Analysis of pancreas tissue in a child positive for islet cell antibodies

Conclusions/interpretation These observations suggest that positivity for ICA alone, even when lasting for more than 1 year, is not associated with inflammatory changes in the islets. However, it is most likely that the pancreatic islets were infected by an enterovirus in this child

Validation of the Munich Actimetry Sleep Detection Algorithm for estimating sleep-wake patterns from activity recordings

© 2021 The Authors. Journal of Sleep Research published by John Wiley & Sons Ltd on behalf of European Sleep Research Society. This is an open access article under the terms of the Creative Commons Attribution‐NonCommercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.Periods of sleep and wakefulness can be estimated from wrist-locomotor activity recordings via algorithms that identify periods of relative activity and inactivity. Here, we evaluated the performance of our Munich Actimetry Sleep Detection Algorithm. The Munich Actimetry Sleep Detection Algorithm uses a moving 24-h threshold and correlation procedure estimating relatively consolidated periods of sleep and wake. The Munich Actimetry Sleep Detection Algorithm was validated against sleep logs and polysomnography. Sleep-log validation was performed on two field samples collected over 54 and 34 days (median) in 34 adolescents and 28 young adults. Polysomnographic validation was performed on a clinical sample of 23 individuals undergoing one night of polysomnography. Epoch-by-epoch analyses were conducted and comparisons of sleep measures carried out via Bland-Altman plots and correlations. Compared with sleep logs, the Munich Actimetry Sleep Detection Algorithm classified sleep with a median sensitivity of 80% (interquartile range [IQR] = 75%-86%) and specificity of 91% (87%-92%). Mean onset and offset times were highly correlated (r = .86-.91). Compared with polysomnography, the Munich Actimetry Sleep Detection Algorithm reached a median sensitivity of 92% (85%-100%) but low specificity of 33% (10%-98%), owing to the low frequency of wake episodes in the night-time polysomnographic recordings. The Munich Actimetry Sleep Detection Algorithm overestimated sleep onset (~21 min) and underestimated wake after sleep onset (~26 min), while not performing systematically differently from polysomnography in other sleep parameters. These results demonstrate the validity of the Munich Actimetry Sleep Detection Algorithm in faithfully estimating sleep-wake patterns in field studies. With its good performance across daytime and night-time, it enables analyses of sleep-wake patterns in long recordings performed to assess circadian and sleep regularity and is therefore an excellent objective alternative to sleep logs in field settings.ASL received a stipend from the Max‐Weber‐Programm (Studienstiftung), AMB received funding from the Graduate School of Systemic Neurosciences Munich, CR received funding from the Fundação para a Ciência e Tecnologia (FCT) PhD research grants (PDE/BDE/114584/2016), LKP received a fellowship from the Coordenação de Aperfeiçoamento Pessoal de Nível Superior (CAPES, Finance Code 001), and NG received research funding from the FoeFoLe program at LMU (registration No. 37/2013).info:eu-repo/semantics/publishedVersio

Stringency of the 2-His–1-Asp Active-Site Motif in Prolyl 4-Hydroxylase

The non-heme iron(II) dioxygenase family of enzymes contain a common 2-His–1-carboxylate iron-binding motif. These enzymes catalyze a wide variety of oxidative reactions, such as the hydroxylation of aliphatic C–H bonds. Prolyl 4-hydroxylase (P4H) is an α-ketoglutarate-dependent iron(II) dioxygenase that catalyzes the post-translational hydroxylation of proline residues in protocollagen strands, stabilizing the ensuing triple helix. Human P4H residues His412, Asp414, and His483 have been identified as an iron-coordinating 2-His–1-carboxylate motif. Enzymes that catalyze oxidative halogenation do so by a mechanism similar to that of P4H. These halogenases retain the active-site histidine residues, but the carboxylate ligand is replaced with a halide ion. We replaced Asp414 of P4H with alanine (to mimic the active site of a halogenase) and with glycine. These substitutions do not, however, convert P4H into a halogenase. Moreover, the hydroxylase activity of D414A P4H cannot be rescued with small molecules. In addition, rearranging the two His and one Asp residues in the active site eliminates hydroxylase activity. Our results demonstrate a high stringency for the iron-binding residues in the P4H active site. We conclude that P4H, which catalyzes an especially demanding chemical transformation, is recalcitrant to change

General practitioners' experience and benefits from patient evaluations

<p>Abstract</p> <p>Background</p> <p>It has now for many years been recognised that patient evaluations should be undertaken as an integral part of the complex task of improving the quality of general practice care. Yet little is known about the general practitioners' (GPs') benefit from patient evaluations. Aim 1 was to study the impact on the GPs of a patient evaluation and subsequent feedback of results presented at a plenary session comprising a study guide for the results and group discussions. Aim 2 was to study possible facilitators and barriers to the implementations of the results raised by the patient evaluation process.</p> <p>Methods</p> <p>A patient evaluation survey of 597 voluntarily participating GPs was performed by means of the EUROPEP questionnaire. Evaluation results were fed back to the GPs as written reports at a single feedback meeting with group discussions of the results. Between 3 and 17 months after the feedback, the 597 GPs received a questionnaire with items addressing their experience with and perceived benefit from the evaluations.</p> <p>Results</p> <p>79.4% of the GPs responded. 33% of the responding GPs reported that the patient evaluation had raised their attention to the patient perspective on the quality of general practice care. Job satisfaction had improved among 26%, and 21% had developed a more positive attitude to patient evaluations. 77% of the GPs reported having learnt from the evaluation. 54% had made changes to improve practice, 82% would recommend a patient evaluation to a colleague and 75% would do another patient evaluation if invited. 14% of the GPs had become less positive towards patient evaluations, and job satisfaction had decreased among 3%.</p> <p>Conclusions</p> <p>We found a significant impact on the GPs regarding satisfaction with the process and attitude towards patient evaluations, GPs' attention to the patients' perspective on care quality and their job satisfaction. Being benchmarked against the average seemed to raise barriers to the concept of patient evaluations and difficulties interpreting the results may have formed a barrier to their implementation which was partly overcome by adding qualitative data to the quantitative results. The GPs' significant willingness to share and discuss the results with others may have served as a facilitator.</p

Evidence of disturbed sleep and mood state in well-trained athletes during short-term intensified training with and without a high carbohydrate nutritional intervention

This is an Accepted Manuscript of an article published by Taylor & Francis in Journal of Sports Sciences on 25th September 2015, available online: http://www.tandfonline.com/10.1080/02640414.2015.1085589.Few studies have investigated the effects of exercise training on sleep physiology in well-trained athletes. We investigated changes in sleep markers, mood state and exercise performance in well-trained cyclists undergoing short-term intensified training and carbohydrate nutritional intervention. Thirteen highly-trained male cyclists (age: 25 ± 6y, (Formula presented.)O2max: 72 ± 5 ml/kg/min) participated in two 9-day periods of intensified training while undergoing a high (HCHO) or moderate (CON) carbohydrate nutritional intervention before, during and after training sessions. Sleep was measured each night via wristwatch actigraphy. Mood state questionnaires were completed daily. Performance was assessed with maximal oxygen uptake ((Formula presented.). Percentage sleep time fell during intensified training (87.9 ± 1.5 to 82.5 ± 2.3%; p < 0.05) despite an increase in time in bed (456 ± 50 to 509 ± 48 min; p = 0.02). Sleep efficiency decreased during intensified training (83.1 ± 5.3 to 77.8 ± 8.6%; p < 0.05). Actual sleep time was significantly higher in CON than HCHO throughout intensified training. Mood disturbance increased during intensified training and was higher in CON than HCHO (p < 0.05). Performance in the (Formula presented.) exercise protocol fell significantly with intensified training. The main findings of this study were that 9-days of intensified training in highly-trained cyclists resulted in significant and progressive declines in sleep quality, mood state and maximal exercise performance

Eco-physiological responses of copepods and pteropods to ocean warming and acidification

We compare physiological responses of the crustacean copepod Calanus pacificus and pelagic pteropod mollusk Limacina helicina to ocean temperatures and pH by measuring biomarkers of oxidative stress, antioxidant defences, and the activity of the respiratory electron transport system in organisms collected on the 2016 West Coast Ocean Acidification cruise in the California Current System. Copepods and pteropods exhibited strong but divergent responses in the same habitat; copepods had higher oxygen-reactive absorbance capacity, glutathione-S-transferase, and total glutathione content. The ratio between reduced to oxidised glutathione was higher in copepods than in pteropods, indicating lower oxidative stress in copepods. Pteropods showed higher activities of glutathione reductase, catalase, and lipid peroxidation, indicating increased antioxidant defences and oxidative stress. Thus, the antioxidant defence system of the copepods has a greater capacity to respond to oxidative stress, while pteropods already face severe stress and show limited capacity to deal with further changes. The results suggest that copepods have higher adaptive potential, owing to their stronger vertical migration behaviour and efficient glutathione metabolism, whereas pteropods run the risk of oxidative stress and mortality under high CO2 conditions. Our results provide a unique dataset and evidence of stress-inducing mechanisms behind pteropod ocean acidification responses.</p

The Structural Features of Trask That Mediate Its Anti-Adhesive Functions

Trask/CDCP1 is a transmembrane protein with a large extracellular and small intracellular domains. The intracellular domain (ICD) undergoes tyrosine phosphorylation by Src kinases during anchorage loss and, when phosphorylated, Trask functions to inhibit cell adhesion. The extracellular domain (ECD) undergoes proteolytic cleavage by serine proteases, although the functional significance of this remains unknown. There is conflicting evidence regarding whether it functions to signal the phosphorylation of the ICD. To better define the structural determinants that mediate the anti-adhesive functions of Trask, we generated a series of deletion mutants of Trask and expressed them in tet-inducible cell models to define the structural elements involved in cell adhesion signaling. We find that the ECD is dispensable for the phosphorylation of the ICD or for the inhibition of cell adhesion. The anti-adhesive functions of Trask are entirely embodied within its ICD and are specifically due to tyrosine phosphorylation of the ICD as this function is completely lost in a phosphorylation-defective tyrosine-phenylalanine mutant. Both full length and cleaved ECDs are fully capable of phosphorylation and undergo phosphorylation during anchorage loss and cleavage is not an upstream signal for ICD phosphorylation. These data establish that the anti-adhesive functions of Trask are mediated entirely through its tyrosine phosphorylation. It remains to be defined what role, if any, the Trask ECD plays in its adhesion functions

Attachment and Entry of Chlamydia Have Distinct Requirements for Host Protein Disulfide Isomerase

Chlamydia is an obligate intracellular pathogen that causes a wide range of diseases in humans. Attachment and entry are key processes in infectivity and subsequent pathogenesis of Chlamydia, yet the mechanisms governing these interactions are unknown. It was recently shown that a cell line, CHO6, that is resistant to attachment, and thus infectivity, of multiple Chlamydia species has a defect in protein disulfide isomerase (PDI) N–terminal signal sequence processing. Ectopic expression of PDI in CHO6 cells led to restoration of Chlamydia attachment and infectivity; however, the mechanism leading to this recovery was not ascertained. To advance our understanding of the role of PDI in Chlamydia infection, we used RNA interference to establish that cellular PDI is essential for bacterial attachment to cells, making PDI the only host protein identified as necessary for attachment of multiple species of Chlamydia. Genetic complementation and PDI-specific inhibitors were used to determine that cell surface PDI enzymatic activity is required for bacterial entry into cells, but enzymatic function was not required for bacterial attachment. We further determined that it is a PDI-mediated reduction at the cell surface that triggers bacterial uptake. While PDI is necessary for Chlamydia attachment to cells, the bacteria do not appear to utilize plasma membrane–associated PDI as a receptor, suggesting that Chlamydia binds a cell surface protein that requires structural association with PDI. Our findings demonstrate that PDI has two essential and independent roles in the process of chlamydial infectivity: it is structurally required for chlamydial attachment, and the thiol-mediated oxido-reductive function of PDI is necessary for entry

Novel HTS Strategy Identifies TRAIL-Sensitizing Compounds Acting Specifically Through the Caspase-8 Apoptotic Axis

Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) is potentially a very important therapeutic as it shows selectivity for inducing apoptosis in cancer cells whilst normal cells are refractory. TRAIL binding to its cognate receptors, Death Receptors-4 and -5, leads to recruitment of caspase-8 and classical activation of downstream effector caspases, leading to apoptosis. As with many drugs however, TRAIL's usefulness is limited by resistance, either innate or acquired. We describe here the development of a novel 384-well high-throughput screening (HTS) strategy for identifying potential TRAIL-sensitizing agents that act solely in a caspase-8 dependent manner. By utilizing a TRAIL resistant cell line lacking caspase-8 (NB7) compared to the same cells reconstituted with the wild-type protein, or with a catalytically inactive point mutant of caspase-8, we are able to identify compounds that act specifically through the caspase-8 axis, rather than through general toxicity. In addition, false positive hits can easily be “weeded out” in this assay due to their activity in cells lacking caspase-8-inducible activity. Screening of the library of pharmacologically active compounds (LOPAC) was performed as both proof-of-concept and to discover potential unknown TRAIL sensitizers whose mechanism is caspase-8 mediated. We identified known TRAIL sensitizers from the library and identified new compounds that appear to sensitize specifically through caspase-8. In sum, we demonstrate proof-of-concept and discovery of novel compounds with a screening strategy optimized for the detection of caspase-8 pathway-specific TRAIL sensitizers. This screen was performed in the 384-well format, but could easily be further miniaturized, allows easy identification of artifactual false positives, and is highly scalable to accommodate diverse libraries