7,887 research outputs found

    Endogenous measures for contextualising large-scale social phenomena: a corpus-based method for mediated public discourse

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    This work presents an interdisciplinary methodology for developing endogenous measures of group membership through analysis of pervasive linguistic patterns in public discourse. Focusing on political discourse, this work critiques the conventional approach to the study of political participation, which is premised on decontextualised, exogenous measures to characterise groups. Considering the theoretical and empirical weaknesses of decontextualised approaches to large-scale social phenomena, this work suggests that contextualisation using endogenous measures might provide a complementary perspective to mitigate such weaknesses. This work develops a sociomaterial perspective on political participation in mediated discourse as affiliatory action performed through language. While the affiliatory function of language is often performed consciously (such as statements of identity), this work is concerned with unconscious features (such as patterns in lexis and grammar). This work argues that pervasive patterns in such features that emerge through socialisation are resistant to change and manipulation, and thus might serve as endogenous measures of sociopolitical contexts, and thus of groups. In terms of method, the work takes a corpus-based approach to the analysis of data from the Twitter messaging service whereby patterns in users’ speech are examined statistically in order to trace potential community membership. The method is applied in the US state of Michigan during the second half of 2018—6 November having been the date of midterm (i.e. non-Presidential) elections in the United States. The corpus is assembled from the original posts of 5,889 users, who are nominally geolocalised to 417 municipalities. These users are clustered according to pervasive language features. Comparing the linguistic clusters according to the municipalities they represent finds that there are regular sociodemographic differentials across clusters. This is understood as an indication of social structure, suggesting that endogenous measures derived from pervasive patterns in language may indeed offer a complementary, contextualised perspective on large-scale social phenomena

    Pollution-induced community tolerance in freshwater biofilms – from molecular mechanisms to loss of community functions

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    Exposure to herbicides poses a threat to aquatic biofilms by affecting their community structure, physiology and function. These changes render biofilms to become more tolerant, but on the downside community tolerance has ecologic costs. A concept that addresses induced community tolerance to a pollutant (PICT) was introduced by Blanck and WĂ€ngberg (1988). The basic principle of the concept is that microbial communities undergo pollution-induced succession when exposed to a pollutant over a long period of time, which changes communities structurally and functionally and enhancing tolerance to the pollutant exposure. However, the mechanisms of tolerance and the ecologic consequences were hardly studied up to date. This thesis addresses the structural and functional changes in biofilm communities and applies modern molecular methods to unravel molecular tolerance mechanisms. Two different freshwater biofilm communities were cultivated for a period of five weeks, with one of the communities being contaminated with 4 ÎŒg L-1 diuron. Subsequently, the communities were characterized for structural and functional differences, especially focusing on their crucial role of photosynthesis. The community structure of the autotrophs was assessed using HPLC-based pigment analysis and their functional alterations were investigated using Imaging-PAM fluorometry to study photosynthesis and community oxygen profiling to determine net primary production. Then, the molecular fingerprints of the communities were measured with meta-transcriptomics (RNA-Seq) and GC-based community metabolomics approaches and analyzed with respect to changes in their molecular functions. The communities were acute exposed to diuron for one hour in a dose-response design, to reveal a potential PICT and uncover related adaptation to diuron exposure. The combination of apical and molecular methods in a dose-response design enabled the linkage of functional effects of diuron exposure and underlying molecular mechanisms based on a sensitivity analysis. Chronic exposure to diuron impaired freshwater biofilms in their biomass accrual. The contaminated communities particularly lost autotrophic biomass, reflected by the decrease in specific chlorophyll a content. This loss was associated with a change in the molecular fingerprint of the communities, which substantiates structural and physiological changes. The decline in autotrophic biomass could be due to a primary loss of sensitive autotrophic organisms caused by the selection of better adapted species in the course of chronic exposure. Related to this hypothesis, an increase in diuron tolerance has been detected in the contaminated communities and molecular mechanisms facilitating tolerance have been found. It was shown that genes of the photosystem, reductive-pentose phosphate cycle and arginine metabolism were differentially expressed among the communities and that an increased amount of potential antioxidant degradation products was found in the contaminated communities. This led to the hypothesis that contaminated communities may have adapted to oxidative stress, making them less sensitive to diuron exposure. Moreover, the photosynthetic light harvesting complex was altered and the photoprotective xanthophyll cycle was increased in the contaminated communities. Despite these adaptation strategies, the loss of autotrophic biomass has been shown to impair primary production. This impairment persisted even under repeated short-term exposure, so that the tolerance mechanisms cannot safeguard primary production as a key function in aquatic systems.:1. The effect of chemicals on organisms and their functions .............................. 1 1.1 Welcome to the anthropocene .......................................................................... 1 1.2 From cellular stress responses to ecosystem resilience ................................... 3 1.2.1 The individual pursuit for homeostasis ....................................................... 3 1.2.2 Stability from diversity ................................................................................. 5 1.3 Community ecotoxicology - a step forward in monitoring the effects of chemical pollution? ................................................................................................................. 6 1.4 Functional ecotoxicological assessment of microbial communities ................... 9 1.5 Molecular tools – the key to a mechanistic understanding of stressor effects from a functional perspective in microbial communities? ...................................... 12 2. Aims and Hypothesis ......................................................................................... 14 2.1 Research question .......................................................................................... 14 2.2 Hypothesis and outline .................................................................................... 15 2.3 Experimental approach & concept .................................................................. 16 2.3.1 Aquatic freshwater biofilms as model community ..................................... 16 2.3.2 Diuron as model herbicide ........................................................................ 17 2.3.3 Experimental design ................................................................................. 18 3. Structural and physiological changes in microbial communities after chronic exposure - PICT and altered functional capacity ................................................. 21 3.1 Introduction ..................................................................................................... 21 3.2 Methods .......................................................................................................... 23 3.2.1 Biofilm cultivation ...................................................................................... 23 3.2.2 Dry weight and autotrophic index ............................................................. 23 3.2.4 Pigment analysis of periphyton ................................................................. 23 3.2.4.1 In-vivo pigment analysis for community characterization ....................... 24 3.2.4.2 In-vivo pigment analysis based on Imaging-PAM fluorometry ............... 24 3.2.4.3 In-vivo pigment fluorescence for tolerance detection ............................. 26 3.2.4.4 Ex-vivo pigment analysis by high-pressure liquid-chromatography ....... 27 3.2.5 Community oxygen metabolism measurements ....................................... 28 3.3 Results and discussion ................................................................................... 29 3.3.1 Comparison of the structural community parameters ............................... 29 3.3.2 Photosynthetic activity and primary production of the communities after selection phase ................................................................................................. 33 3.3.3 Acquisition of photosynthetic tolerance .................................................... 34 3.3.4 Primary production at exposure conditions ............................................... 36 3.3.5 Tolerance detection in primary production ................................................ 37 3.4 Summary and Conclusion ........................................................................... 40 4. Community gene expression analysis by meta-transcriptomics ................... 41 4.1 Introduction to meta-transcriptomics ............................................................... 41 4.2. Methods ......................................................................................................... 43 4.2.1 Sampling and RNA extraction................................................................... 43 4.2.2 RNA sequencing analysis ......................................................................... 44 4.2.3 Data assembly and processing................................................................. 45 4.2.4 Prioritization of contigs and annotation ..................................................... 47 4.2.5 Sensitivity analysis of biological processes .............................................. 48 4.3 Results and discussion ................................................................................... 48 4.3.1 Characterization of the meta-transcriptomic fingerprints .......................... 49 4.3.2 Insights into community stress response mechanisms using trend analysis (DRomic’s) ......................................................................................................... 51 4.3.3 Response pattern in the isoform PS genes .............................................. 63 4.5 Summary and conclusion ................................................................................ 65 5. Community metabolome analysis ..................................................................... 66 5.1 Introduction to community metabolomics ........................................................ 66 5.2 Methods .......................................................................................................... 68 5.2.1 Sampling, metabolite extraction and derivatisation................................... 68 5.2.2 GC-TOF-MS analysis ............................................................................... 69 5.2.3 Data processing and statistical analysis ................................................... 69 5.3 Results and discussion ................................................................................... 70 5.3.1 Characterization of the metabolic fingerprints .......................................... 70 5.3.2 Difference in the metabolic fingerprints .................................................... 71 5.3.3 Differential metabolic responses of the communities to short-term exposure of diuron ............................................................................................................ 73 5.4 Summary and conclusion ................................................................................ 78 6. Synthesis ............................................................................................................. 79 6.1 Approaches and challenges for linking molecular data to functional measurements ...................................................................................................... 79 6.2 Methods .......................................................................................................... 83 6.2.1 Summary on the data ............................................................................... 83 6.2.2 Aggregation of molecular data to index values (TELI and MELI) .............. 83 6.2.3 Functional annotation of contigs and metabolites using KEGG ................ 83 6.3 Results and discussion ................................................................................... 85 6.3.1 Results of aggregation techniques ........................................................... 85 6.3.2 Sensitivity analysis of the different molecular approaches and endpoints 86 6.3.3 Mechanistic view of the molecular stress responses based on KEGG functions ............................................................................................................ 89 6.4 Consolidation of the results – holistic interpretation and discussion ............... 93 6.4.1 Adaptation to chronic diuron exposure - from molecular changes to community effects.............................................................................................. 93 6.4.2 Assessment of the ecological costs of Pollution-induced community tolerance based on primary production ............................................................. 94 6.5 Outlook ............................................................................................................ 9

    Identifizierung prÀdiktiver und prognostischer Biomarker in unterschiedlichen Tumorkompartimenten des ösophagealen Adenokarzinoms

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    Das ösophageale Adenokarzinom zeigt eine global steigende Inzidenz und hat mit einer 5-Jahres-Überlebensrate von weniger als 25% eine schlechte Prognose. Personalisierte TherapieansĂ€tze sind selten und prognostische/prĂ€diktive Biomarker des Tumormikromilieus sind unzureichend charakterisiert. Die kumulative Promotion nĂ€hert sich dieser Problematik in drei unterschiedlichen Schwerpunkten. 1. Zur Identifizierung Kompartiment-spezifischer Biomarker wurde eine Methode entwickelt, welche als kostengĂŒnstige Alternative zum sc-Seq Expressionsprofile individueller Zelltypen generiert. Dabei erfolgt die Extraktion der RNA nicht aus Einzelzellen, sondern aus flowzytometrisch-getrennten Zellkompartimenten. Die Separation der Proben in Epithelzellen, Immunzellen und Fibroblasten wurde durch verschiedene Verfahren validiert und eine suffiziente Ausbeute an RNA auch fĂŒr kleine Gewebemengen gezeigt. 2. Biomarker des Immunzellkompartiments als therapeutische Angriffspunkte wurden in einem Patientenkollektiv von bis zu 551 Patienten auf ihre Bedeutung beim EAC ĂŒberprĂŒft. Es zeigte sich eine Expression der Immuncheckpoints LAG3, VISTA und IDO auf TILs durch IHC und RNA-Sonden basierte Verfahren in einem relevanten Anteil (LAG3: 11,4%, VISTA: 29%, IDO: 52,6%). Es konnte eine prognostisch gĂŒnstige Bedeutung der VISTA, LAG3 und IDO Expression gezeigt werden. Durch den Vergleich von Genexpressionsprofilen aus therapienaiven und vorbehandelten Tumoren konnte zudem ein immunsuppressiver Effekt von neoadjuvanten Therapiekonzepten auf das Tumormikromilieu des EACs gezeigt werden. Dabei kam es zur verminderten Expression von Checkpoints und Anzahl TILs nach (Radio-) Chemotherapie. 3. Im Tumorzellkompartiment wurde die Rolle von Amplifikationen in ErbB-Rezeptor abhĂ€ngigen Signalwegen durch FISH-Technik und Immunhistochemie evaluiert. Es fanden sich KRAS Amplifikationen in 17,1%, PIK3CA Amplifikationen in 5% sowie eine HER2/neu-Überexpression in 14,9% der untersuchten Tumore

    Victims' Access to Justice in Trinidad and Tobago: An exploratory study of experiences and challenges of accessing criminal justice in a post-colonial society

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    This thesis investigates victims' access to justice in Trinidad and Tobago, using their own narratives. It seeks to capture how their experiences affected their identities as victims and citizens, alongside their perceptions of legitimacy regarding the criminal justice system. While there have been some reforms in the administration of criminal justice in Trinidad and Tobago, such reforms have not focused on victims' accessibility to the justice system. Using grounded theory methodology, qualitative data was collected through 31 in-depth interviews with victims and victim advocates. The analysis found that victims experienced interpersonal, structural, and systemic barriers at varying levels throughout the criminal justice system, which manifested as institutionalized secondary victimization, silencing and inequality. This thesis argues that such experiences not only served to appropriate conflict but demonstrates that access is often given in a very narrow sense. Furthermore, it shows a failure to encompass access to justice as appropriated conflicts are left to stagnate in the system as there is often very little resolution. Adopting a postcolonial lens to analyse victims' experiences, the analysis identified othering practices that served to institutionalize the vulnerability and powerlessness associated with victim identities. Here, it is argued that these othering practices also affected the rights consciousness of victims, delegitimating their identities as citizens. Moreover, as a result of their experiences, victims had mixed perceptions of the justice system. It is argued that while the system is a legitimate authority victims' endorsement of the system is questionable, therefore victims' experiences suggest that there is a reinforcement of the system's legal hegemony. The findings suggest that within the legal system of Trinidad and Tobago, legacies of colonialism shape the postcolonial present as the psychology and inequalities of the past are present in the interactions and processes of justice. These findings are relevant for policymakers in Trinidad and Tobago and other regions. From this study it is recognized that, to improve access to justice for victims, there needs to be a move towards victim empowerment that promotes resilience and enhances social capital. Going forward it is noted that there is a need for further research

    Targeting Fusion Proteins of HIV-1 and SARS-CoV-2

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    Viruses are disease-causing pathogenic agents that require host cells to replicate. Fusion of host and viral membranes is critical for the lifecycle of enveloped viruses. Studying viral fusion proteins can allow us to better understand how they shape immune responses and inform the design of therapeutics such as drugs, monoclonal antibodies, and vaccines. This thesis discusses two approaches to targeting two fusion proteins: Env from HIV-1 and S from SARS-CoV-2. The first chapter of this thesis is an introduction to viruses with a specific focus on HIV-1 CD4 mimetic drugs and antibodies against SARS-CoV-2. It discusses the architecture of these viruses and fusion proteins and how small molecules, peptides, and antibodies can target these proteins successfully to treat and prevent disease. In addition, a brief overview is included of the techniques involved in structural biology and how it has informed the study of viruses. For the interested reader, chapter 2 contains a review article that serves as a more in-depth introduction for both viruses as well as how the use of structural biology has informed the study of viral surface proteins and neutralizing antibody responses to them. The subsequent chapters provide a body of work divided into two parts. The first part in chapter 3 involves a study on conformational changes induced in the HIV-1 Env protein by CD4-mimemtic drugs using single particle cryo-EM. The second part encompassing chapters 4 and 5 includes two studies on antibodies isolated from convalescent COVID-19 donors. The former involves classification of antibody responses to the SARS-CoV-2 S receptor-binding domain (RBD). The latter discusses an anti-RBD antibody class that binds to a conserved epitope on the RBD and shows cross-binding and cross-neutralization to other coronaviruses in the sarbecovirus subgenus.</p

    Towards personalized immunotherapy : development of in vitro models for imaging natural killer cell behavior in the tumor microenvironment

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    Tremendous advances in the tumor immunology field have transformed immunotherapy from a promising approach to a standard clinical practice. However, a subset of cancer patients is non-responsive to immunotherapy. More research is therefore needed to understand the mechanisms underlying tumor resistance to immunotherapeutic treatments. The aim of this doctoral work was to develop new tools to study the mechanisms of cancer immunosurveillance and to test immunotherapeutic treatments in vitro. In this thesis, I describe the methods developed, and I discuss the main biological findings obtained by using these methods. The thesis is organized as follows. A short historical background of immunotherapy is provided in Chapter 1. Chapter 2 describes the principles of NK cell-mediated cancer immunosurveillance, and provides an overview on rare cancers, mainly focusing on sarcoma. The research aims are listed in Chapter 3. In Chapter 4, I describe the cell culture methods and cell analysis techniques relevant for my doctoral work. In Chapter 5, I describe the methods we developed to culture tumor spheroids in vitro using ultrasonic standing waves in microwell chips, focusing on the theory, design, and applications. Chapter 6 and Chapter 7 focus on the biological findings obtained using our platform in combination with traditional immunological methods, followed by future implementations discussed in Chapter 8. The constituent papers are provided at the end of the thesis. In Paper I, we combined the use of the microwell chip, ultrasonic standing waves and a protein-repellent polymer coating to enable the production of spheroids from multiple cell types. In absence of cell adhesion to the chip, spheroids could be collected and further analyzed by off-the-chip techniques. In Paper II, we designed a novel multichambered microwell chip to perform multiplexed fluorescence screening of two- or three-dimensional cell cultures. The platform allows the direct assessment of drug or immune cell cytotoxic efficacy, making it a promising tool for individualized cytotoxicity tests for personalized medicine. In Paper III, we investigate the function of PVR receptors in NK cells interacting with renal carcinoma spheroids, and the impact of PVR in NK cell-based cellular immunotherapy. We demonstrated that variations in PVR expression are primarily recognized by the inhibitory receptor TIGIT, while DNAM-1 strongly contributes to NK cell activation mainly through PVR-independent mechanisms. We performed NK cell-based cytotoxicity assays against renal carcinoma spheroids in the microwell chip. Anti-TIGIT treatment was effective only for TIGIThigh NK cells both when used as monotherapy or in combination with other drugs, suggesting that only a fraction of patients might respond to anti-TIGIT therapy. In Paper IV, a similar approach was used with primary sarcomas. We cultured patient-derived sarcoma spheroids and tested NK cell-based immunotherapy in the microwell chip, either alone or in combination with antibody therapy, and we identified promising treatment combinations. In Paper V, we applied the use of expansion microscopy to visualize NK cells infiltrating renal carcinoma spheroids. In conclusion, our multi-disciplinary work shows the development of new imaging-based platform and its use to study the mechanisms of NK cell-mediated tumor surveillance and for personalized therapy

    Towards a non-equilibrium thermodynamic theory of ecosystem assembly and development

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    Non-equilibrium thermodynamics has had a significant historic influence on the development of theoretical ecology, even informing the very concept of an ecosystem. Much of this influence has manifested as proposed extremal principles. These principles hold that systems will tend to maximise certain thermodynamic quantities, subject to the other constraints they operate under. A particularly notable extremal principle is the maximum entropy production principle (MaxEPP); that systems maximise their rate of entropy production. However, these principles are not robustly based in physical theory, and suffer from treating complex ecosystems in an extremely coarse manner. To address this gap, this thesis derives a limited but physically justified extremal principle, as well as carrying out a detailed investigation of the impact of non-equilibrium thermodynamic constraints on the assembly of microbial communities. The extremal principle we obtain pertains to the switching between states in simple bistable systems, with switching paths that generate more entropy being favoured. Our detailed investigation into microbial communities involved developing a novel thermodynamic microbial community model, using which we found the rate of ecosystem development to be set by the availability of free-energy. Further investigation was carried out using this model, demonstrating the way that trade-offs emerging from fundamental thermodynamic constraints impact the dynamics of assembling microbial communities. Taken together our results demonstrate that theory can be developed from non-equilibrium thermodynamics, that is both ecologically relevant and physically well grounded. We find that broad extremal principles are unlikely to be obtained, absent significant advances in the field of stochastic thermodynamics, limiting their applicability to ecology. However, we find that detailed consideration of the non-equilibrium thermodynamic mechanisms that impact microbial communities can broaden our understanding of their assembly and functioning.Open Acces

    Contemporary, decadal, and millennial-scale permafrost- and vegetation dynamics and carbon release in an alpine region of Jotunheimen, Norway

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    Climatic warming in northern alpine regions facilitates the thawing of permafrost, the associated release of soil carbon into the atmosphere, and the altitudinal shifts in vegetation patterns. Here, a multi-disciplinary approach is adopted to investigate the response of an alpine permafrost landscape (Jotunheimen, Norway, with focus on GaldhĂžpiggen) to climatic changes over long- to medium timescales. First, a gas analyser is used to explore how ecosystem respiration is affected by ecosystem (soil and vegetation) and geomorphological (cryogenic disturbance) factors during the peak growing season. A palaeoecological record is then analysed to infer the past dynamics of the alpine tree lines and the lower limit of permafrost on GaldhĂžpiggen over the millennial- and centennial scales. Finally, remotely sensed satellite imagery is combined with observed air temperatures to create a model that provides an estimation of land surface temperatures over the past six decades. The model is then used to predict surface ‘greenness’ over the same period. Palynological evidence from GaldhĂžpiggen indicates that the altitudinal limits of alpine tree lines have shifted by hundreds of metres in response to climatic changes over the millennial scale. Since 1957, the model predictions indicate substantial increases in land surface temperatures and growing season surface ‘greenness’ (i.e., vegetation abundance) in Jotunheimen, but the change has not been spatially uniform. The highest increases were recorded over the low- and mid-alpine heaths above the tree line (1050-1500 m a.s.l.), which was attributed to increased shrub cover. This trend could facilitate carbon release from the ground, as peak growing season ecosystem respiration was found to be most strongly controlled by soil microclimate and plant growth forms. The likely future scenario in response to warming in Jotunheimen will be continued permafrost degradation, with higher altitudes (≄1500 m a.s.l.) experiencing decreased cryoturbation, increased shrub encroachment and higher surface CO2 emissions

    Statistical Learning for Gene Expression Biomarker Detection in Neurodegenerative Diseases

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    In this work, statistical learning approaches are used to detect biomarkers for neurodegenerative diseases (NDs). NDs are becoming increasingly prevalent as populations age, making understanding of disease and identification of biomarkers progressively important for facilitating early diagnosis and the screening of individuals for clinical trials. Advancements in gene expression profiling has enabled the exploration of disease biomarkers at an unprecedented scale. The work presented here demonstrates the value of gene expression data in understanding the underlying processes and detection of biomarkers of NDs. The value of novel approaches to previously collected -omics data is shown and it is demonstrated that new therapeutic targets can be identified. Additionally, the importance of meta-analysis to improve power of multiple small studies is demonstrated. The value of blood transcriptomics data is shown in applications to researching NDs to understand underlying processes using network analysis and a novel hub detection method. Finally, after demonstrating the value of blood gene expression data for investigating NDs, a combination of feature selection and classification algorithms were used to identify novel accurate biomarker signatures for the diagnosis and prognosis of Parkinson’s disease (PD) and Alzheimer’s disease (AD). Additionally, the use of feature pools based on previous knowledge of disease and the viability of neural networks in dimensionality reduction and biomarker detection is demonstrated and discussed. In summary, gene expression data is shown to be valuable for the investigation of ND and novel gene biomarker signatures for the diagnosis and prognosis of PD and AD
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