System of System Integration for Hyperspectral Imaging Microscopy

Hyperspectral imaging (HSI) has become a leading tool in the medical field due to its capabilities for providing assessments of tissue pathology and separation of fluorescence signals. Acquisition speeds have been slow due to the need to acquire signal in many spectral bands and the light losses associated with technologies of spectral filtering. Traditional methods resulted in limited signal strength which placed limitations on time sensitive and photosensitive assays. For example, the distribution of cyclic adenosine monophosphate (cAMP) is largely undetermined because current microscope technologies lack the combination of speed, resolution, and spectral ability to accurately measure Forster resonance energy transfer (FRET). The work presented in this dissertation assesses the feasibility of integrating excitation-scanning hyperspectral imaging methods in widefield and confocal microscopy as a potential solution to improving acquisition speeds without compromising sensitivity and specificity. Our laboratory has previously proposed excitation-scanning approaches to improve signal-to-noise ratio (SNR) and showed that by using excitation-scanning, most-to-all emitted light at each excitation wavelength band can be detected which in turn, increases the SNR. This dissertation describes development and early feasibility studies for two novel prototype concepts as an alternative excitation-scanning HSI technology that may xvi increase acquisition speeds without compromising sensitivity or specificity. To achieve this, two new technologies for excitation-scanning HSI were conceptually designed: - LED-based spectral illumination for widefield microscopy - Supercontinuum-laser-based spectral illumination for spinning disk confocal microscopy. Next, design concepts were theoretically evaluated and optimized, leading to prototype testing. To evaluate the performance of each concept, prototype systems were integrated with other systems and subsystems, calibrated and feasibility assays were executed. This dissertation is divided into three main sections: 1) early development feasibility results of an excitation-scanning widefield system of systems prototype utilizing LED-based HSI, 2) Excitation-scanning HSI and image analysis methods used for endmember identification in fluorescence microscopy studies, and 3) early development feasibility of an excitation-scanning confocal SoS prototype utilizing a supercontinuum laser light source. Integration and testing results proved initial feasibility of both LED-based and broadband-based SoSs. The LED-based light source was successfully tested on a widefield microscope, while the broadband light source system was successfully tested on a confocal microscope. Feasibility for the LED-based system showed that further optical transmission optimization is needed to achieve high acquisition rates without compromising sensitivity or specificity. Early feasibility study results for the broadband-based system showed a successful proof of concept. Findings presented in this dissertation are expected to impact the fields of cellular physiology, medical sciences, and clinical diagnostics by providing the ability for high speed, high sensitivity microscopic imaging with spectroscopic discrimination

The Influence of Gender Stereotypes on the Growth of Gender Inequality and Domestic Violence in Russia in the Context of the COVID-19 Pandemic

The present article is concerned with the influence of gender stereotypes on gender inequality and violence against women in modern Russia as well as the response of government institutions and civil society organisations to domestic violence incidents under lockdown. Conclusions on the role of stereotypes in the growth of inequality during the COVID-19 pandemic are based on findings of the research carried out by the Russian Public Opinion Research Center (VTsIOM) and the Institute of Socio-Economic Studies of Population of the Federal Center of Theoretical and Applied Sociology of the Russian Academy of Sciences. The COVID-19 pandemic revealed profound vulnerabilities concerning the state of women and exacerbated the current issues of gender discrimination. Today, discrimination has become obvious, and, to a certain degree, the state has recognised its prevalence in the labour market as well as in the areas of political activities and career advancement. However, existence of gender discrimination is still negated when it comes to issues of violence against women and reproductive rights. In general, the measures that have been implemented that aim to reduce women\u27s vulnerability are fragmentary and inadequate

The GW/WG repeats of Drosophila GW182 function as effector motifs for miRNA-mediated repression

The control of messenger RNA (mRNA) function by micro RNAs (miRNAs) in animal cells requires the GW182 protein. GW182 is recruited to the miRNA repression complex via interaction with Argonaute protein, and functions downstream to repress protein synthesis. Interaction with Argonaute is mediated by GW/WG repeats, which are conserved in many Argonaute-binding proteins involved in RNA interference and miRNA silencing, from fission yeast to mammals. GW182 contains at least three effector domains that function to repress target mRNA. Here, we analyze the functions of the N-terminal GW182 domain in repression and Argonaute1 binding, using tethering and immunoprecipitation assays in Drosophila cultured cells. We demonstrate that its function in repression requires intact GW/WG repeats, but does not involve interaction with the Argonaute1 protein, and is independent of the mRNA polyadenylation status. These results demonstrate a novel role for the GW/WG repeats as effector motifs in miRNA-mediated repressio

Recalculation of dose for each fraction of treatment on TomoTherapy.

OBJECTIVE: The VoxTox study, linking delivered dose to toxicity requires recalculation of typically 20-37 fractions per patient, for nearly 2000 patients. This requires a non-interactive interface permitting batch calculation with multiple computers. METHODS: Data are extracted from the TomoTherapy(®) archive and processed using the computational task-management system GANGA. Doses are calculated for each fraction of radiotherapy using the daily megavoltage (MV) CT images. The calculated dose cube is saved as a digital imaging and communications in medicine RTDOSE object, which can then be read by utilities that calculate dose-volume histograms or dose surface maps. The rectum is delineated on daily MV images using an implementation of the Chan-Vese algorithm. RESULTS: On a cluster of up to 117 central processing units, dose cubes for all fractions of 151 patients took 12 days to calculate. Outlining the rectum on all slices and fractions on 151 patients took 7 h. We also present results of the Hounsfield unit (HU) calibration of TomoTherapy MV images, measured over an 8-year period, showing that the HU calibration has become less variable over time, with no large changes observed after 2011. CONCLUSION: We have developed a system for automatic dose recalculation of TomoTherapy dose distributions. This does not tie up the clinically needed planning system but can be run on a cluster of independent machines, enabling recalculation of delivered dose without user intervention. ADVANCES IN KNOWLEDGE: The use of a task management system for automation of dose calculation and outlining enables work to be scaled up to the level required for large studies.JES is supported by Cancer Research UK through the Cambridge Cancer Centre. MR, AB and KH are supported by Cancer Research UK through the VoxTox Research Programme. NGB is supported by the NIHR Cambridge Biomedical Research Centre.This is the author accepted manuscript. The final version is available from British Institute of Radiology via http://dx.doi.org/10.1259/bjr.2015077

2011-2012 Master Class - Jon Kimura Parker (Piano)

Jon Kimura Parker Performance (January 15, 2012) - Programhttps://spiral.lynn.edu/conservatory_masterclasses/1066/thumbnail.jp

ATR maintains chromosomal integrity during postnatal cerebellar neurogenesis and is required for medulloblastoma formation

Microcephaly and medulloblastoma may both result from mutations that compromise genomic stability. We report that ATR, which is mutated in the microcephalic disorder Seckel syndrome, sustains cerebellar growth by maintaining chromosomal integrity during postnatal neurogenesis. Atr deletion in cerebellar granule neuron progenitors (CGNPs) induced proliferation-associated DNA damage, p53 activation, apoptosis and cerebellar hypoplasia in mice. Co-deletions of either p53 or Bax and Bak prevented apoptosis in Atr-deleted CGNPs, but failed to fully rescue cerebellar growth. ATR-deficient CGNPs had impaired cell cycle checkpoint function and continued to proliferate, accumulating chromosomal abnormalities. RNA-Seq demonstrated that the transcriptional response to ATR-deficient proliferation was highly p53 dependent and markedly attenuated by p53 co-deletion. Acute ATR inhibition in vivo by nanoparticle-formulated VE-822 reproduced the developmental disruptions seen with Atr deletion. Genetic deletion of Atr blocked tumorigenesis in medulloblastoma-prone SmoM2 mice. Our data show that p53-driven apoptosis and cell cycle arrest – and, in the absence of p53, non-apoptotic cell death – redundantly limit growth in ATR-deficient progenitors. These mechanisms may be exploited for treatment of CGNP-derived medulloblastoma using ATR inhibition

Assisted protein folding at low temperature: evolutionaryadaptation of the Antarctic fish chaperonin CCT and its clientproteins

Eukaryotic ectotherms of the Southern Ocean face energetic challenges to protein folding assisted by the cytosolic chaperonin CCT. We hypothesize that CCT and its client proteins (CPs) have co-evolved molecular adaptations that facilitate CCT–CP interaction and the ATP-driven folding cycle at low temperature. To test this hypothesis, we compared the functional and structural properties of CCT–CP systems from testis tissues of an Antarctic fish, Gobionotothen gibberifrons (Lo¨nnberg) (habitat/body T=-1.9 to +2˚C), and of the cow (body T=37˚C). We examined the temperature dependence of the binding of denatured CPs (bactin, b-tubulin) by fish and bovine CCTs, both in homologous and heterologous combinations and at temperatures between 24˚C and 20˚C, in a buffer conducive to binding of the denatured CP to the open conformation of CCT. In homologous combination, the percentage of G. gibberifrons CCT bound to CP declined linearly with increasing temperature, whereas the converse was true for bovine CCT. Binding of CCT to heterologous CPs was low, irrespective of temperature. When reactions were supplemented with ATP, G. gibberifrons CCT catalyzed the folding and release of actin at 2˚C. The ATPase activity of apo-CCT from G. gibberifrons at 4˚C was, 2.5-fold greater than that of apo-bovine CCT, whereas equivalent activities were observed at 20˚C. Based on these results, we conclude that the catalytic folding cycle of CCT from Antarctic fishes is partially compensated at their habitat temperature, probably by means of enhanced CP-binding affinity and increased flexibility of the CCT subunits

Accumulated dose to the rectum, measured using dose-volume histograms and dose-surface maps, is different from planned dose in all patients treated with radiotherapy for prostate cancer.

OBJECTIVE: We sought to calculate accumulated dose (DA) to the rectum in patients treated with radiotherapy for prostate cancer. We were particularly interested in whether dose-surface maps (DSMs) provide additional information to dose-volume histograms (DVHs). METHODS: Manual rectal contours were obtained for kilovoltage and daily megavoltage CT scans for 10 participants from the VoxTox study (380 scans). Daily delivered dose recalculation was performed using a ray-tracing algorithm. Delivered DVHs were summated to create accumulated DVHs. The rectum was considered as a cylinder, cut and unfolded to produce daily delivered DSMs; these were summated to produce accumulated DSMs. RESULTS: Accumulated dose-volumes were different from planned in all participants. For one participant, all DA levels were higher and all volumes were larger than planned. For four participants, all DA levels were lower and all volumes were smaller than planned. For each of these four participants, ≥1% of pixels on the accumulated DSM received ≥5 Gy more than had been planned. CONCLUSION: Differences between accumulated and planned dose-volumes were seen in all participants. DSMs were able to identify differences between DA and planned dose that could not be appreciated from the DVHs. Further work is needed to extract the dose data embedded in the DSMs. These will be correlated with toxicity as part of the VoxTox Programme. ADVANCES IN KNOWLEDGE: DSMs are able to identify differences between DA and planned dose that cannot be appreciated from DVHs alone and should be incorporated into future studies investigating links between DA and toxicity.JES is supported by Cancer Research UK through the Cambridge Cancer Centre. NGB is supported by the NIHR Cambridge Biomedical Research Centre. KH, MR and AMB are supported by the VoxTox Research Programme, which is funded by Cancer Research UK.This is the final version of the article. It first appeared from the British Institute of Radiology via http://dx.doi.org/10.1259/bjr.2015024

The role of GαO-mediated signaling in the rostral ventrolateral medulla oblongata in cardiovascular reflexes and control of cardiac ventricular excitability.

The heart is controlled by the sympathetic and parasympathetic limbs of the autonomic nervous system with inhibitory signaling mechanisms recruited in both limbs. The aim of this study was to determine the role of inhibitory heterotrimeric G proteins in the central nervous mechanisms underlying autonomic control of the heart and its potential role in arrhythmogenesis. Mice with conditional deletion of the inhibitory heterotrimeric G protein GαO in the presympathetic area of the rostral ventral lateral medulla (RVLM) were generated to determine the role of GαO-mediated signalling in autonomic control and electrophysiological properties of the heart. GαO deletion within the RVLM was not associated with changes in heart rate (HR) or the arterial blood pressure at rest (home cage, normal behavior). However, exposure to stressful conditions (novel environment, hypoxia, or hypercapnia) in these mice was associated with abnormal HR responses and an increased baroreflex gain when assessed under urethane anesthesia. This was associated with shortening of the ventricular effective refractory period. This phenotype was reversed by systemic beta-adrenoceptor blockade, suggesting that GαO depletion in the RVLM increases central sympathetic drive. The data obtained support the hypothesis that GαO-mediated signaling within the presympathetic circuits of the RVLM contributes to the autonomic control of the heart. GαO deficiency in the RVLM has a significant impact on cardiovascular responses to stress, cardiovascular reflexes and electrical properties of the heart.This research was supported by the Medical Research Council (MRC Clinical Research Training Fellowship to RA), British Heart Foundation (Ref: RG/14/4/30736), Wellcome Trust (Wellcome Trust Senior Research Fellowship to AVG; Ref: 095064), and by the Intramural Research Program of the National Institutes of Health, National Institute of Environmental Health Sciences (Project Z01- ES-101643 to LB). This work was facilitated by the National Institute for Health Research Barts Cardiovascular Biomedical Research Unit

A case study evaluation of implementation of a care pathway to support normal birth in one English birth centre: anticipated benefits and unintended consequences

Background: The policy drive for the UK National Health Service (NHS) has focused on the need for high quality services informed by evidence of best practice. The introduction of care pathways and protocols to standardise care and support implementation of evidence into practice has taken place across the NHS with limited evaluation of their impact. A multi-site case study evaluation was undertaken to assess the impact of use of care pathways and protocols on clinicians, service users and service delivery. One of the five sites was a midwifery-led Birth Centre, where an adapted version of the All Wales Clinical Pathway for Normal Birth had been implemented. Methods: The overarching framework was realistic evaluation. A case study design enabled the capture of data on use of the pathway in the clinical setting, use of multiple methods of data collection and opportunity to study and understand the experiences of clinicians and service users whose care was informed by the pathway. Women attending the Birth Centre were recruited at their 36 week antenatal visit. Episodes of care during labour were observed, following which the woman and the midwife who cared for her were interviewed about use of the pathway. Interviews were also held with other key stakeholders from the study site. Qualitative data were content analysed. Results: Observations were undertaken of four women during labour. Eighteen interviews were conducted with clinicians and women, including the women whose care was observed and the midwives who cared for them, senior midwifery managers and obstetricians. The implementation of the pathway resulted in a number of anticipated benefits, including increased midwifery confidence in skills to support normal birth and promotion of team working. There were also unintended consequences, including concerns about a lack of documentation of labour care and negative impact on working relationships with obstetric and other midwifery colleagues. Women were unaware their care was informed by a care pathway. Conclusion: Care pathways are complex interventions which generate a number of consequences for practice. Those considering introduction of pathways need to ensure all relevant stakeholders are engaged with this and develop robust evaluation strategies to accompany implementation