Abstract
Improvements in cost and speed of next generation sequencing (NGS) have provided a new pathway for delivering disease diagnosis, molecular typing, and detection of antimicrobial resistance ...(AMR). Numerous published methods and protocols exist, but a lack of harmonisation has hampered meaningful comparisons between results produced by different methods/protocols vital for global genomic diagnostics and surveillance. As an exemplar, this study evaluated the sensitivity and specificity of five well-established in-silico AMR detection software where the genotype results produced from running a panel of 436
Escherichia coli
were compared to their AMR phenotypes, with the latter used as gold-standard. The pipelines exploited previously known genotype–phenotype associations. No significant differences in software performance were observed. As a consequence, efforts to harmonise AMR predictions from sequence data should focus on: (1) establishing universal minimum to assess performance thresholds (e.g. a control isolate panel, minimum sensitivity/specificity thresholds); (2) standardising AMR gene identifiers in reference databases and gene nomenclature; (3) producing consistent genotype/phenotype correlations. The study also revealed limitations of in-silico technology on detecting resistance to certain antimicrobials due to lack of specific fine-tuning options in bioinformatics tool or a lack of representation of resistance mechanisms in reference databases. Lastly, we noted user friendliness of tools was also an important consideration. Therefore, our recommendations are timely for widespread standardisation of bioinformatics for genomic diagnostics and surveillance globally.
The proximal musculo‐tendinous junction (MTJ) is a common site of hamstring strain injury but the anatomy of this region is not well defined. A morphometric analysis of the proximal MTJs of biceps ...femoris long head (BFlh), semitendinosus (ST), and semimembranosus (SM) was undertaken from dissection of 10 thighs from five male cadavers and magnetic resonance imaging of 20 thighs of 10 active young men. The length, volume, and cross‐sectional area of the proximal tendon, MTJ and muscle belly, and muscle‐tendon interface area were calculated. In both groups, MTJs were reconstructed three‐dimensionally. The proximal tendons and MTJs were expansive, particularly within SM and BFlh. Morphology varied between muscles although length measurements within individual muscles were similar in cadavers and young men. Semimembranosus had the longest proximal tendon (cadavers: mean 33.6 ± 2.0 cm; young men: mean 31.7 ± 1.6 cm) and MTJ (>20 cm in both groups) and the greatest muscle‐tendon interface area, followed by BFlh and ST. Mean muscle belly volumes were more than three times greater in young men than elderly male cadavers (P < 0.001). These unique morphometric data contribute to a better understanding of hamstring anatomy, an important factor in the pathogenesis of hamstring strain injury.
Hypothermia and hypothermic preconditioning are known to be profoundly cardioprotective, but the molecular mechanisms of this protection have not been fully explained. In this study, temperature ...preconditioning (16 °C) was found to be cardioprotective in isolated adult rat ventricular myocytes, enhancing contractile recovery and preventing calcium dysregulation after oxidative stress. Hypothermic preconditioning preserved mitochondrial function by delaying the pathological opening of the mitochondrial permeability transition pore (mPTP), whereas transient mPTP flickering remained unaltered. For the first time, reactive oxygen species (ROS) from the mitochondria are shown to be released exclusively during the hypothermic episodes of the temperature-preconditioning protocol. Using a mitochondrially targeted ROS biosensor, ROS release was shown during the brief bursts to 16 °C of temperature preconditioning. The ROS scavenger N-(2-mercaptopropionyl) glycine attenuated ROS accumulation during temperature preconditioning, abolishing the protective delay in mPTP opening. Temperature preconditioning induces ROS-dependant phosphorylation of the prosurvival kinase extracellular signal-regulated kinase (ERK)1/2. ERK1/2 activation was shown to be downstream of ROS release, as the presence of a ROS scavenger during temperature preconditioning completely blocked ERK1/2 activation. The cardioprotective effects of temperature preconditioning on mPTP opening were completely lost by inhibiting ERK1/2 activation. Thus, mitochondrial ROS release and ERK1/2 activation are both necessary to signal the cardioprotective effects of temperature preconditioning in cardiac myocytes.
Raman microspectroscopy was applied to monitor the intracellular redox state of myoglobin and cytochrome c from isolated adult rat cardiomyocytes during hypoxia and reoxygenation. The nitrite ...reductase activity of myoglobin leads to the production of nitric oxide in cells under hypoxic conditions, which is linked to the inhibition of mitochondrial respiration. In this work, the subsequent reoxygenation of cells after hypoxia is shown to lead to increased levels of oxygen-bound myoglobin relative to the initial levels observed under normoxic conditions. Increased levels of reduced cytochrome c in ex vivo cells are also observed during hypoxia and reoxygenation by Raman microspectroscopy. The cellular response to reoxygenation differed dramatically depending on the method used in the preceding step to create hypoxic conditions in the cell suspension, where a chemical agent, sodium dithionite, leads to reduction of cytochromes in addition to removal of dissolved oxygen, and bubbling-N2 gas leads to displacement of dissolved oxygen only. These results have an impact on the assessment of experimental simulations of hypoxia in cells. The spectroscopic technique employed in this work will be used in the future as an analytical method to monitor the effects of varying levels of oxygen and nutrients supplied to cardiomyocytes during either the preconditioning of cells or the reperfusion of ischaemic tissue.
Background
Emergent decompression, either by closed reduction or surgical decompression, of the acutely compressed cervical spinal cord is recognized as important in minimizing the neurological ...outcomes of these injuries. The aim of this study is to optimize New Zealand's capability to perform closed reductions.
Methods
Regional hospitals throughout New Zealand were surveyed on their capability to reduce acute cervical spine dislocations using traction. A systematic literature review was performed to investigate aspects of patient selection, reduction technique and the outcomes achieved with closed reduction of such injuries. This information was combined with our local experience to develop an evidence‐based guideline.
Results
Most (12/14) of the regional centres throughout New Zealand have recent experience, remain willing and have the resources necessary to reduce appropriate cervical spine injuries using traction. Fourteen published studies from nine countries detail a 75% success rate from 363 cases of attempted closed reduction, with the greatest neurological recovery noted in patients with shorter time period from injury to reduction. One patient suffered neurological worsening. The published protocols were compared and coupled with our local practice to create an online, step‐by‐step, evidence‐based reference to help clinicians in regional hospitals perform a safe and successful closed reduction.
Conclusion
To optimize the capability of inexperienced personnel to perform closed reductions in a safe and timely manner, we have developed an online, step‐by‐step, evidence‐based reference (www.closedreduction.co.nz). This forms part of New Zealand's strategy to achieve urgent cord decompression for appropriate cervical spinal cord injuries.
'Artifacts of Thinking: Reading Arendt’s 'Denktagebuch'' offers a path through Hannah Arendt’s recently published 'Denktagebuch', or 'Book of Thoughts.' In this book a number of innovative Arendt ...scholars come together to ask how we should think about these remarkable writings in the context of Arendt’s published writing and broader political thinking. Unique in its form, the 'Denktagebuch' offers brilliant insights into Arendt’s practice of thinking and writing. 'Artifacts of Thinking' provides an introduction to the 'Denktagebuch' as well as a glimpse of these fascinating but untranslated fragments that reveal not only Arendt’s understanding of “the life of the mind” but her true lived experience of it.
The loss of intracellular potassium is a pivotal step in the induction of apoptosis but the mechanisms underlying this response are poorly understood. Here we report caspase-dependent stimulation of ...potassium channels by the Fas receptor in a human Jurkat T cell line. Receptor activation with Fas ligand for 30 min increased the amplitude of voltage-activated potassium currents 2-fold on average. This produces a sustained outward current, ∼10 pA, at physiological membrane potentials during Fas ligand-induced apoptosis. Both basal and Fas ligand-induced currents were blocked completely by toxins that selectively inhibit Kv1.3 potassium channels. Kv1.3 stimulation required the expression of Fas-associated death domain protein and activation of caspase 8, but did not require activation of caspase 3 or protein synthesis. Furthermore, Kv1.3 stimulation by Fas ligand was prevented by chronic stimulation of protein kinase C with 20 nm phorbol 12-myristate 13-acetate during Fas ligand treatment, which also blocks apoptosis. Thus, Fas ligand increases Kv1.3 channel activity through the same canonical apoptotic signaling cascade that is required for potassium efflux, cell shrinkage, and apoptosis.
Postmastectomy radiotherapy in patients with four or more positive axillary nodes reduces breast cancer mortality, but its role in patients with one to three involved nodes is controversial. We ...assessed the effects of postmastectomy radiotherapy on quality of life (QOL) in women with intermediate-risk breast cancer.
SUPREMO is an open-label, international, parallel-group, randomised, controlled trial. Women aged 18 years or older with intermediate-risk breast cancer (defined as pT1–2N1; pT3N0; or pT2N0 if also grade III or with lymphovascular invasion) who had undergone mastectomy and, if node positive, axillary surgery, were randomly assigned (1:1) to receive chest wall radiotherapy (50 Gy in 25 fractions or a radiobiologically equivalent dose of 45 Gy in 20 fractions or 40 Gy in 15 fractions) or no radiotherapy. Randomisation was done with permuted blocks of varying block length, and stratified by centre, without masking of patients or investigators. The primary endpoint is 10-year overall survival. Here, we present 2-year results of QOL (a prespecified secondary endpoint). The QOL substudy, open to all UK patients, consists of questionnaires (European Organisation for Research and Treatment of Cancer QLQ-C30 and QLQ-BR23, Body Image Scale, Hospital Anxiety and Depression Scale HADS, and EQ-5D-3L) completed before randomisation, and at 1, 2, 5, and 10 years. The prespecified primary outcomes within this QOL substudy were global QOL, fatigue, physical function, chest wall symptoms, shoulder and arm symptoms, body image, and anxiety and depression. Data were analysed by intention to treat, using repeated mixed-effects methods. This trial is registered with the ISRCTN registry, number ISRCTN61145589.
Between Aug 4, 2006, and April 29, 2013, 1688 patients were enrolled internationally and randomly assigned to receive chest wall radiotherapy (n=853) or not (n=835). 989 (79%) of 1258 patients from 111 UK centres consented to participate in the QOL substudy (487 in the radiotherapy group and 502 in the no radiotherapy group), of whom 947 (96%) returned the baseline questionnaires and were included in the analysis (radiotherapy, n=471; no radiotherapy, n=476). At up to 2 years, chest wall symptoms were worse in the radiotherapy group than in the no radiotherapy group (mean score 14·1 SD 15·8 in the radiotherapy group vs 11·6 14·6 in the no radiotherapy group; effect estimate 2·17, 95% CI 0·40–3·94; p=0·016); however, there was an improvement in both groups between years 1 and 2 (visit effect −1·34, 95% CI −2·36 to −0·31; p=0·010). No differences were seen between treatment groups in arm and shoulder symptoms, body image, fatigue, overall QOL, physical function, or anxiety or depression scores.
Postmastectomy radiotherapy led to more local (chest wall) symptoms up to 2 years postrandomisation compared with no radiotherapy, but the difference between groups was small. These data will inform shared decision making while we await survival (trial primary endpoint) results.
Medical Research Council, European Organisation for Research and Treatment of Cancer, Cancer Australia, Dutch Cancer Society, Trustees of Hong Kong and Shanghai Banking Corporation.
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