Search results for: Marcel Kohler

Authors: Marcel Schneider, Nils Nießen

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Microscopic simulation programs enable the description of the two processes of railway operation and the previous timetabling. Occupation conflicts are often solved based on defined train priorities on both process levels. These conflict resolutions produce knock-on delays for the involved trains. The sum of knock-on delays is commonly used to evaluate the quality of railway operations. It is either compared to an acceptable level-of-service or the delays are evaluated economically by linearly monetary functions. It is impossible to properly evaluate dispatching decisions without a well-founded objective function. This paper presents a new approach for evaluation of dispatching decisions. It uses models of choice of transport and considers the behaviour of the end-costumers. These models evaluate the knock-on delays in more detail than linearly monetary functions and consider other competing modes of transport. The new approach pursues the coupling of a microscopic model of railway operation with the macroscopic model of choice of transport. First it will be implemented for the railway operations process, but it can also be used for timetabling. The evaluation considers the possibility to change over to other transport modes by the end-costumers. The new approach first looks at the rail-mounted and road transport, but it can also be extended to air transport. The split of the end-costumers is described by the modal-split. The reactions by the end-costumers have an effect on the revenues of the railway undertakings. Various travel purposes has different pavement reserves and tolerances towards delays. Longer journey times affect besides revenue changes also additional costs. The costs depend either on time or track and arise from circulation of workers and vehicles. Only the variable values are summarised in the contribution margin, which is the base for the monetary evaluation of the delays. The contribution margin is calculated for different resolution decisions of the same conflict. The conflict resolution is improved until the monetary loss becomes minimised. The iterative process therefore determines an optimum conflict resolution by observing the change of the contribution margin. Furthermore, a monetary value of each dispatching decision can also be determined.

Keywords: choice of transport, knock-on delays, monetary evaluation, railway operations

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Authors: Ines Raies, Eric Kohler, Marc Fleury, Béatrice Ledésert

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In this work, the impact of clay minerals on the formation damage of sandstone reservoirs is studied to provide a better understanding of the problem of deep geothermal reservoir permeability reduction due to fine particle dispersion and migration. In some situations, despite the presence of filters in the geothermal loop at the surface, particles smaller than the filter size (<1 µm) may surprisingly generate significant permeability reduction affecting in the long term the overall performance of the geothermal system. Our study is carried out on cores from a Triassic reservoir in the Paris Basin (Feigneux, 60 km Northeast of Paris). Our goal is to first identify the clays responsible for clogging, a mineralogical characterization of these natural samples was carried out by coupling X-Ray Diffraction (XRD), Scanning Electron Microscopy (SEM) and Energy Dispersive X-ray Spectroscopy (EDS). The results show that the studied stratigraphic interval contains mostly illite and chlorite particles. Moreover, the spatial arrangement of the clays in the rocks as well as the morphology and size of the particles, suggest that illite is more easily mobilized than chlorite by the flow in the pore network. Thus, based on these results, illite particles were prepared and used in core flooding in order to better understand the factors leading to the aggregation and deposition of this type of clay particles in geothermal reservoirs under various physicochemical and hydrodynamic conditions. First, the stability of illite suspensions under geothermal conditions has been investigated using different characterization techniques, including Dynamic Light Scattering (DLS) and Scanning Transmission Electron Microscopy (STEM). Various parameters such as the hydrodynamic radius (around 100 nm), the morphology and surface area of aggregates were measured. Then, core-flooding experiments were carried out using sand columns to mimic the permeability decline due to the injection of illite-containing fluids in sandstone reservoirs. In particular, the effects of ionic strength, temperature, particle concentration and flow rate of the injected fluid were investigated. When the ionic strength increases, a permeability decline of more than a factor of 2 could be observed for pore velocities representative of in-situ conditions. Further details of the retention of particles in the columns were obtained from Magnetic Resonance Imaging and X-ray Tomography techniques, showing that the particle deposition is nonuniform along the column. It is clearly shown that very fine particles as small as 100 nm can generate significant permeability reduction under specific conditions in high permeability porous media representative of the Triassic reservoirs of the Paris basin. These retention mechanisms are explained in the general framework of the DLVO theory

Keywords: geothermal energy, reinjection, clays, colloids, retention, porosity, permeability decline, clogging, characterization, XRD, SEM-EDS, STEM, DLS, NMR, core flooding experiments

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Authors: E. M. D. Akuoko, Eliana Vasquez Osorio, Marcel Van Herk, Marianne Aznar

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Radiotherapy is an integral part of treatment for many patients with breast cancer. However, side effects can occur, e.g., fibrosis or erythema. If patients at higher risks of radiation-induced side effects could be identified before treatment, they could be given more individual information about the risks and benefits of radiotherapy. We hypothesize that breast density is correlated with the risk of side effects and present a novel method for automatic evaluation based on radiotherapy planning CT scans. Methods: 799 supine CT scans of breast radiotherapy patients were available from the REQUITE dataset. The methodology was first established in a subset of 114 patients (cohort 1) before being applied to the whole dataset (cohort 2). All patients were scanned in the supine position, with arms up, and the treated breast (ipsilateral) was identified. Manual experts contour available in 96 patients for both the ipsilateral and contralateral breast in cohort 1. Breast tissue was segmented using atlas-based automatic contouring software, ADMIRE® v3.4 (Elekta AB, Sweden). Once validated, the automatic segmentation method was applied to cohort 2. Breast density was then investigated by thresholding voxels within the contours, using Otsu threshold and pixel intensity ranges based on Hounsfield units (-200 to -100 for fatty tissue, and -99 to +100 for fibro-glandular tissue). Volumetric breast density (VBD) was defined as the volume of fibro-glandular tissue / (volume of fibro-glandular tissue + volume of fatty tissue). A sensitivity analysis was performed to verify whether calculated VBD was affected by the choice of breast contour. In addition, we investigated the correlation between volumetric breast density (VBD) and patient age and breast size. VBD values were compared between ipsilateral and contralateral breast contours. Results: Estimated VBD values were 0.40 (range 0.17-0.91) in cohort 1, and 0.43 (0.096-0.99) in cohort 2. We observed ipsilateral breasts to be denser than contralateral breasts. Breast density was negatively associated with breast volume (Spearman: R=-0.5, p-value < 2.2e-16) and age (Spearman: R=-0.24, p-value = 4.6e-10). Conclusion: VBD estimates could be obtained automatically on a large CT dataset. Patients’ age or breast volume may not be the only variables that explain breast density. Future work will focus on assessing the usefulness of VBD as a predictive variable for radiation-induced side effects.

Keywords: breast cancer, automatic image segmentation, radiotherapy, big data, breast density, medical imaging

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Authors: Ni Putu Evi Wijayanti, Made Darmiati, Ni Ketut Wiwiek Agustina, Putu Gde Arie Yudhistira, Marcel Hardono

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This research study discusses the tourists’ perception to white water rafting service quality in Bali (Case Study: Ayung River). The aim is to determine the tourists’ perception to: firstly, the services quality of white water rafting trip in Bali, secondly, is to determine which dimensions of the service quality that need to take main handling priority in accordance with the level of important service of white water rafting company’s working performance toward the service quality of rafting in Bali especially on Ayung Riveri, lastly, is to know the efforts are needed to improve the service quality of white water rafting trip for tourist in Bali, specifically on Ayung River. This research uses the concept of the service quality with five principal dimensions, namely: Tangibles, Reliability, Responsiveness, Assurance, Empathy. Location of the research is tourist destination area of the Ayung River, that lies between the boundary of Badung Regency at Western part and Gianyar Regency eastern side. There are three rafting companies located on the Ayung River. This research took 100 respondents who were selected as a sample by using purposive sampling method. Data were collected through questionnaires distributed to domestic tourists then tabulated using the weighting scale (Likert scale) and analyzed using analysis of the benefit performance (important performance analysis) in the form of Cartesian diagram. The results of the research are translated into three points. Firstly, there are 23 indicators assessed by the service aspect of domestic tourists where the highest value is the aspect of familiarity between the tourist and employees with points (0.29) and the lowest score is the aspect of the clarity of the Ayung River water discharge value (-0.35). This shows that the indicator has not been fully able to meet the expectations of service aspects of the rating. Secondly, the dimensions of service quality that requires serious attention is the dimension of tangibles. The third point is the efforts that needs to be done adapted to the results of the Cartesian diagram breaks down into four quadrants. Based on the results of the research suggested to the manager of the white water rafting tour in order to continuously improve the service quality to tourists, performing new innovations in terms of product variations, provide insight and training to its employees to increase their competence, especially in the field of excellent service so that the satisfaction rating can be achieved.

Keywords: perception, rafting, service quality, tourist satisfaction

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Authors: Mareen Brosterhaus, Antje Hammer, Steffen Kalina, Stefan Grau, Anjali A. Roeth, Hany Ashmawy, Thomas Gross, Marcel Binnebosel, Wolfram T. Knoefel, Tanja Manser

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Background: The identification of critical incidents in hospitals is an essential component of improving patient safety. To date, various methods have been used to measure and characterize such critical incidents. These methods are often viewed by physicians and nurses as external quality assurance, and this creates obstacles to the reporting events and the implementation of recommendations in practice. One way to overcome this problem is to use tools that directly involve staff in measuring indicators of quality and safety of care in the department. One such instrument is the global trigger tool (GTT), which helps physicians and nurses identify adverse events by systematically reviewing randomly selected patient records. Based on so-called ‘triggers’ (warning signals), indications of adverse events can be given. While the tool is already used internationally, its implementation in German hospitals has been very limited. Objectives: This study aimed to assess the feasibility and potential of the global trigger tool for identifying adverse events in German hospitals. Methods: A total of 120 patient records were randomly selected from two surgical, and one neurosurgery, departments of three university hospitals in Germany over a period of two months per department between January and July, 2017. The records were reviewed using an adaptation of the German version of the Institute for Healthcare Improvement Global Trigger Tool to identify triggers and adverse event rates per 1000 patient days and per 100 admissions. The severity of adverse events was classified using the National Coordinating Council for Medication Error Reporting and Prevention. Results: A total of 53 adverse events were detected in the three departments. This corresponded to adverse event rates of 25.5-72.1 per 1000 patient-days and from 25.0 to 60.0 per 100 admissions across the three departments. 98.1% of identified adverse events were associated with non-permanent harm without (Category E–71.7%) or with (Category F–26.4%) the need for prolonged hospitalization. One adverse event (1.9%) was associated with potentially permanent harm to the patient. We also identified practical challenges in the implementation of the tool, such as the need for adaptation of the global trigger tool to the respective department. Conclusions: The global trigger tool is feasible and an effective instrument for quality measurement when adapted to the departmental specifics. Based on our experience, we recommend a continuous use of the tool thereby directly involving clinicians in quality improvement.

Keywords: adverse events, global trigger tool, patient safety, record review

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Authors: Marcel Heimar Ribeiro Utiyama, Fernanda Caveiro Correia, Dario Henrique Alliprandini

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The importance of the correct allocation of improvement programs is of growing interest in recent years. Due to their limited resources, companies must ensure that their financial resources are directed to the correct workstations in order to be the most effective and survive facing the strong competition. However, to our best knowledge, the literature about allocation of improvement programs does not analyze in depth this problem when the flow shop process has two capacity constrained resources. This is a research gap which is deeply studied in this work. The purpose of this work is to identify the best strategy to allocate improvement programs in a flow shop with two capacity constrained resources. Data were collected from a flow shop process with seven workstations in an industrial control and automation company, which process 13.690 units on average per month. The data were used to conduct a simulation with the System Dynamics-Factory Physics model. The main variables considered, due to their importance on lead time reduction, were the mean time between failures and the mean time to repair. The lead time reduction was the output measure of the simulations. Ten different strategies were created: (i) focused time to repair improvement, (ii) focused time between failures improvement, (iii) distributed time to repair improvement, (iv) distributed time between failures improvement, (v) focused time to repair and time between failures improvement, (vi) distributed time to repair and between failures improvement, (vii) hybrid time to repair improvement, (viii) hybrid time between failures improvements, (ix) time to repair improvement strategy towards the two capacity constrained resources, (x) time between failures improvement strategy towards the two capacity constrained resources. The ten strategies tested are variations of the three main strategies for improvement programs named focused, distributed and hybrid. Several comparisons among the effect of the ten strategies in lead time reduction were performed. The results indicated that for the flow shop analyzed, the focused strategies delivered the best results. When it is not possible to perform a large investment on the capacity constrained resources, companies should use hybrid approaches. An important contribution to the academy is the hybrid approach, which proposes a new way to direct the efforts of improvements. In addition, the study in a flow shop with two strong capacity constrained resources (more than 95% of utilization) is an important contribution to the literature. Another important contribution is the problem of allocation with two CCRs and the possibility of having floating capacity constrained resources. The results provided the best improvement strategies considering the different strategies of allocation of improvement programs and different positions of the capacity constrained resources. Finally, it is possible to state that both strategies, hybrid time to repair improvement and hybrid time between failures improvement, delivered best results compared to the respective distributed strategies. The main limitations of this study are mainly regarding the flow shop analyzed. Future work can further investigate different flow shop configurations like a varying number of workstations, different number of products or even different positions of the two capacity constrained resources.

Keywords: allocation of improvement programs, capacity constrained resource, hybrid strategy, lead time, mean time to repair, mean time between failures

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Authors: Chris Cadman, Marcel Strauss

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Background: In 2020, the University Hospital Wishaw in the United Kingdom became the centre for trauma and orthopaedics within its health board. This resulted in the majority of patients with suspected cauda equina syndrome (CES) being assessed and imaged at this site, putting an increased demand on MR imaging and displacing other previous activity. Following this transition, imaging requests for CES did not always follow national guidelines and would often be missing important clinical and safety information. There also appeared to be a very low positive scan rate compared with previously reported studies. In an attempt to improve patient selection and reduce the burden of CES imaging at this site clinical audit was performed. Methods: A total of 250 consecutive patients imaged to assess for CES were evaluated. Patients had to have presented to either the emergency or orthopaedic department acutely with a presenting complaint of suspected CES. Patients were excluded if they were not admitted acutely or were assessed by other clinical specialities. In total, 233 patients were included. Requests were assessed for appropriate clinical history, accurate and complete clinical assessment and MRI safety information. Clinical assessment was allocated a score of 1-6 based on information relating to history of pain, level of pain, dermatomes/myotomes affected, peri-anal paraesthesia/anaesthesia, anal tone and post-void bladder volume with each element scoring one point. Images were assessed for positive findings of CES, acquired spinal stenosis or nerve root compression. Results: Overall, 73% of requests had a clear clinical history of CES. The urgency of the request for imaging was given in 23% of cases. The mean clinical assessment score was 3.7 out of a total of 6. Overall, 2% of scans were positive for CES, 29% had acquired spinal stenosis and 30% had nerve root compression. For patients with CES, 75% had acute neurological signs compared with 68% of the study population. CES patients had a mean clinical history score of 5.3 compared with 3.7 for the study population. Overall, 95% of requests had appropriate MRI safety information. Discussion: it study included 233 patients who underwent specialist assessment and referral for MR imaging for suspected CES. Despite the serious nature of this condition, a large proportion of imaging requests did not have a clear clinical query of CES and the level of urgency was not given, which could potentially lead to a delay in imaging and treatment. Clinical examination was often also incomplete, which can make triaging of patients presenting with similar symptoms challenging. The positive rate for CES was only 2%, much below other studies which had positive rates of 6–40% with a large meta-analysis finding a mean positive rate of 19%. These findings demonstrate an opportunity to improve the quality of imaging requests for suspected CES. This may help to improve patient selection for imaging and result in a positive rate for CES imaging that is more in line with other centres.

Keywords: cauda equina syndrome, acute back pain, MRI, spine

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Authors: Geanderson Eduardo Ambrósio, Dênis Antônio Da Cunha, Marcel Viana Pires

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Historically, human development has been based on economic gains associated with intensive energy activities, which often are exhaustive in the emission of Greenhouse Gases (GHGs). It requires the establishment of targets for mitigation of GHGs in order to disassociate the human development from emissions and prevent further climate change. Brazil presents itself as one of the most GHGs emitters and it is of critical importance to discuss such reductions in intra-national framework with the objective of distributional equity to explore its full mitigation potential without compromising the development of less developed societies. This research displays some incipient considerations about which Brazil’s micro-regions should reduce, when the reductions should be initiated and what its magnitude should be. We started with the methodological assumption that human development and GHGs emissions arise in the future as their behavior was observed in the past. Furthermore, we assume that once a micro-region became developed, it is able to maintain gains in human development without the need of keep growing GHGs emissions rates. The human development index and the carbon dioxide equivalent emissions (CO2e) were extrapolated to the year 2050, which allowed us to calculate when the micro-regions will become developed and the mass of GHG’s emitted. The results indicate that Brazil must throw 300 GT CO2e in the atmosphere between 2011 and 2050, of which only 50 GT will be issued by micro-regions before it’s develop and 250 GT will be released after development. We also determined national mitigation targets and structured reduction schemes where only the developed micro-regions would be required to reduce. The micro-region of São Paulo, the most developed of the country, should be also the one that reduces emissions at most, emitting, in 2050, 90% less than the value observed in 2010. On the other hand, less developed micro-regions will be responsible for less impactful reductions, i.e. Vale do Ipanema will issue in 2050 only 10% below the value observed in 2010. Such methodological assumption would lead the country to issue, in 2050, 56.5% lower than that observed in 2010, so that the cumulative emissions between 2011 and 2050 would reduce by 130 GT CO2e over the initial projection. The fact of associating the magnitude of the reductions to the level of human development of the micro-regions encourages the adoption of policies that favor both variables as the governmental planner will have to deal with both the increasing demand for higher standards of living and with the increasing magnitude of reducing emissions. However, if economic agents do not act proactively in local and national level, the country is closer to the scenario in which emits more than the one in which mitigates emissions. The research highlighted the importance of considering the heterogeneity in determining individual mitigation targets and also ratified the theoretical and methodological feasibility to allocate larger share of contribution for those who historically emitted more. It is understood that the proposals and discussions presented should be considered in mitigation policy formulation in Brazil regardless of the adopted reduction target.

Keywords: greenhouse gases, human development, mitigation, intensive energy activities

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Authors: Marcel Schulze, Behrem Aslan, Silke Lux, Alexandra Philipsen

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Objective: Patients with Attention Deficit/Hyperactivity Disorder (ADHD) often report that they are being flooded by sensory impressions. Studies investigating sensory processing show hypersensitivity for sensory inputs across the senses in children and adults with ADHD. Especially the auditory modality is affected by deficient acoustical inhibition and modulation of signals. While studying unimodal signal-processing is relevant and well-suited in a controlled laboratory environment, everyday life situations occur multimodal. A complex interplay of the senses is necessary to form a unified percept. In order to achieve this, the unimodal sensory modalities are bound together in a process called multisensory integration (MI). In the current study we investigate MI in an adult ADHD sample using the McGurk-effect – a well-known illusion where incongruent speech like phonemes lead in case of successful integration to a new perceived phoneme via late top-down attentional allocation . In ADHD neuronal dysregulation at rest e.g., aberrant within or between network functional connectivity may also account for difficulties in integrating across the senses. Therefore, the current study includes resting-state functional connectivity to investigate a possible relation of deficient network connectivity and the ability of stimulus integration. Method: Twenty-five ADHD patients (6 females, age: 30.08 (SD:9,3) years) and twenty-four healthy controls (9 females; age: 26.88 (SD: 6.3) years) were recruited. MI was examined using the McGurk effect, where - in case of successful MI - incongruent speech-like phonemes between visual and auditory modality are leading to a perception of a new phoneme. Mann-Whitney-U test was applied to assess statistical differences between groups. Echo-planar imaging-resting-state functional MRI was acquired on a 3.0 Tesla Siemens Magnetom MR scanner. A seed-to-voxel analysis was realized using the CONN toolbox. Results: Susceptibility to McGurk was significantly lowered for ADHD patients (ADHDMdn:5.83%, ControlsMdn:44.2%, U= 160.5, p=0.022, r=-0.34). When ADHD patients integrated phonemes, reaction times were significantly longer (ADHDMdn:1260ms, ControlsMdn:582ms, U=41.0, p<.000, r= -0.56). In functional connectivity medio temporal gyrus (seed) was negatively associated with primary auditory cortex, inferior frontal gyrus, precentral gyrus, and fusiform gyrus. Conclusion: MI seems to be deficient for ADHD patients for stimuli that need top-down attentional allocation. This finding is supported by stronger functional connectivity from unimodal sensory areas to polymodal, MI convergence zones for complex stimuli in ADHD patients.

Keywords: attention-deficit hyperactivity disorder, audiovisual integration, McGurk-effect, resting-state functional connectivity

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Authors: Marcel Meinhardt, Manfred Keuser, Thomas Braml

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Cracking of reinforced concrete is a complex phenomenon induced by direct loads or restraints affecting reinforced concrete structures as soon as the tensile strength of the concrete is exceeded. Hence it is important to predict where cracks will be located and how they will propagate. The bond theory and the crack formulas in the actual design codes, for example, DIN EN 1992-1-1, are all based on the assumption that the reinforcement bars are embedded in homogeneous concrete without taking into account the influence of transverse reinforcement and the real stress situation. However, it can often be observed that real structures such as walls, slabs or beams show a crack spacing that is orientated to the transverse reinforcement bars or to the stirrups. In most Finite Element Analysis studies, the smeared crack approach is used for crack prediction. The disadvantage of this model is that the typical strain localization of a crack on element level can’t be seen. The crack propagation in concrete is a discontinuous process characterized by different factors such as the initial random distribution of defects or the scatter of material properties. Such behavior presupposes the elaboration of adequate models and methods of simulation because traditional mechanical approaches deal mainly with average material parameters. This paper concerned with the modelling of the initiation and the propagation of cracks in reinforced concrete structures considering the influence of transverse reinforcement and the real stress distribution in reinforced concrete (R/C) beams/plates in bending action. Therefore, a parameter study was carried out to investigate: (I) the influence of the transversal reinforcement to the stress distribution in concrete in bending mode and (II) the crack initiation in dependence of the diameter and distance of the transversal reinforcement to each other. The numerical investigations on the crack initiation and propagation were carried out with a 2D reinforced concrete structure subjected to quasi static loading and given boundary conditions. To model the uncertainty in the tensile strength of concrete in the Finite Element Analysis correlated normally and lognormally distributed random filed with different correlation lengths were generated. The paper also presents and discuss different methods to generate random fields, e.g. the Covariance Matrix Decomposition Method. For all computations, a plastic constitutive law with softening was used to model the crack initiation and the damage of the concrete in tension. It was found that the distributions of crack spacing and crack widths are highly dependent of the used random field. These distributions are validated to experimental studies on R/C panels which were carried out at the Laboratory for Structural Engineering at the University of the German Armed Forces in Munich. Also, a recommendation for parameters of the random field for realistic modelling the uncertainty of the tensile strength is given. The aim of this research was to show a method in which the localization of strains and cracks as well as the influence of transverse reinforcement on the crack initiation and propagation in Finite Element Analysis can be seen.

Keywords: crack initiation, crack modelling, crack propagation, cracks, numerical simulation, random fields, reinforced concrete, stochastic

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Authors: Marcel Verwey, Anna M. Joubert, Elsie M. Nolte, Wolfgang Dohle, Barry V. L. Potter, Anne E. Theron

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A tetrahydroisoquinolinone (THIQ) core can be used to mimic the A,B-ring of colchicine site-binding microtubule disruptors such as 2-methoxyestradiol in the design of anti-cancer agents. Steroidomimeric microtubule disruptors were synthesized by introducing C'2 and C'3 of the steroidal A-ring to C'6 and C'7 of the THIQ core and by introducing a decorated hydrogen bond acceptor motif projecting from the steroidal D-ring to N'2. For this in vitro study, four non-steroidal THIQ-based analogues were investigated and comparative studies were done between the non-sulphamoylated compound STX 3450 and the sulphamoylated compounds STX 2895, STX 3329 and STX 3451. The objective of this study was to investigate the modes of cell death induced by these four THIQ-based analogues in A549 lung carcinoma epithelial cells and metastatic breast adenocarcinoma MDA-MB-231 cells. Cytotoxicity studies to determine the half maximal growth inhibitory concentrations were done using spectrophotometric quantification via crystal violet staining and lactate dehydrogenase (LDH) assays. Microtubule integrity and morphologic changes of exposed cells were investigated using polarization-optical transmitted light differential interference contrast microscopy, transmission electron microscopy and confocal microscopy. Flow cytometric quantification was used to determine apoptosis induction and the effect that THIQ-based analogues have on cell cycle progression. Signal transduction pathways were elucidated by quantification of the mitochondrial membrane integrity, cytochrome c release and caspase 3, -6 and -8 activation. Induction of autophagic cell death by the THIQ-based analogues was investigated by morphological assessment of fluorescent monodansylcadaverine (MDC) staining of acidic vacuoles and by quantifying aggresome formation via flow cytometry. Results revealed that these non-steroidal microtubule disrupting analogues inhibited 50% of cell growth at nanomolar concentrations. Immunofluorescence microscopy indicated microtubule depolarization and the resultant mitotic arrest was further confirmed through cell cycle analysis. Apoptosis induction via the intrinsic pathway was observed due to depolarization of the mitochondrial membrane, induction of cytochrome c release as well as, caspase 3 activation. Potential involvement of programmed cell death type II was observed due to the presence of acidic vacuoles and aggresome formation. Necrotic cell death did not contribute significantly, indicated by stable LDH levels. This in vitro study revealed the induction of the intrinsic apoptotic pathway as well as possible involvement of autophagy after exposure to these THIQ-based analogues in both MDA-MB-231- and A549 cells. Further investigation of this series of anticancer drugs still needs to be conducted to elucidate the temporal, mechanistic and functional crosstalk mechanisms between the two observed programmed cell deaths pathways.

Keywords: apoptosis, autophagy, cancer, microtubule disruptor

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Authors: Alena Semeradtova, Marcel Stofik, Lucie Mareckova, Petr Maly, Ondrej Stanek, Jan Maly

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Recently, novel strategies based on so-called molecular evolution were shown to be effective for the production of various peptide ligand libraries with high affinities to molecular targets of interest comparable or even better than monoclonal antibodies. The major advantage of these peptide scaffolds is mainly their prevailing low molecular weight and simple structure. This study describes a new high-affinity binding molecules based immunesensor using a simple optical system for human serum albumin (HSA) detection as a model molecule. We present a comparison of two variants of recombinant binders based on albumin binding domain of the protein G (ABD) performed on micropatterned glass chip. Binding domains may be tailored to any specific target of interest by molecular evolution. Micropatterened glass chips were prepared using UV-photolithography on chromium sputtered glasses. Glass surface was modified by (3-aminopropyl)trietoxysilane and biotin-PEG-acid using EDC/NHS chemistry. Two variants of high-affinity binding molecules were used to detect target molecule. Firstly, a variant is based on ABD domain fused with TolA chain. This molecule is in vivo biotinylated and each molecule contains one molecule of biotin and one ABD domain. Secondly, the variant is ABD domain based on streptavidin molecule and contains four gaps for biotin and four ABD domains. These high-affinity molecules were immobilized to the chip surface via biotin-streptavidin chemistry. To eliminate nonspecific binding 1% bovine serum albumin (BSA) or 6% fetal bovine serum (FBS) were used in every step. For both variants range of measured concentrations of fluorescently labelled HSA was 0 – 30 µg/ml. As a control, we performed a simultaneous assay without high-affinity binding molecules. Fluorescent signal was measured using inverse fluorescent microscope Olympus IX 70 with COOL LED pE 4000 as a light source, related filters, and camera Retiga 2000R as a detector. The fluorescent signal from non-modified areas was substracted from the signal of the fluorescent areas. Results were presented in graphs showing the dependence of measured grayscale value on the log-scale of HSA concentration. For the TolA variant the limit of detection (LOD) of the optical immunosensor proposed in this study is calculated to be 0,20 µg/ml for HSA detection in 1% BSA and 0,24 µg/ml in 6% FBS. In the case of streptavidin-based molecule, it was 0,04 µg/ml and 0,07 µg/ml respectively. The dynamical range of the immunosensor was possible to estimate just in the case of TolA variant and it was calculated to be 0,49 – 3,75 µg/ml and 0,73-1,88 µg/ml respectively. In the case of the streptavidin-based the variant we didn´t reach the surface saturation even with the 480 ug/ml concentration and the upper value of dynamical range was not estimated. Lower value was calculated to be 0,14 µg/ml and 0,17 µg/ml respectively. Based on the obtained results, it´s clear that both variants are useful for creating the bio-recognizing layer on immunosensors. For this particular system, it is obvious that the variant based on streptavidin molecule is more useful for biosensing on glass planar surfaces. Immunosensors based on this variant would exhibit better limit of detection and wide dynamical range.

Keywords: high affinity binding molecules, human serum albumin, optical immunosensor, protein G, UV-photolitography

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Authors: Lacramioara Ochiuz, Manuela Hortolomei, Aurelia Vasile, Iulian Stoleriu, Marcel Popa, Cristian Peptu

Abstract:

Local antibiotherapy is one of the most effective acne therapies. Erythromycin (ER) is a macrolide antibiotic topically administered for over 30 years in the form of gel, ointment or hydroalcoholic solution for the acne therapy. The use of ER as a base for topical dosage forms raises some technological challenges due to the physicochemical properties of this substance. The main disadvantage of ER is the poor water solubility (2 mg/mL) that limits both formulation using hydrophilic bases and skin permeability. Cyclodextrins (CDs) are biocompatible cyclic oligomers of glucose, with hydrophobic core and hydrophilic exterior. CDs are used to improve the bioavailability of drugs by increasing their solubility and/or their rate of dissolution after including the poorly water soluble substances (such as ER) in the hydrophobic cavity of CDs. Adding CDs leads to the increase of solubility and improved stability of the drug substance, increased permeability of substances of low water solubility, decreased toxicity and even to active dose reduction as a result of increased bioavailability. CDs increase skin tolerability by reducing the irritant effect of certain substances. We have included ER to lactide modified β-cyclodextrin, in order to improve the therapeutic effect of topically administered ER. The aims of the present study were to synthesise and describe a new complex with prolonged release of ER with lactide modified β-cyclodextrin (CD-LA_E), to investigate the CD-LA_E complex by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR), to analyse the effect of semisolid base on the in vitro and ex vivo release characteristics of ER in the CD-LA_E complex by assessing the permeability coefficient and the release kinetics by fitting on mathematical models. SEM showed that, by complexation, ER changes its crystal structure and enters the amorphous phase. FTIR analysis has shown that certain specific bands of some groups in the ER structure move during the incapsulation process. The structure of the CD-LA_E complex has a molar ratio of 2.12 to 1 between lactide modified β-cyclodextrin and ER. The three semisolid bases (2% Carbopol, 13% Lutrol 127 and organogel based on Lutrol and isopropyl myristate) show a good capacity for incorporating the CD-LA_E complex, having a content of active ingredient ranging from 98.3% to 101.5% as compared to the declared value of 2% ER. The results of the in vitro dissolution test showed that the ER solubility was significantly increased by CDs incapsulation. The amount of ER released from the CD-LA_E gels was in the range of 76.23% to 89.01%, whereas gels based on ER released a maximum percentage of 26.01% ER. The ex vivo dissolution test confirms the increased ER solubility achieved by complexation, and supports the assumption that the use of this process might increase ER permeability. The highest permeability coefficient was obtained in ER released from gel based on 2% Carbopol: in vitro 33.33 μg/cm2/h, and ex vivo 26.82 μg/cm2/h, respectively. The release kinetics of complexed ER is performed by Fickian diffusion, according to the results obtained by fitting the data in the Korsmeyer-Peppas model.

Keywords: erythromycin, acne, lactide, cyclodextrin

Procedia PDF Downloads 266

Authors: Elsie M. Nolte, Annie M. Joubert, Roy Lakier, Maryke Etsebeth, Jolene M. Helena, Marcel Verwey, Laurence Lafanechere, Anne E. Theron

Abstract:

Treatment regimens for breast- and lung cancers may include both radiation- and chemotherapy. Ideally, a pharmaceutical agent which selectively sensitizes cancer cells to gamma (γ)-radiation would allow administration of lower doses of each modality, yielding synergistic anti-cancer benefits and lower metastasis occurrence, in addition to decreasing the side-effect profiles. A range of 2-methoxyestradiol (2-ME) analogues, namely 2-ethyl-3-O-sulphamoyl-estra-1,3,5 (10) 15-tetraene-3-ol-17one (ESE-15-one), 2-ethyl-3-O-sulphamoyl-estra-1,3,5(10),15-tetraen-17-ol (ESE-15-ol) and 2-ethyl-3-O-sulphamoyl-estra-1,3,5(10)16-tetraene (ESE-16) were in silico-designed by our laboratory, with the aim of improving the parent compound’s bioavailability in vivo. The main effect of these compounds is the disruption of microtubule dynamics with a resultant mitotic accumulation and induction of programmed cell death in various cancer cell lines. This in vitro study aimed to determine the cellular responses involved in the radiation sensitization effects of these analogues at low doses in breast- and lung cancer cell lines. The oestrogen receptor positive MCF-7-, oestrogen receptor negative MDA-MB-231- and triple negative BT-20 breast cancer cell lines as well as the A549 lung cancer cell line were used. The minimal compound- and radiation doses able to induce apoptosis were determined using annexin-V and cell cycle progression markers. These doses (cell line dependent) were used to pre-sensitize the cancer cells 24 hours prior to 6 gray (Gy) radiation. Experiments were conducted on samples exposed to the individual- as well as the combination treatment conditions in order to determine whether the combination treatment yielded an additive cell death response. Morphological studies included light-, fluorescence- and transmission electron microscopy. Apoptosis induction was determined by flow cytometry employing annexin V, cell cycle analysis, B-cell lymphoma 2 (Bcl-2) signalling, as well as reactive oxygen species (ROS) production. Clonogenic studies were performed by allowing colony formation for 10 days post radiation. Deoxyribonucleic acid (DNA) damage was quantified via γ-H2AX foci and micronuclei quantification. Amplification of the p53 signalling pathway was determined by western blot. Results indicated that exposing breast- and lung cancer cells to nanomolar concentrations of these analogues 24 hours prior to γ-radiation induced more cell death than the compound- and radiation treatments alone. Hypercondensed chromatin, decreased cell density, a damaged cytoskeleton and an increase in apoptotic body formation were observed in cells exposed to the combination treatment condition. An increased number of cells present in the sub-G1 phase as well as increased annexin-V staining, elevation of ROS formation and decreased Bcl-2 signalling confirmed the additive effect of the combination treatment. In addition, colony formation decreased significantly. p53 signalling pathways were significantly amplified in cells exposed to the analogues 24 hours prior to radiation, as was the amount of DNA damage. In conclusion, our results indicated that pre-treatment of breast- and lung cancer cells with low doses of 2-ME analogues sensitized breast- and lung cancer cells to γ-radiation and induced apoptosis more so than the individual treatments alone. Future studies will focus on the effect of the combination treatment on non-malignant cellular counterparts.

Keywords: cancer, microtubule dynamics, radiation therapy, radiosensitization

Procedia PDF Downloads 207