Semi-automated Analyses Research Articles

Semi-automated finite element analyses of surgically treated acetabular fractures to investigate the biomechanical behaviour of patient-specific compared to conventional implants

BackgroundIn acetabular fracture surgery, understanding the biomechanical behaviour of fractures and implants is beneficial for clinical decision-making about implant selection and postoperative (early) weightbearing protocols. This study outlines a novel approach for creating finite element models (FEA) from actual clinical cases. Our objectives were to (1) create a detailed semi-automatic three-dimensional FEA of a patient with a transverse posterior wall acetabular fracture and (2) biomechanically compare patient-specific implants with manually bent off-the-shelf implants.MethodsA computational study was performed in which we developed three finite element models. The models were derived from clinical imaging data of a 20-year-old male with a transverse posterior wall acetabular fracture treated with a patient-specific implant. This implant was designed to fit the patient's anatomy and fracture configuration, allowing for optimal placement and predetermined screw trajectories. The three FEA models included an intact hemipelvis for baseline comparison, one with a fracture fixated with a patient-specific implant, and another with a conventional implant. Two loading conditions were investigated: standing up and peak walking forces. Von Mises stress and displacement patterns in bone, implants and screws were analysed to assess the biomechanical behaviour of fracture fixation with either a patient-specific versus a conventional implant.ResultsThe finite element models demonstrated that for a transverse posterior wall type fracture, a patient-specific implant resulted in lower peak stresses in the bone (30 MPa and 56 MPa) in standing-up and peak walking scenario, respectively, compared to the conventional implant model (46 MPa and 90 MPa). The results suggested that patient-specific implant could safely withstand standing-up and walking after surgery, with maximum von Mises stresses in the implant of 156 MPa and 371 MPa, respectively. The results from the conventional implant indicate a likelihood of implant failure, with von Mises stresses in the implant (499 MPa and 1000 MPa) exceeding the yield stress of stainless steel.ConclusionThis study presents a workflow for conducting finite element analysis of real clinical cases in acetabular fracture surgery. This concept of personalized biomechanical fracture and implant assessment can eventually be applied in clinical settings to guide implant selection, compare conventional implants with innovative patient-specific ones, optimizing implant designs (including shape, size, materials, screw positions), and determine whether immediate full weight-bearing can be safely permitted.

Associations between medications and carotid artery plaque morphology using semi-automated CTA analyses.

Associations between medications and carotid artery plaque morphology using semi-automated CTA analyses.

Metaphern am Lebensende: Resilienz als Widerstand oder Akzeptanz

The article provides an overview of the research project "Metaphor in end-of-life care", which was conducted at Lancaster University (UK) between 2011 and 2014. The goals of the project were to show a)how patients, family members and healthcare professionals use metaphors to talk about their experiences with and expectations of end-of-life care, b)what experiences and needs are suggested by the use of metaphors by these groups and c)what contribution the results can make to communication in the healthcare system. Acorpus consisting of interviews with the various groups of people and their postings in online fora were subject to both manual and semi-automated analyses. The results show that especially the online communications by patients and family members are characterized by violence and journey metaphors. The use of metaphors by these groups suggests aneed for solidarity and community and reflects both positive and negative personal experiences. Resilience can be understood as resistance against illness or as acceptance of it.

NIMG-71. REAL-WORLD ASSESSMENT OF IDH1 INHIBITOR IVOSIDENIB IN IDH MUTANT LOWER GRADE GLIOMAS – THE JOHNS HOPKINS EXPERIENCE

Abstract BACKGROUND As clinical trials are ongoing to establish the role of IDH inhibitors in the treatment of lower-grade gliomas, long-term data is not yet available. Hence, here we share our real-world experience with off-label ivosidenib. METHODS In this single-institution retrospective analysis, we included chemo/radiotherapy-naïve, gadolinium non-enhancing WHO Grade 2/3 IDH1-mutated astrocytomas (AS) and oligodendrogliomas (OG) that were treated with off-label ivosidenib for ≥ 5 months for active tumor growth. We performed semi-automated volumetric analyses on T2/FLAIR images in 3D Slicer v4.11. Annualized growth-rates before and on ivosidenib were calculated, and time to initial response (reduction in growth-rate), growth arrest (growth-rate ≤ 0cc/yr) and shrinkage in patients with confirmatory data available (≥ 1 confirmatory follow-up scan). Progression-free survival (PFS), at 6 and 12 months were assessed (progression defined as ≥ 40% volume increase per RANO 3D). RESULTS We studied 12 patients (2F, median age 46yrs [range 26-60], 8/4 AS/OG; 4 grade 3) with an average of 17.2 months (5-33) follow-up on ivosidenib. Eleven (92%) patients had confirmatory data. Response rate was 82% (9/11) with median time of 4.9 (3.5-6.3) months to initial response and 11.1 (4.1-21.4) months to growth arrest. Growth arrest occurred 3-folds faster in OG than in AS (4.2±0.8 vs.12.5±4.2 months, p=0.014). Mean best response was 11.4±12.8% volume reduction, after a median of 12.1±9.3 months on ivosidenib. Mean PFS was 24.4 months (CI95%18.8-29.9). PFS-6 and PFS-12 were 91%, and 88% (7/8), respectively. No major toxicity was noted. There were 3 discontinuations, two for radiographic progression, one for increased seizure frequency. CONCLUSIONS In this small cohort, ivosidenib was well-tolerated with a high response rate; however, responses took up to a year to become evident and seemingly faster in oligodendrogliomas. These data suggest that ivosidenib might require more than 5 months of commitment and volumetrics may help the detection of early response.

Advanced Image Analysis Methods for Automated Segmentation of Subnuclear Chromatin Domains.

The combination of ever-increasing microscopy resolution with cytogenetical tools allows for detailed analyses of nuclear functional partitioning. However, the need for reliable qualitative and quantitative methodologies to detect and interpret chromatin sub-nuclear organization dynamics is crucial to decipher the underlying molecular processes. Having access to properly automated tools for accurate and fast recognition of complex nuclear structures remains an important issue. Cognitive biases associated with human-based curation or decisions for object segmentation tend to introduce variability and noise into image analysis. Here, we report the development of two complementary segmentation methods, one semi-automated (iCRAQ) and one based on deep learning (Nucl.Eye.D), and their evaluation using a collection of A. thaliana nuclei with contrasted or poorly defined chromatin compartmentalization. Both methods allow for fast, robust and sensitive detection as well as for quantification of subtle nucleus features. Based on these developments, we highlight advantages of semi-automated and deep learning-based analyses applied to plant cytogenetics.

UAV applications to assess short-term dynamics of slow-moving landslides under dense forest cover

Abstract The paper presents a methodology to rapidly assess and map the landslide kinematics in areas with dense vegetation cover. The method uses aerial imagery collected with UAVs (Unmanned Aerial Vehicles) and their derived products obtained from the structure from motion technique. The landslide analysed in the current paper occurred in the spring of 2021 and is located in Livadea village from Curvature Subcarpathians, Romania. This landslide affected the houses in the vicinity, and people were relocated because of the risk of landslide reactivation. To mitigate the landslide consequences, a preliminary investigation based on UAV imagery and geological-geomorphological field surveys was carried out to map the active parts of the landslide and establish evacuation measures. Three UAV flights were performed between 6 May and 10 June using DJI Phantom 4 and Phantom 4 RTK UAVs (Real-Time Kinematic Unmanned Aerial Vehicles). Because it is a densely forested area, semi-automated analyses of the landslide kinematics and change detection analysis were not possible. Instead, the landslide displacement rates and the changes in terrain morphology were assessed by manually interpolating the landmarks, mostly tilted trees, collected from all three UAV flights. The results showed an average displacement of approximately 20 m across the landslides, with maximum values reaching 45 m in the transport area and minimum values below 1 m in the toe area. This approach proved quick and efficient for rapid landslide investigations in a densely forested area when fast response and measures are necessary to reduce the landslide consequences.

Hate speech’s double damage: A semi-automated approach toward direct and indirect targets

Democracies around the world have been facing increasing challenges with hate speech online as it contributes to a tense and thus less discursive public sphere. In that, hate speech online targets free speech both directly and indirectly, through harassments and explicit harm as well as by informing a vicious environment of irrationality, misrepresentation, or disrespect. Consequently, platforms have implemented varying means of comment-moderation techniques, depending both on policy regulations and on the quantity and quality of hate speech online. This study seeks to provide descriptive measures between direct and indirect targets in light of different incentives and practices of moderation on both social media and news outlets. Based on three distinct samples from German Twitter, YouTube, and a set of four news outlets, it applies semi-automated content analyses using a set of five cross-sample classifiers. Thereby, the largest amounts of visible hate speech online depict rather implicit devaluations of ideas or behavior. More explicit forms of hate speech online, such as insult, slander, or vulgarity, are only rarely observable and accumulate around certain events (Twitter) or single videos (YouTube). Moreover, while hate speech on Twitter and YouTube tends to target particular groups or individuals, hate speech below news articles shows a stronger focus on debates. Potential reasons and implications are discussed in light of political and legal efforts in Germany.

Abstract 9821: Fully Automated Measurements of Right Ventricular Strain in Children: Validation Using Conventional Manual Technique

Introduction: Fully automated strain measurements have been developed for the left ventricle. However, they have not been studied in the right ventricle (RV) of adults or children. Hypothesis: Automated strain measurements of RV will be comparable with manual strain measurements. Methods: We prospectively enrolled 94 patients, who underwent clinically indicated echocardiograms (mean age: 13.0±4.4 years). Strain measurements included RV free wall longitudinal strain (RVFWLS) and RV “global” longitudinal strain (RVGLS), both measured from the RV-focused apical 4-chamber view. Strain parameters were measured using three methods: 1) Fully automated (AutoStrain, Phillips Medical Systems), 2) Semiautomated (manual correction after automated measurements), 3) Manual measurements (2D-CPA, Tomtec). We also recorded the time required for measurements in all patients. Automated measurements relied on an anatomical intelligence program that was trained by adult datasets. Results: Fully automated measurements were feasible in all cases. However, 40% of patients required manual correction after automated measurements, mostly at the apex and RV lateral annulus at endsystole. The results are depicted in Table 1. Fully automated RVFWS showed moderate to good correlation with manual measurements (r=0.71, p<0.001). Semiautomated measurements showed a stronger correlation (r=0.78, p<0.001) and less bias than fully automated measurements ( Figure 1 ). The time needed for fully and semi-automated analyses were 9±2.6 and 68±25 seconds, respectively (p<0.001). Conclusions: Automated strain may provide rapid and clinically feasible means to assess RV function in children. Presently, a significant number of patients need manual correction, especially at endsystole. We speculate that accuracy of AutoStrain may improve in the future with inclusion of pediatric training datasets, as the anatomical intelligence program currently relies on adult datasets.

Testing temporal benchmarks effects on the implementation of the new Brazilian Forest Act

Native vegetation in private lands plays an important role in providing ecosystem services and safeguarding biodiversity worldwide. Legal protection rules of this vegetation are thus crucial. In Brazil, since 1934, there has been a requirement for landowners to preserve a percentage of their land covered with native vegetation, the Legal Reserve. However, the 1934 legislation applies only to arboreal vegetation, while the protection of savannahs and grasslands begins after 1965 and 1989. The reference year for the application of Legal Reserves can thus have significant impacts on the amount of native vegetation legally protected. Here we tested the effects on Legal Reserve cover in São Paulo State, considering two different legal benchmarks as starting points: 1934 and 1965. Given that the 1934 law precedes the first aerial mapping in Brazil, we developed a methodology to estimate past native vegetation cover through a probabilistic approach. The forest deficit considering 1934 as the initial point is 3% lower than the one starting with the 1965 benchmark. Comparing both scenarios, the 1934 led to a reduction of 1255 farms with Legal Reserve deficit from a total of 30,417. For both scenarios, Legal Reserve deficits were concentrated in the West, Northwest, and Mid-west regions of the state. The outcomes showed that the benchmark change does not significantly affect the total area of Legal Reserves protection, the number of farms potentially benefited by this mechanism, and the amount of native vegetation deficit. However, the use of the 1934 as an initial date for considering protection of Legal Reserve can delay the implementation of the law due to a time-consuming farm-by-farm analysis, once the probabilistic map has an intrinsic limitation for an automatic process. Thus, the environmental gains with the adoption of 1934 as the initial date do not overcome the limitations of using a probabilistic map, suggesting that effective law enforcement depends on reliable and more recent baselines, allowing semi-automated analyses. This scientific evidence can be adopted by the other Brazilian States that have not yet defined the initial legal benchmark, balancing the advantages and disadvantages of adopting the 1934 Forest Act.

Correlation between computed tomography angiography and histology of carotid artery atherosclerosis: Can semi-automated imaging software predict a plaque's composition?

Using computed tomography angiography to differentiate between components of carotid atherosclerotic lesions remains largely elusive. This study sought to validate a semi-automated software for computed tomography angiography plaque analysis using histologic comparisons. A retrospective review was performed of consecutive patients that underwent a carotid endarterectomy, with pre-procedural computed tomography angiography imaging of the cervical arterial vasculature available for review. Images were evaluated using a commercially-available software package, which produced segmented analyses of intraplaque components (e.g. intraplaque hemorrhage, lipid-rich necrotic core, and calcifications). On imaging, each component was assessed in terms of its (1) presence or absence, and (2) both volume and proportion of the total plaque volume (if present). On histological evaluation of carotid endarterectomy specimens, each component was evaluated as an estimated proportion of total plaque volume. Of 80 included patients, 30 (37.5%) were female. The average age was 69.7 years (SD = 9.1). Based on imaging, intraplaque hemorrhage was the smallest contributor to plaque composition (1.2% of volumes on average). Statistically significant linear associations were noted between the proportion of intraplaque hemorrhage, lipid-rich necrotic core, and calcifications on histology and the volume of each component on imaging (p values ranged from 0.0008 to 0.01). Area under curve were poor for intraplaque hemorrhage and lipid-rich necrotic core (0.59 and 0.61, respectively) and acceptable for calcifications (0.73). Semi-automated analyses of computed tomography angiography have limited diagnostic accuracy in the detection of intraplaque hemorrhage and lipid-rich necrotic core in carotid artery plaques. However, volumetric imaging measurements of different components corresponded with histologic analysis.

Using tri-axial accelerometer loggers to identify spawning behaviours of large pelagic fish

BackgroundTri-axial accelerometers have been used to remotely describe and identify in situ behaviours of a range of animals without requiring direct observations. Datasets collected from these accelerometers (i.e. acceleration, body position) are often large, requiring development of semi-automated analyses to classify behaviours. Marine fishes exhibit many “burst” behaviours with high amplitude accelerations that are difficult to interpret and differentiate. This has constrained the development of accurate automated techniques to identify different “burst” behaviours occurring naturally, where direct observations are not possible.MethodsWe trained a random forest machine learning algorithm based on 624 h of accelerometer data from six captive yellowtail kingfish during spawning periods. We identified five distinct behaviours (swim, feed, chafe, escape, and courtship), which were used to train the model based on 58 predictive variables.ResultsOverall accuracy of the model was 94%. Classification of each behavioural class was variable; F1 scores ranged from 0.48 (chafe) – 0.99 (swim). The model was subsequently applied to accelerometer data from eight free-ranging kingfish, and all behaviour classes described from captive fish were predicted by the model to occur, including 19 events of courtship behaviours ranging from 3 s to 108 min in duration.ConclusionOur findings provide a novel approach of applying a supervised machine learning model on free-ranging animals, which has previously been predominantly constrained to direct observations of behaviours and not predicted from an unseen dataset. Additionally, our findings identify typically ambiguous spawning and courtship behaviours of a large pelagic fish as they naturally occur.

The structure of environmental governance: How public policies connect and partition California's oil and gas policy landscape

In this paper, we ask how the written composition of public policies structure an environmental governance system. We answer this question using semi-automated text analyses of 22 state-level policies governing oil and gas development in California between 2007 and 2017. The findings portray an environmental governance system that is both partitioned and connected into different focal areas (called “targeted action situations”) through certain actors, issues, and rules. We conclude with substantive insights about California's oil and gas governance system, as well as theoretical and methodological contributions for analyzing the composition of public policy to advance knowledge about hybrid governance.

Semi-Automated Cell and Tissue Analyses Reveal Regionally Specific Morphological Alterations of Immune and Neural Cells in a Porcine Middle Cerebral Artery Occlusion Model of Stroke.

Histopathological analysis of cellular changes in the stroked brain provides critical information pertaining to inflammation, cell death, glial scarring, and other dynamic injury and recovery responses. However, commonly used manual approaches are hindered by limitations in speed, accuracy, bias, and the breadth of morphological information that can be obtained. Here, a semi-automated high-content imaging (HCI) and CellProfiler histological analysis method was developed and used in a Yucatan miniature pig permanent middle cerebral artery occlusion (pMCAO) model of ischemic stroke to overcome these limitations. Evaluation of 19 morphological parameters in IBA1+ microglia/macrophages, GFAP+ astrocytes, NeuN+ neuronal, FactorVIII+ vascular endothelial, and DCX+ neuroblast cell areas was conducted on porcine brain tissue 4 weeks post pMCAO. Out of 19 morphological parameters assessed in the stroke perilesional and ipsilateral hemisphere regions (38 parameters), a significant change in measured IBA1+ parameters, GFAP+ parameters, NeuN+ parameters, FactorVIII+ parameters, and DCX+ parameters were observed in stroked vs. non-stroked animals. Principal component analysis (PCA) and correlation analyses demonstrated that stroke-induced significant and predictable morphological changes that demonstrated strong relationships between IBA1+, GFAP+, and NeuN+ areas. Ultimately, this unbiased, semi-automated HCI and CellProfiler histopathological analysis approach revealed regional and cell specific morphological signatures of immune and neural cells after stroke in a highly translational porcine model. These identified features can provide information of disease pathogenesis and evolution with high resolution, as well as be used in therapeutic screening applications.

Volumetric MicroCT Intensity Histograms of Fatty Infiltration Correlate with the Mechanical Strength of Rotator Cuff Repairs: An Ex Vivo Rabbit Model.

Fatty infiltration of the rotator cuff occurs after injury to the tendon and results in a buildup of adipose in the muscle. Fatty infiltration may be a biomarker for predicting future injuries and mechanical properties after tendon repair. As such, quantifying fatty infiltration accurately could be a relevant metric for determining the success of tendon repairs. Currently, fatty infiltration is quantified by an experienced observer using the Goutallier or Fuchs staging system, but because such score-based quantification systems rely on subjective assessments, newer techniques using semiautomated analyses in CT and MRI were developed and have met with varying degrees of success. However, semiautomated analyses of CT and MRI results remain limited in cases where only a few two-dimensional slices of tissue are examined and applied to the three-dimensional (3-D) tissue structure. We propose that it is feasible to assess fatty infiltration within the 3-D volume of muscle and tendon in a semiautomated fashion by selecting anatomic features and examining descriptive metrics of intensity histograms collected from a cylinder placed within the central volume of the muscle and tendon of interest. (1) Do descriptive metrics (mean and SD) of intensity histograms from microCT images correlate with the percentage of fat present in muscle after rotator cuff repair? (2) Do descriptive metrics of intensity histograms correlate with the maximum load during mechanical testing of rotator cuff repairs? We developed a custom semiautomated program to generate intensity histograms based on user-selected anatomic features. MicroCT images were obtained from 12 adult female New Zealand White rabbits (age 8 to 12 months, weight 3.7 kg ± 5 kg) that were randomized to surgical repair or sham repair of an induced infraspinatus defect. Intensity histograms were generated from images of the operative and contralateral intact shoulder in these rabbits which were presented to the user in a random order without identifying information to minimize sources of bias. The mean and SD of the intensity histograms were calculated and compared with the total percentage of the volume threshold as fat. Patterns of fat identified were qualitatively compared with histologic samples to confirm that thresholding was detecting fat. We conducted monotonic tensile strength-to-failure tests of the humeral-infraspinatus bone-tendon-muscle complex, and evaluated associations between histogram mean and SDs and maximum load. The total percentage of fat was negatively correlated with the intensity histogram mean (Pearson correlation coefficient -0.92; p < 0.001) and positively with intensity histogram SD (Pearson correlation coefficient 0.88; p < 0.001), suggesting that the increase in fat leads to a reduction and wider variability in volumetric tissue density. The percentage of fat content was also negatively correlated with the maximum load during mechanical testing (Pearson correlation coefficient -78; p = 0.001), indicating that as the percentage of fat in the volume increases, the mechanical strength of the repair decreases. Furthermore, the intensity histogram mean was positively correlated with maximum load (Pearson correlation coefficient 0.77; p = 0.001) and histogram SD was negatively correlated with maximum load (Pearson correlation coefficient -0.72; p = 0.004). These correlations were strengthened by normalizing maximum load to account for animal size (Pearson correlation coefficient 0.86 and -0.9, respectively), indicating that as histogram mean decreases, the maximum load of the repair decreases and as histogram spread increases, the maximum load decreases. In this ex vivo rabbit model, a semiautomated approach to quantifying fat on microCT images was a noninvasive way of quantifying fatty infiltration associated with the strength of tendon healing. Histogram-derived variables may be useful as surrogate measures of repair strength after rotator cuff repair. The preclinical results presented here provide a foundation for future studies to translate this technique to patient studies and additional imaging modalities. This semiautomated method provides an accessible approach to quantification of fatty infiltration by users of varying experience and can be easily adapted to any intensity-based imaging approach. To translate this approach to clinical practice, this technique should be calibrated for MRI or conventional CT imaging and applied to patient scans. Further investigations are needed to assess the correlation of volumetric intensity histogram descriptive metrics to clinical mechanical outcomes.

Automated high content image analysis of dendritic arborization in primary mouse hippocampal and rat cortical neurons in culture

BackgroundPrimary neuronal cell cultures are useful for studying mechanisms that influence dendritic morphology during normal development and in response to various stressors. However, analyzing dendritic morphology is challenging, particularly in cultures with high cell density, and manual methods of selecting neurons and tracing dendritic arbors can introduce significant bias, and are labor-intensive. To overcome these challenges, semi-automated and automated methods are being developed, with most software solutions requiring computer-assisted dendrite tracing with subsequent quantification of various parameters of dendritic morphology, such as Sholl analysis. However fully automated approaches for classic Sholl analysis of dendritic complexity are not currently available. New MethodThe previously described Omnisphero software, was extended by adding new functions to automatically assess dendritic mass, total length of the dendritic arbor and the number of primary dendrites, branch points, and terminal tips, and to perform Sholl analysis. ResultsThe new functions for assessing dendritic morphology were validated using primary mouse hippocampal and rat cortical neurons transfected with a fluorescently tagged MAP2 cDNA construct. These functions allow users to select specific populations of neurons as a training set for subsequent automated selection of labeled neurons in high-density cultures. Comparison with Existing Semi-Automated MethodsCompared to manual or semi-automated analyses of dendritic arborization, the new functions increase throughput while significantly decreasing researcher bias associated with neuron selection, tracing, and thresholding. ConclusionThese results demonstrate the importance of using unbiased automated methods to mitigate experimenter-dependent bias in analyzing dendritic morphology.

The decline of organizational sociology? An empirical analysis of research trends in leading journals across half a century

Recent works see organizational sociology at the brink of irrelevance. Against this backdrop, in this article the authors want to explore the current state of organizational sociology empirically. They employ a variety of manual, automated and semi-automated content analyses to examine research articles published in generalist sociology journals since the 1950s. Contrary to contemporary pessimistic assessments, the results indicate that organizational sociology has not significantly declined over time. However, the study finds an increasing concentration on quantitative research designs, business-related topics, and only two dominant theory perspectives – neo-institutionalism and the network approach. A multifaceted decrease in variety rather than an absolute decline could be the right diagnosis.

Abstract PD9-08: Breast cancer liver metastases vascularize by vessel co-option, not angiogenesis, and have a desert immune phenotype: A histopathological and gene expression study

Abstract Phase 3 trials of bevacizumab combined with chemotherapy in metastatic breast cancer have consistently failed to demonstrate a survival benefit for the addition of bevacizumab. When cancers metastasize to highly vascular organs (including the liver), they can utilize vessel co-option, instead of angiogenesis, as a mechanism to obtain a vascular supply (1). We have repeatedly shown by histopathological analyses that almost all (95%, 2 cohorts) breast cancer liver metastases utilize vessel co-option instead of angiogenesis to vascularize (2,3). The prevalence of vessel co-option in breast cancer could explain, at least in part, why anti-angiogenic therapy has been a disappointing therapeutic approach in metastatic breast cancer. Animal models of non-angiogenic liver and lung metastases also displayed resistance to anti-VEGF treatment (3,4). We have now undertaken a gene expression study (mRNA sequencing) of targeted samples at the tumor-liver interface to discover gene expression patterns and signaling pathways that are associated with non-angiogenic growth of metastatic cancer in the liver (n = 70). A network to detect biological themes of non-angiogenic growth was built by gene set enrichment analysis. Key components of this network are: cancer cell motility and invasion, epithelial-to-mesenchymal transition, stemness and proliferation. This contrasts with the network of angiogenic liver metastases of which the most important components are inflammation and ECM remodeling. Semi-automated image analyses of CD8-immunostained section of liver metastases confirms that non-angiogenic liver metastases have a significantly lower density of CD8-positive cytotoxic T-lymphocytes at the tumor-liver interface when compared with angiogenic liver metastases (300 cells/mm2 and 1000 cells/mm2, respectively (p&amp;lt;0.0001)). In addition, a clear CXCL13-driven B-cell gene expression signature is associated with angiogenic growth of liver mets but is absent in non-angiogenic growth of breast cancer liver metastases. Gene expression patterns that may be play a role in vessel co-option are the up-regulation of LAMA3, LAMB3, LAMC2, coding for the 3 subunits of laminin-5, and of ITGA3, ITGB1, ITGA6 and ITGB4, coding for both integrin-receptors of laminin-5. This supports the concept of 'adhesive' vessel co-option during which cancer cells use the basement membrane of the co-opted blood vessels as a soil (5). In addition, the claudin-2 gene (CLDN2) is significantly overexpressed in non-angiogenic liver metastases which is consistent with earlier reports on the role of claudin-2 during breast cancer metastasis to the liver (6). In conclusion, we provide evidence, based on morphology and gene expression, for the almost exclusive non-angiogenic growth of breast cancer liver metastases. In addition, non-angiogenic breast cancer liver metastases are characterized by a desert immune phenotype. Both observations can have an impact on the treatment strategy of patients with metastatic breast cancer.

Proof of concept for quantitative urine NMR metabolomics pipeline for large-scale epidemiology and genetics.

BackgroundQuantitative molecular data from urine are rare in epidemiology and genetics. NMR spectroscopy could provide these data in high throughput, and it has already been applied in epidemiological settings to analyse urine samples. However, quantitative protocols for large-scale applications are not available.MethodsWe describe in detail how to prepare urine samples and perform NMR experiments to obtain quantitative metabolic information. Semi-automated quantitative line shape fitting analyses were set up for 43 metabolites and applied to data from various analytical test samples and from 1004 individuals from a population-based epidemiological cohort. Novel analyses on how urine metabolites associate with quantitative serum NMR metabolomics data (61 metabolic measures; n = 995) were performed. In addition, confirmatory genome-wide analyses of urine metabolites were conducted (n = 578). The fully automated quantitative regression-based spectral analysis is demonstrated for creatinine and glucose (n = 4548).ResultsIntra-assay metabolite variations were mostly <5%, indicating high robustness and accuracy of urine NMR spectroscopy methodology per se. Intra-individual metabolite variations were large, ranging from 6% to 194%. However, population-based inter-individual metabolite variations were even larger (from 14% to 1655%), providing a sound base for epidemiological applications. Metabolic associations between urine and serum were found to be clearly weaker than those within serum and within urine, indicating that urinary metabolomics data provide independent metabolic information. Two previous genome-wide hits for formate and 2-hydroxyisobutyrate were replicated at genome-wide significance.ConclusionQuantitative urine metabolomics data suggest broad novelty for systems epidemiology. A roadmap for an open access methodology is provided.

Interactions of tropomyosin Tpm1.1 on a single actin filament: A method for extraction and processing of high resolution TIRF microscopy data.

Skeletal muscle tropomyosin (Tpm1.1) is an elongated, rod-shaped, alpha-helical coiled-coil protein that forms continuous head-to-tail polymers along both sides of the actin filament. In this study we use single molecule fluorescence TIRF microscopy combined with a microfluidic device and fluorescently labelled proteins to measure Tpm1.1 association to and dissociation from single actin filaments. Our experimental setup allows us to clearly resolve Tpm1.1 interactions on both sides of the filaments. Here we provide a semi-automated method for the extraction and quantification of kymograph data for individual actin filaments bound at different Tpm1.1 concentrations. We determine boundaries on the kymograph on each side of the actin filament, based on intensity thresholding, performing fine manual editing of the boundaries (if needed) and extracting user defined kinetic properties of the system. Using our analytical tools we can determine (i) nucleation point(s) and rates, (ii) elongation rates of Tpm1.1, (iii) identify meeting points after the saturation of filament, and when dissociation occurs, (iv) initiation point(s), (v) the final dissociation point(s), as well as (vi) dissociation rates.All of these measurements can be extracted from both sides of the filament, allowing for the determination of possible differences in behaviour on the two sides of the filament, and across concentrations. The robust and repeatable nature of the method allows quantitative, semi-automated analyses to be made of large studies of acto-tropomyosin interactions, as well as for other actin binding proteins or filamentous structures, opening the way for dissection of the dynamics underlying these interactions.

Quantitative MRI volumetry in the detection of mesial temporal sclerosis in patients with refractory epilepsy

One third of patients who suffer from epilepsy are resistant to anti-epileptic medication, known as refractory epilepsy. These patients may be considered for surgical resection, where a portion of brain tissue which is believed to be involved in the seizure onset is removed. In the most common form of epilepsy, temporal lobe epilepsy (TLE), structures such as the hippocampus are often found to be involved. An indicator that the hippocampus may be related to the seizure onset is the presence of mesial temporal sclerosis (mTS), which is noted by a loss of internal architecture, reduced hippocampal volume and hyperintensity on T2-weighted MRI. The published literature indicates that a radiological evaluation of mTS supported by hippocampal volumetry can indicate a better postoperative outcome for patients. This project evaluates the use of software packages to produce volumetric estimates of these TL structures. These estimates were obtained for patients whose radiological reports indicate the presence and/or suspicion of mTS and a normal cohort. A comparison between Freesurfer and more specific hippocampal volumetric software packages was made. A database of normal MRI scans was compiled, this is age and gender matched for robust comparison and to reduce systematic error associated with comparing volumetric data. The software packages showed concordance with radiological reports by indicating a reduced hippocampal volume. Following on from this work and further validation of semi-automated volumetric analyses, it is hoped that this approach will be integrated into the pre-surgical evaluation workflow with the aim of improving the postoperative prognosis for patients.