant mitochondria, opening a new door for the study of thioredoxins as well as mitochondria.