A machine learning‑based image segmentation method to quantify in vitro osteoclast culture endpoints

Quantification of in vitro osteoclast cultures (e.g. cell number) often relies on manual counting methods. These approaches are labour intensive, time consuming and result in substantial inter- and intra-user variability. This study aimed to develop and validate an automated workflow to robustly quantify in vitro osteoclast cultures. Using ilastik, a machine learning-based image analysis software, images of tartrate resistant acid phosphatase-stained mouse osteoclasts cultured on dentine discs were used to train the ilastik-based algorithm. Assessment of algorithm training showed that osteoclast numbers strongly correlated between manual- and automatically quantified values (r = 0.87). Osteoclasts were consistently faithfully segmented by the model when visually compared to the original reflective light images. The ability of this method to detect changes in osteoclast number in response to different treatments was validated using zoledronate, ticagrelor, and co-culture with MCF7 breast cancer cells. Manual and automated counting methods detected a 70% reduction (p < 0.05) in osteoclast number, when cultured with 10 nM zoledronate and a dose-dependent decrease with 1-10 μM ticagrelor (p < 0.05). Co-culture with MCF7 cells increased osteoclast number by ≥ 50% irrespective of quantification method. Overall, an automated image segmentation and analysis workflow, which consistently and sensitively identified in vitro osteoclasts, was developed. Advantages of this workflow are (1) significantly reduction in user variability of endpoint measurements (93%) and analysis time (80%); (2) detection of osteoclasts cultured on different substrates from different species; and (3) easy to use and freely available to use along with tutorial resources

Catatonic Episodes Related to Substance Use: A Cross-Sectional Study Using Electronic Healthcare Records

OBJECTIVE: Substance use has increasingly been linked to the onset of catatonic episodes; however, no large observational studies have examined this association. This study aimed to identify catatonic episodes temporally associated with acute intoxication, withdrawal or chronic substance use, investigate which substances were involved, and compare clinical characteristics of substance-related and non-substance-related catatonic episodes. METHODS: This study retrospectively identified all catatonic episodes recorded in an electronic case register hosted at a large secondary mental health trust in London, UK. Episodes were categorized as substance-related if the clinical record reported either a positive urine drug screen, an ICD-10 diagnosis of a mental or behavioral disorder due to substance use, or documented substance use between two weeks prior to the catatonic episode and the date of the catatonic episode. RESULTS: 108 of 2130 catatonic episodes (5.1%) were deemed substance-related. The number of contemporaneously reported substance-related episodes increased between 2007 and 2016 [r = 0.72, p = 0.02]. Episodes in the context of acute intoxication (n = 54) were most frequently related to cannabis (n = 31) or cocaine (n = 5) use, whilst those in the context of drug withdrawal (n = 8) were most commonly related to alcohol, opioids and benzodiazepines. There were 50 episodes of catatonia associated with chronic substance use without intoxication or withdrawal, of which the majority were related to cannabis use (n = 37). 21 episodes had overlapping intoxication, withdrawal and chronic use of different substances within an episode. Compared to catatonic episodes not related to substance use, episodes of substance-related catatonia occurred in individuals who were younger (mean age 31.3 years [SD 12.2] vs 35.7 years [SD 16.3], p = 0.01) and more likely to be men (74.0% vs 54.3%, p < 0.001). The clinical features of catatonia were similar between the two groups. CONCLUSIONS: A relatively small proportion of catatonic episodes were temporally associated with reported substance use within their electronic records. Substance-related catatonic episodes were mostly related to cannabis use, but other substances including cocaine, alcohol, opioids and benzodiazepines were sometimes implicated. This is likely an underestimate of substance-related catatonia use due to issues with documentation and appropriate investigation

Tumor-Induced IL-6 Reprograms Host Metabolism to Suppress Anti-tumor Immunity

In patients with cancer, the wasting syndrome, cachexia, is associated with caloric deficiency. Here, we describe tumor-induced alterations of the host metabolic response to caloric deficiency that cause intratumoral immune suppression. In pre-cachectic mice with transplanted colorectal cancer or autochthonous pancreatic ductal adenocarcinoma (PDA), we find that IL-6 reduces the hepatic ketogenic potential through suppression of PPARalpha, the transcriptional master regulator of ketogenesis. When these mice are challenged with caloric deficiency, the resulting relative hypoketonemia triggers a marked rise in glucocorticoid levels. Multiple intratumoral immune pathways are suppressed by this hormonal stress response. Moreover, administering corticosterone to elevate plasma corticosterone to a level that is lower than that occurring in cachectic mice abolishes the response of mouse PDA to an immunotherapy that has advanced to clinical trials. Therefore, tumor-induced IL-6 impairs the ketogenic response to reduced caloric intake, resulting in a systemic metabolic stress response that blocks anti-cancer immunotherapy.We also thank the University of Cambridge, Cancer Research UK, the CRUK Cambridge Institute Core Facilities, and Hutchison Whampoa Limited. This work was also supported by the Lustgarten Foundation for Pancreatic Cancer Research, the Ludwig Institute for Cancer Research, the NIHR Biomedical Research Centre, and the Cambridge ECMC. T.R.F. was supported by the Rosetrees Trust and the Cambridge School of Clinical Medicine’s MB/PhD Programme, T.J. was supported by the Wellcome Trust Translational Medicine and Therapeutics Programme and the University of Cambridge Department of Oncology (RJAG/076), C.M.C. was supported by the Cambridge University Hospitals NHS Foundation Trust, E.W.R. was supported by the CRI Irvington Postdoctoral Fellowship Program, and A.P.C. was supported by the Medical Research Council (MRC) Metabolic Diseases Unit (MRC_MC_UU_12012/1). D.T.F. is a Distinguished Scholar of the Lustgarten Foundation

Lithium atom interferometer using laser diffraction : description and experiments

We have built and operated an atom interferometer of the Mach-Zehnder type. The atomic wave is a supersonic beam of lithium seeded in argon and the mirrors and beam-splitters for the atomic wave are based on elastic Bragg diffraction on laser standing waves at 671 nm. We give here a detailed description of our experimental setup and of the procedures used to align its components. We then present experimental signals, exhibiting atomic interference effects with a very high visibility, up to 84.5 %. We describe a series of experiments testing the sensitivity of the fringe visibility to the main alignment defects and to the magnetic field gradient.Comment: 8 avril 200

Parenting in a warming world: thermoregulatory responses to heat stress in an endangered seabird

This is the final version. Available on open access from Oxford University Press via the DOI in this recordThe frequency of extreme weather events, including heat waves, is increasing with climate change. The thermoregulatory demands resulting from hotter weather can have catastrophic impacts on animals, leading to mass mortalities. Although less dramatic, animals also experience physiological costs below, but approaching, critical temperature thresholds. These costs may be particularly constraining during reproduction, when parents must balance thermoregulation against breeding activities. Such challenges should be acute among seabirds, which often nest in locations exposed to high solar radiation and predation risk. The globally endangered bank cormorant Phalacrocorax neglectus breeds in southern Africa in the winter, giving little scope for poleward or phenological shifts in the face of increasing temperatures. Physiological studies of endangered species sensitive to human disturbance, like the bank cormorant, are challenging, because individuals cannot be captured for experimental research. Using a novel, non-invasive, videographic approach, we investigated the thermoregulatory responses of this seabird across a range of environmental temperatures at three nesting colonies. The time birds spent gular fluttering, a behaviour enhancing evaporative heat loss, increased with temperature. Crouching or standing birds spent considerably less time gular fluttering than birds sitting on nests (ca 30% less at 22°C), showing that postural adjustments mediate exposure to heat stress and enhance water conservation. Crouching or standing, however, increases the vulnerability of eggs and chicks to suboptimal temperatures and/or expose nest contents to predation, suggesting that parents may trade-off thermoregulatory demands against offspring survival. We modelled thermoregulatory responses under future climate scenarios and found that nest-bound bank cormorants will gular flutter almost continuously for several hours a day by 2100. The associated increase in water loss may lead to dehydration, forcing birds to prioritize survival over breeding, a trade-off that would ultimately deteriorate the conservation status of this species.National Research FoundationDST-NRF Centre of Excellence fund at the FitzPatrick Institute of African OrnithologyLeiden Conservation Foundatio

The development of a new measure of quality of life in the management of gastro-oesophageal reflux disease: the Reflux Questionnaire.

INTRODUCTION This paper reports on the development of a new measure of health-related quality of life for use among patients with gastro-oesophageal reflux disease (GORD), funded as part of the REFLUX trial. This is a large UK multi centre trial that aims to compare the clinical and cost effectiveness of minimal access surgery with best medical treatment for patients with GORD within the NHS. Method Potential items were identified via a series of interviews and focus groups carried out with patients who were receiving/had received medical or surgical treatment for GORD. The final measure consisted of 31 items covering 7 categories (Heartburn; Acid reflux; Wind; Eating and swallowing; Bowel movements; Sleep; Work, physical and social activities). The measure produced two outputs: a quality of life score (RQLS) and five Reflux symptom scores. Reliability (internal consistency), criterion validity with the SF-36 and, sensitivity to change in terms of relationship with reported change in prescribed medication were assessed amongst a sample of 794 patients recruited into the trial. RESULTS The measure was shown to be internally consistent, to show criterion validity with the SF-36 and sensitive to changes in patients use of prescribed medication at baseline and 3 month follow-up. DISCUSSION The Reflux questionnaire is a new self-administered questionnaire for use amongst patients with GORD. Initial findings suggest that the new measure is valid, reliable, acceptable to respondents and simple to administer in both a clinical and research context

Can HRCT be used as a marker of airway remodelling in children with difficult asthma?

BACKGROUND: Whole airway wall thickening on high resolution computed tomography (HRCT) is reported to parallel thickening of the bronchial epithelial reticular basement membrane (RBM) in adult asthmatics. A similar relationship in children with difficult asthma (DA), in whom RBM thickening is a known feature, may allow the use of HRCT as a non-invasive marker of airway remodelling. We evaluated this relationship in children with DA. METHODS: 27 children (median age 10.5 [range 4.1-16.7] years) with DA, underwent endobronchial biopsy from the right lower lobe and HRCT less than 4 months apart. HRCTs were assessed for bronchial wall thickening (BWT) of the right lower lobe using semi-quantitative and quantitative scoring techniques. The semi-quantitative score (grade 0-4) was an overall assessment of BWT of all clearly identifiable airways in HRCT scans. The quantitative score (BWT %; defined as [airway outer diameter - airway lumen diameter]/airway outer diameter x100) was the average score of all airways visible and calculated using electronic endpoint callipers. RBM thickness in endobronchial biopsies was measured using image analysis. 23/27 subjects performed spirometry and the relationships between RBM thickness and BWT with airflow obstruction evaluated. RESULTS: Median RBM thickness in endobronchial biopsies was 6.7(range 4.6-10.0) microm. Median qualitative score for BWT of the right lower lobe was 1(range 0-1.5) and quantitative score was 54.3 (range 48.2-65.6)%. There was no relationship between RBM thickness and BWT in the right lower lobe using either scoring technique. No relationship was found between FEV1 and BWT or RBM thickness. CONCLUSION: Although a relationship between RBM thickness and BWT on HRCT has been found in adults with asthma, this relationship does not appear to hold true in children with D