Apnoeic oxygenation during anaesthesia has traditionally been limited by the rapid increase in carbon dioxide and subsequent decrease in pH. Using a Transnasal Humidified Rapid-Insufflation ...Ventilatory Exchange (THRIVE) technique a slower increase in carbon dioxide than earlier studies was seen. Notably, apnoeic oxygenation using THRIVE has not been systematically evaluated with arterial blood gases or in patients undergoing laryngeal surgery. The primary aim of this study was to characterize changes in arterial PO2, PCO2 and pH during apnoeic oxygenation using THRIVE under general anaesthesia.
Adult patients, (ASA I-II), undergoing shorter laryngeal surgery under general anaesthesia, were oxygenated during apnoea using THRIVE, 100% oxygen, 40–70 litres min−1. A cohort was randomized to hyperventilate during pre-oxygenation. Vital parameters and blood gases were monitored.
Thirty-one patients, age 51 (34–76) yr, BMI 25 (4) were included. Mean apnoea time was 22.5 (4.5) min. Patients were well oxygenated, SpO2 was never below 91%. The increase in PaCO2 and end-tidal CO2 during apnoea was 0.24 (0.05) and 0.12 (0.04) kPa min−1, respectively. Hyperventilation during pre-oxygenation generated no difference in PaCO2 at the end of apnoea compared with normoventilation.
This physiological study of apnoeic oxygenation using THRIVE during laryngeal surgery shows that this technique is able to keep patients with mild systemic disease and a BMI <30 well oxygenated for a period of up to 30 min. The THRIVE concept makes it possible to extend the apnoeic window but monitoring of CO2 and/or pH is recommended.
NCT02706431.
Mammalian mitochondrial DNA (mtDNA) encodes 13 proteins that are essential for the function of the oxidative phosphorylation system, which is composed of four respiratory-chain complexes and ...adenosine triphosphate (ATP) synthase. Remarkably, the maintenance and expression of mtDNA depend on the mitochondrial import of hundreds of nuclear-encoded proteins that control genome maintenance, replication, transcription, RNA maturation, and mitochondrial translation. The importance of this complex regulatory system is underscored by the identification of numerous mutations of nuclear genes that impair mtDNA maintenance and expression at different levels, causing human mitochondrial diseases with pleiotropic clinical manifestations. The basic scientific understanding of the mechanisms controlling mtDNA function has progressed considerably during the past few years, thanks to advances in biochemistry, genetics, and structural biology. The challenges for the future will be to understand how mtDNA maintenance and expression are regulated and to what extent direct intramitochondrial cross talk between different processes, such as transcription and translation, is important.
An increased prevalence of asthma/recurrent wheeze (RW), clinical allergy and allergic sensitisation up to age 13 years has previously been reported in subjects hospitalised with respiratory ...syncytial virus (RSV) bronchiolitis in their first year of life compared with matched controls. A study was undertaken to examine whether these features persist into early adulthood, to report longitudinal wheeze and allergy patterns, and to see how large and small airway function relates to RSV infection and asthma.
Follow-up at age 18 years was performed in 46 of 47 subjects with RSV and 92 of 93 controls. Assessments included questionnaire, clinical examination, skin prick tests, serum IgE antibodies to inhaled allergens, blood eosinophils, fraction of exhaled nitric oxide (FeNO), spirometry, multiple breath washout (lung clearance index, LCI) and dry air hyperventilation challenge.
Increased prevalence of asthma/RW (39% vs 9%), clinical allergy (43% vs 17%) and sensitisation to perennial allergens (41% vs 14%) were present at age 18 in the RSV cohort compared with controls. Persistent/relapsing wheeze associated with early allergic sensitisation predominated in the RSV cohort compared with controls (30% vs 1%). Spirometric function was reduced in subjects with RSV with or without current asthma, but not in asthmatic controls. LCI was linked only to current asthma, airway hyperresponsiveness and FeNO.
Severe early RSV bronchiolitis is associated with an increased prevalence of allergic asthma persisting into early adulthood. Small airway dysfunction (LCI) is related to current asthma and airway inflammation but not to RSV bronchiolitis. Reduced spirometry after RSV may reflect airway remodelling.
Mitochondrial DNA (mtDNA) encodes for proteins required for oxidative phosphorylation, and mutations affecting the genome have been linked to a number of diseases as well as the natural ageing ...process in mammals. Human mtDNA is replicated by a molecular machinery that is distinct from the nuclear replisome, but there is still no consensus on the exact mode of mtDNA replication. We here demonstrate that the mitochondrial single-stranded DNA binding protein (mtSSB) directs origin specific initiation of mtDNA replication. MtSSB covers the parental heavy strand, which is displaced during mtDNA replication. MtSSB blocks primer synthesis on the displaced strand and restricts initiation of light-strand mtDNA synthesis to the specific origin of light-strand DNA synthesis (OriL). The in vivo occupancy profile of mtSSB displays a distinct pattern, with the highest levels of mtSSB close to the mitochondrial control region and with a gradual decline towards OriL. The pattern correlates with the replication products expected for the strand displacement mode of mtDNA synthesis, lending strong in vivo support for this debated model for mitochondrial DNA replication.
Lateral resolution that exceeds the classical diffraction limit by a factor of two is achieved by using spatially structured illumination in a wide‐field fluorescence microscope. The sample is ...illuminated with a series of excitation light patterns, which cause normally inaccessible high‐resolution information to be encoded into the observed image. The recorded images are linearly processed to extract the new information and produce a reconstruction with twice the normal resolution. Unlike confocal microscopy, the resolution improvement is achieved with no need to discard any of the emission light. The method produces images of strikingly increased clarity compared to both conventional and confocal microscopes.
It is important to detect children with Early Symptomatic Syndromes Eliciting Neurodevelopmental Clinical Examinations (ESSENCE) in order to implement early intervention and support for the child and ...family. Standardized instruments for assessment in different contexts of behaviour problems, engagement and psychosocial health obtain an objective picture of the preschool child's mental health.
To explore and compare parents', preschool teachers' and child health care psychologists' assessment of behaviour, everyday function, engagement, social interaction and psychosocial health in children with ESSENCE symptoms.
Parents of 152 children (114 boys and 38 girls, 4.5 ± 1 years) with ESSENCE symptoms, 155 preschool teachers and 8 child psychologists participated. Parents and preschool teachers assessed externalizing and internalizing behavioural problems using the Strengths and Difficulties Questionnaire (SDQ), including the SDQ supplement for assessing the impact of behavioral problems on daily function. Preschool teachers also assessed engagement and social interaction using the Children's Engagement Questionnaire (CEQ), and the child psychologists assessed psychosocial health with the Child Psychosocial Health Assessment (LillaLAPS) and template in conversations with parents of children with neurodevelopmental problems.
Parents', preschool teachers' and child psychologists' assessment of the child's ESSENCE symptoms overall agreed. Both parents and preschool teachers see a strength in the child's social abilities. Differences in mean values show that parents assess more conduct, emotional symptoms and problems in daily life and more social skills, compared to the preschool teachers rating more peer problems.
It is important to consider different contexts to identify the child's need for support in everyday life. Expanded use of validated screening instruments in clinical practice would promote detection of children not already identified as exhibiting neurodevelopmental problems.
Deletions and duplications in mitochondrial DNA (mtDNA) cause mitochondrial disease and accumulate in conditions such as cancer and age-related disorders, but validated high-throughput methodology ...that can readily detect and discriminate between these two types of events is lacking. Here we establish a computational method, MitoSAlt, for accurate identification, quantification and visualization of mtDNA deletions and duplications from genomic sequencing data. Our method was tested on simulated sequencing reads and human patient samples with single deletions and duplications to verify its accuracy. Application to mouse models of mtDNA maintenance disease demonstrated the ability to detect deletions and duplications even at low levels of heteroplasmy.
Summary
Structured‐illumination microscopy allows widefield fluorescence imaging with resolution beyond the classical diffraction limit. Its linear form extends resolution by a factor of two, and its ...nonlinear form by an in‐principle infinite factor, the effective resolution in practice being determined by noise. In this paper, we analyse the noise properties and achievable resolution of linear and nonlinear 1D and 2D patterned SIM from a frequency–space perspective. We develop an analytical theory for a general case of linear or nonlinear fluorescent imaging, and verify the analytical calculations with numerical simulation for a special case where nonlinearity is produced by photoswitching of fluorescent labels. We compare the performance of two alternative implementations, using either two‐dimensional (2D) illumination patterns or sequentially rotated one‐dimensional (ID) patterns. We show that 1D patterns are advantageous in the linear case, and that in the nonlinear case 2D patterns provide a slight signal‐to‐noise advantage under idealised conditions, but perform worse than 1D patterns in the presence of nonswitchable fluorescent background.
Lay Description
Structured‐illumination microscopy (SIM) is a high‐resolution light microscopy technique that allows imaging of fluorescence at a resolution about twice the classical diffraction limit. There are various ways that the illumination can be structured, but it is not obvious how the choice of illumination pattern affects the final image quality, especially in view of the noise. We present a detailed performance analysis considering two illumination techniques: sequential illumination with line‐gratings that are shifted and rotated during image acquisition and two‐dimensional (2D) illumination structures requiring only shift operations. Our analysis is based on analytical theory, supported by simulations of images considering noise. We also extend our analysis to a nonlinear variant of SIM, with which enhanced resolution can be achieved, limited only by noise. This includes nonlinear SIM based on the light‐induced switching of the fluorescent molecules between a bright and a dark state. We find sequential illumination with line‐gratings to be advantageous in ordinary (linear) SIM, whereas 2D patterns provides a slight signal‐to‐noise advantage under idealised conditions in nonlinear SIM if there is no nonswitching background.
The mitochondrial transcription initiation machinery in humans consists of three proteins: the RNA polymerase (POLRMT) and two accessory factors, transcription factors A and B2 (TFAM and TFB2M, ...respectively). This machinery is required for the expression of mitochondrial DNA and the biogenesis of the oxidative phosphorylation system. Previous experiments suggested that TFB2M is required for promoter melting, but conclusive experimental proof for this effect has not been presented. Moreover, the role of TFB2M in promoter unwinding has not been discriminated from that of TFAM. Here we used potassium permanganate footprinting, DNase I footprinting, and in vitro transcription from the mitochondrial light-strand promoter to study the role of TFB2M in transcription initiation. We demonstrate that a complex composed of TFAM and POLRMT was readily formed at the promoter but alone was insufficient for promoter melting, which only occurred when TFB2M joined the complex. We also show that mismatch bubble templates could circumvent the requirement of TFB2M, but TFAM was still required for efficient initiation. Our findings support a model in which TFAM first recruits POLRMT to the promoter, followed by TFB2M binding and induction of promoter melting.
Dabigatran is an oral, reversible thrombin inhibitor that has shown promising results in large clinical trials. Laboratory monitoring is not needed but the effects on common coagulation assays are ...incompletely known. Dabigatran was added to plasma from healthy subjects in the concentration range 0-1,000 μg/l and analysed using several reagents for activated thromboplastin time (APTT), prothrombin time (PT), fibrinogen, antithrombin, and activated protein C resistance. Typical trough concentrations are about 50 μg/l, peak concentrations 100-300 μg/l. At 100 μg/l all APTT-results were prolonged. The concentration required to double APTT ranged between 227 and 286 μg/l, the responses for all five reagents were similar. PT-reagents were much less affected with almost no samples above INR 1.2 at 100 μg/l. The effect was sample dilution dependent with PT Quick type more sensitive than PT Owren type methods. If a patient on dabigatran has prolonged APTT, >90 seconds, and Quick PT INR>2 or Owren PT INR>1.5 over-dosing or accumulation of dabigatran should be considered. Two of four fibrinogen reagents underestimated the fibrinogen concentration considerably at expected peak concentration. Methods based on inhibition of thrombin over-estimated the antithrombin concentration, but not Xa-based. The APC-resistance methods over-estimated the APC-ratio, which may lead to miss-classification of factor V Leiden patients as being normal. Different coagulation assays, and even different reagents within an assay group, display variable effects at therapeutic concentrations of dabigatran. Some of these assay variations are of clinical importance, thus knowledge is needed for a correct interpretation of results.
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