Development of examination objectives for the Korean paramedic and emergency medical technician examination: a survey study.

PURPOSE: The duties of paramedics and emergency medical technicians (P&EMTs) are continuously changing due to developments in medical systems. This study presents evaluation goals for P&EMTs by analyzing their work, especially the tasks that new P&EMTs (with less than 3 years' experience) find difficult, to foster the training of P&EMTs who could adapt to emergency situations after graduation. METHODS: A questionnaire was created based on prior job analyses of P&EMTs. The survey questions were reviewed through focus group interviews, from which 253 task elements were derived. A survey was conducted from July 10, 2023 to October 13, 2023 on the frequency, importance, and difficulty of the 6 occupations in which P&EMTs were employed. RESULTS: The P&EMTs' most common tasks involved obtaining patients' medical histories and measuring vital signs, whereas the most important task was cardiopulmonary resuscitation (CPR). The task elements that the P&EMTs found most difficult were newborn delivery and infant CPR. New paramedics reported that treating patients with fractures, poisoning, and childhood fever was difficult, while new EMTs reported that they had difficulty keeping diaries, managing ambulances, and controlling infection. CONCLUSION: Communication was the most important item for P&EMTs, whereas CPR was the most important skill. It is important for P&EMTs to have knowledge of all tasks; however, they also need to master frequently performed tasks and those that pose difficulties in the field. By deriving goals for evaluating P&EMTs, changes could be made to their education, thereby making it possible to train more capable P&EMTs.

Toxicity of diclofenac sodium salt in Yucatan minipigs (Sus scrofa) following 4 weeks of daily intramuscular administration.

Diclofenac sodium salt (DSS) is a widely used nonsteroidal anti-inflammatory drug. The present study was performed under good laboratory practice (GLP) regulations to investigate the toxicity of DSS after 4 weeks of repeated intramuscular administration at doses of 0, 2, 10, or 20 mg/kg/day in 32 minipigs and to evaluate the DSS effect following a 2-week recovery period. Dose-related clinical signs and alterations of hematological or clinical chemistry parameters, organ weight, and macroscopic as well as histopathological findings in hepatic, renal, gastrointestinal, skin and injection sites were observed in both sexes' animals of the 10 or 20 mg/kg/day group. With the exception of the skin-related findings, most symptoms showed a tendency to resolve after the 2-week recovery period. The systemic exposure (AUClast) of DSS in plasma showed similar pattern to the increase rate of the dose and similar values between males and females except for the female 20 mg/kg dose group (56 %) on Day1. The systemic exposure showed a decreasing trend in the 10 or 20 mg/kg group after 4-week of repeated administration compared to Day1. The no-observed-adverse-effect level of DSS in this study was considered to be 2 mg/kg/day in both male and female minipigs.

Detecting protein and post-translational modifications in single cells with iDentification and qUantification sEparaTion (DUET).

While technologies for measuring transcriptomes in single cells have matured, methods for measuring proteins and their post-translational modification (PTM) states in single cells are still being actively developed. Unlike nucleic acids, proteins cannot be amplified, making detection of minute quantities from single cells difficult. Here, we develop a strategy to detect targeted protein and its PTM isoforms in single cells. We barcode the proteins from single cells by tagging them with oligonucleotides, pool barcoded cells together, run bulk gel electrophoresis to separate protein and its PTM isoform and quantify their abundances by sequencing the oligonucleotides associated with each protein species. We used this strategy, iDentification and qUantification sEparaTion (DUET), to measure histone protein H2B and its monoubiquitination isoform, H2Bub, in single yeast cells. Our results revealed the heterogeneities of H2B ubiquitination levels in single cells from different cell-cycle stages, which is obscured in ensemble measurements.

Author Correction: Detecting protein and post-translational modifications in single cells with iDentification and qUantification sEparaTion (DUET).

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

An assessment of the accuracy of a novel weight estimation device for children.

BACKGROUND: We sought to validate the accuracy and assess the efficacy of a newly developed electronic weight estimation device (ie, the rolling tape) for paediatric weight estimation. METHODS: We enrolled a convenience sample of children aged <17 years presenting to our emergency department who volunteered to participate in the study. The children's heights and weights were measured, and three researchers estimated these values using the rolling tape and Broselow tape at 5 min intervals. The weight estimates of researcher 1, researcher 2 and the Broselow tape were compared with measured values, and mean percentage error (MPE), root mean square error (RMSE) and percentage of estimates within 10% of the actual measured values were calculated. For 30 randomly selected subjects, we compared the time interval from the start of the measurement to the time that orders for epinephrine, defibrillation dose and instrument size could be given in a simulated arrest scenario. RESULTS: We enrolled 906 children (median age 4.0 years). For researcher 1, researcher 2 and the Broselow tape, MPE values were 0.11% (RMSE 2.61 kg), 1.41% (RMSE, 2.61 kg) and 1.72% (RMSE 5.41 kg), respectively, and the percentages of children with predictions within 10% of their actual weight were 75.1%, 75.7% and 60.6%, respectively. In the 30 simulated cases, the mean time for measurement to ordering was significantly shorter (25.8 s vs 35.5 s, p<0.001) for the rolling tape compared with the Broselow tape method. CONCLUSIONS: The rolling tape is a good weight estimation tool for children compared with other methods. The rolling tape method significantly decreased the time from weight estimation to orders for essential drug dose, instrument size and defibrillation dose for resuscitation.

Smartphone-Based Urine Reagent Strip Test in the Emergency Department.

BACKGROUND: Although a smartphone could be used for a urine reagent strip test, few studies have reported on the reliability of the test in a clinical setting. The objective of our study was to access the smartphone-based urine reagent strip test in the clinical emergency department (ED). MATERIALS AND METHODS: We developed a smartphone-based urine reagent strip reader for a rapid and accurate screening of leukocyte esterase (LE) and nitrite (NIT) in urine. The developed reader was evaluated with the clinical urine samples (n = 81). The detection performance of the reader for LE and NIT was evaluated to assess reliability of the reader; turnaround times (TATs) for analysis and the time for the entire study procedure were also calculated to assess the efficiency of the reader. A photometric analyzer (model US-3100R Plus(®); Eiken Chemical, Ltd., Tokyo, Japan) was used as a reference. RESULTS: The proposed reader showed high accuracy (85.2% for LE and 97.5% for NIT), exhibiting close agreement with the true values (κ = 0.903 for LE; κ = 1.000 for NIT). The reader also exhibited a lower median TAT for analysis than the photometric analyzer (3.0 min versus 33.0 min; p < 0.001). This reduction of TAT in the reader was even more evident considering the required time for delivery of urine samples for the photometric analyzer (3.0 min versus 62.0 min; p < 0.001). CONCLUSIONS: Our results demonstrated the clinical capability of a smartphone-based urine reagent strip test, and this reader is expected to enable a more rapid and reliable colorimetric test for screening of LE and NIT at the clinical setting and the point of care.

Molecular features distinguish ten neuronal types in the mouse superficial superior colliculus.

The superior colliculus (SC) is a midbrain center involved in controlling head and eye movements in response to inputs from multiple sensory modalities. Visual inputs arise from both the retina and visual cortex and converge onto the superficial layer of the SC (sSC). Neurons in the sSC send information to deeper layers of the SC and to thalamic nuclei that modulate visually guided behaviors. Presently, our understanding of sSC neurons is impeded by a lack of molecular markers that define specific cell types. To better understand the identity and organization of sSC neurons, we took a systematic approach to investigate gene expression within four molecular families: transcription factors, cell adhesion molecules, neuropeptides, and calcium binding proteins. Our analysis revealed 12 molecules with distinct expression patterns in mouse sSC: cadherin 7, contactin 3, netrin G2, cadherin 6, protocadherin 20, retinoid-related orphan receptor ß, brain-specific homeobox/POU domain protein 3b, Ets variant gene 1, substance P, somatostatin, vasoactive intestinal polypeptide, and parvalbumin. Double labeling experiments, by either in situ hybridization or immunostaining, demonstrated that the 12 molecular markers collectively define 10 different sSC neuronal types. The characteristic positions of these cell types divide the sSC into four distinct layers. The 12 markers identified here will serve as valuable tools to examine molecular mechanisms that regulate development of sSC neuronal types. These markers could also be used to examine the connections between specific cell types that form retinocollicular, corticocollicular, or colliculothalamic pathways. J. Comp. Neurol. 524:2300-2321, 2016. © 2016 Wiley Periodicals, Inc.

Enzymatic synthesis of 3-O-α-maltosyl-l-ascorbate using an engineered cyclodextrin glucanotransferase.

A mutant derived from a cyclodextrin glucantransferase with an alanine residue as its acid/base catalyst residue (CGT-E284A) catalyzed regioselective glycosylation at 3-OH of l-ascorbic acid using α-maltosyl fluoride (αG2F) and l-ascorbic acid as the donor and acceptor, respectively, yielding 3-O-α-maltosyl-l-ascorbate (AA3αG2). The optimum conditions were determined by high-performance liquid chromatography analysis with 20mM αG2F and 40mM l-ascorbic acid as the substrates at pH 7.5 and 25°C with 1mg/ml of the enzyme for 24h. Calcium ions bound in CGT-E284A played an important role in the transglycosylation. CGT-E284A exhibited typical saturation kinetic behaviour for αG2F at a fixed acceptor concentration (40mM), and substrate inhibition by l-ascorbic acid was observed at high l-ascorbic acid concentrations (>60mM). AA3αG2 was isolated from a preparative scale reaction with a yield of 29%, and it showed extremely high stability under oxidative conditions.

Transglycosylation of engineered cyclodextrin glucanotransferases as O-glycoligases.

An O-glycoligase is a hydrolytically impaired mutant of a retaining α-glycosidase in which the catalytic acid/base has been removed, but which can still perform transglycosylation when incubated with activated glycosyl fluoride donor sugars. In this paper, we describe another example, wherein a cyclodextrin glucanotransferase mutant (CGT-E284A) with an alanine residue at its general acid/base catalyst position (Glu284), was constructed. This mutant was hydrolytically inactive, but exhibited significant transglycosylation activity using α-maltosyl fluoride (αG2F) as donor, and either 4-nitrophenyl glucosides or maltosides as acceptors. To improve transglycosylation activity, a site-saturation mutagenesis library at Glu284 was created. Through a thin-layer chromatography-based screening process, two mutants were identified; (1) a mutant with a glycine residue at Glu284 (CGT-E284G) exhibiting improved transglycosylation activity compared with the original alanine mutant and (2) a mutant with a serine residue with residual hydrolytic activity. Kinetic analysis revealed that 4-nitrophenyl maltosides were better acceptors than 4-nitrophenyl glucosides. Transglycosylation activities of CGT-E284A and CGT-E284G were inhibited at high concentrations (>0.8 mM) of the acceptor sugars. In contrast, typical saturation kinetic behavior was observed upon varying the donor (αG2F) concentration at a fixed acceptor concentration (0.8 mM). The catalytic efficiencies (apparent k(cat)/K(M)) of CGT-E284G were generally three- to sixfold higher than those of CGT-E284A. Due to the rate at high concentrations of the acceptors, higher transglycosylation yields were achieved at a low concentration of the acceptors (69-84% at 1 mM) compared to those at a higher concentration (22-36% at 10mM).

Structural and biochemical characterization of the broad substrate specificity of Bacteroides thetaiotaomicron commensal sialidase.

Sialidases release the terminal sialic acid residue from a wide range of sialic acid-containing polysaccharides. Bacteroides thetaiotaomicron, a symbiotic commensal microbe, resides in and dominates the human intestinal tract. We characterized the recombinant sialidase from B. thetaiotaomicron (BTSA) and demonstrated that it has broad substrate specificity with a relative activity of 97, 100 and 64 for 2,3-, 2,6- and 2,8-linked sialic substrates, respectively. The hydrolysis activity of BTSA was inhibited by a transition state analogue, 2-deoxy-2,3-dehydro-N-acetyl neuraminic acid, by competitive inhibition with a Ki value of 35µM. The structure of BSTA was determined at a resolution of 2.3Å. This structure exhibited a unique carbohydrate-binding domain (CBM) at its N-terminus (a.a. 23-190) that is adjacent to the catalytic domain (a.a. 191-535). The catalytic domain has a conserved arginine triad with a wide-open entrance for the substrate that exposes the catalytic residue to the surface. Unlike other pathogenic sialidases, the polysaccharide-binding site in the CBM is near the active site and possibly holds and positions the polysaccharide substrate directly at the active site. The structural feature of a wide substrate-binding groove and closer proximity of the polysaccharide-binding site to the active site could be a unique signature of the commensal sialidase BTSA and provide a molecular basis for its pharmaceutical application.

The monoterpenoids citral and geraniol are moderate inhibitors of CYP2B6 hydroxylase activity.

Monoterpenes are found in the volatile essence of flowers, plants oils, and herbal medicines. Some are commonly used as food additives and fragrance components, and many are found in cosmetics, soaps, cleaning products, disinfectants, preservatives, and medicines. We have recently discovered a moderate inhibitory effect of borneol and isoborneol toward CYP2B6-catalyzed bupropion hydroxylase activity. Based on that result, we expanded our study to evaluate the inhibitory effects of 22 monoterpenoids on CYP2B6 activity in vitro. Among the monoterpenoids screened, borneol, camphor, cineole, isoborneol, menthol, and perillaldehyde showed slight inhibition of CYP2B6-catalyzed bupropion hydroxylation, displaying greater than 50% inhibition at 50muM. Citral and geraniol strongly inhibited CYP2B6 hydroxylase activity in a competitive manner, with K(i) values of 6.8 and 10.3muM, respectively, which are higher than the K(i) (1.8muM) of the well-known CYP2B6-selective inhibitor thio-TEPA. These in vitro data indicate that high amounts of these two monoterpenoids might interact with drugs that are metabolized by CYP2B6. The in vivo pharmacokinetics of these compounds should be examined to determine whether the inhibition of CYP2B6 activity by monoterpenoids has clinical relevance.

The contributions of cytochromes P450 3A4 and 3A5 to the metabolism of the phosphodiesterase type 5 inhibitors sildenafil, udenafil, and vardenafil.

The role of the genetically polymorphic CYP3A5 in the metabolism of CYP3A substrates is unclear. We investigated the contributions of the CYP3A4 and CYP3A5 isoforms to the metabolism of the phosphodiesterase type 5 inhibitors (PDE5Is) sildenafil, udenafil, and vardenafil. In vitro incubation studies of sildenafil N-demethylation, udenafil N-dealkylation, and vardenafil N-deethylation were conducted using recombinant CYP3A enzymes and 15 human liver microsome (HLM) preparations with predetermined CYP3A5 genotypes. Recombinant CYP3A4 and CYP3A5 both produced N-desalkyl metabolites of sildenafil, udenafil, and vardenafil. The catalytic efficiency (Cl(int) = V(max)/apparent K(m)) of the rCYP3A5 isoform for vardenafil N-deethylation was about 3.2-fold that of rCYP3A4, whereas the intrinsic clearance rates for N-dealkylation of both sildenafil and udenafil were similar between rCYP3A5 and rCYP3A4. The metabolite formation activity was higher in HLMs heterozygous for the CYP3A5*3 allele (n = 9) than in HLMs homozygous for CYP3A5*3 (n = 6). These findings suggest that CYP3A5 and CYP3A4 play a significant role in the metabolism of PDE5Is. The genetic polymorphism of CYP3A5 may contribute to interindividual variability in the disposition of PDE5Is, especially vardenafil. Further in vivo studies are needed to confirm the effects of CYP3A5 genotypes on the pharmacokinetics of PDE5Is.