Dissertations / Theses on the topic 'Heterogeneous disease'

Functional genomics approaches offer an unparalleled window into cellular biology. It is particularly challenging to apply these methods to heterogeneous tissues, in which multiple cell types or transcriptomic identities co-exist. I applied single cell and bulk functional genomics approaches in an attempt to understand aspects of the biology underlying cerebral cortex and thymic epithelial cells. These are both extremely heterogeneous tissues but the nature of the heterogeneity differs: in cortex there are a large number of different cell types, whereas in thymic epithelial cells, despite a limited number of cell types, individual cells express virtually every gene in the transcriptome. In Chapter 1, I provide a general overview of the currently available functional genomics tools and briefly review relevant aspects of cortical and thymic biology. The first three results chapters examine functional genomics approaches to cerebral cortex. Chapter 2 details the application of single cell functional genomics techniques to examine the extent to which stem cell-derived cortical neurons resemble primary human cortical neurons. In Chapter 3, I use RNA sequencing in cortical neurons derived from wild-type or PSEN1 mutant stem cells in an attempt to identify an Alzheimer's disease-relevant gene signature and to assess how much variability there is between batches of stem cell differentiation. Chapter 4 applies DNase-seq footprinting to primary brain tissue in order to understand how common variants associated with altered gene expression or susceptibility to neurological disease might exert their effects. The last two results chapters focus on thymic epithelial cells. In Chapter 5, I integrate multiple functional genomics datasets to identify a set of genes targeted by Foxn1, a criticial transcription factor in thymus development and function. Chapter 6 use single cell RNA-seq from mature medullary thymic epithelial cells to gain an insight into how promiscuous gene expression in the thymus is orchestrated at the level of individual cells. Finally, in Chapter 7, I summarise the implications of my findings for cortical and thymic biology and speculate on the potential directions that functional genomics may take in the future to better understand heterogeneous tissues.

The application of spatial methods to epidemic estimation and prediction problems is a vibrant and active area of research. In many cases, however, well thought out and laboratory supported models for epidemic patterns may be easy to specify but extremely difficult to fit efficiently. While this problem exists in many scientific disciplines, epidemic modeling is particularly prone to this challenge due to the rate at which the problem scope grows as a function of the size of the spatial and temporal domains involved. An additional barrier to widespread use of spatiotemporal epidemic models is the lack of user friendly software packages capable of fitting them. In particular, compartmental epidemic models are easy to understand, but in most cases difficult to fit. This class of epidemic models describes a set of states, or compartments, which captures the disease progression in a population. This dissertation attempts to expand the problem scope to which spatio-temporal compartmental epidemic models are applicable both computationally and practically. In particular, a general family of spatially heterogeneous SEIRS models is developed alongside a software library with the dual goals of high computational performance and ease of use in fitting models in this class. We emphasize the task of model specification, and develop a framework describing the components of epidemic behavior. In addition, we establish methods to estimate and interpret reproductive numbers, which are of fundamental importance to the study of infectious disease. Finally, we demonstrate the application of these techniques both under simulation, and in the context of a diverse set of real diseases, including Ebola Virus Disease, Smallpox, Methicillin-resistant Staphylococcus aureus, and Influenza.

Thesis (Ph.D.)--Indiana University, School of Health, Physical Education, and Recreation, 2006.
"Title from dissertation home page (viewed July 2, 2007)." Source: Dissertation Abstracts International, Volume: 67-06, Section: B, page: 3087. Advisers: David Lohrmann; Erick Janssen.

Thesis: M. Eng., Massachusetts Institute of Technology, Department of Biological Engineering, May, 2020
Cataloged from PDF version of thesis.
Includes bibliographical references (pages. 85-97).
Tau neurofibrillary tangles or aggregates are a common neuropathological feature found in a number of neurodegenerative conditions, including Alzheimer's disease. Understanding the kinetics of this aggregate build up, how it varies across patients, and how aggregation might be influenced by intracellular pathways is critical for both a deeper knowledge of these disorders as well as identification of potential therapeutic targets. To this end, I employed an in vitro tau aggregation assay to study the kinetics of tau aggregation as it relates to aggregates in sporadic Alzheimer's disease. I found that the formation of aggregates was a logistic process, with a lag phase, an exponential rise phase, and a plateau phase. Aggregation kinetics varied significantly between different cases of sporadic Alzheimer's disease, paralleling the heterogeneity that is observed in the clinical presentation of Alzheimer's disease. Likewise, I found that inhibition of intracellular pathways of macroautophagy and endosomal microautopahgy heterogeneously increased tau aggregation and changed tau aggregation kinetics, dependent upon the case of Alzheimer's disease. These results inform that tau aggregates vary significantly not just between disorders, but even within disorders, and that protein degradation pathways uniquely process aggregates, perhaps potentiated by further molecular differences in aggregate structure or composition.
by Tarun Kamath.
M. Eng.
M.Eng. Massachusetts Institute of Technology, Department of Biological Engineering

The history of mitigation programs to contain vector-borne diseases is a story of successes and failures. Due to the complex interplay among multiple factors that determine disease dynamics, the general principles for timely and specific intervention for incidence reduction or eradication of life-threatening diseases has yet to be determined. This research discusses computational methods developed to assist in the understanding of complex relationships affecting vector-borne disease dynamics. A computational framework to assist public health practitioners with exploring the dynamics of vector-borne diseases, such as malaria and dengue in homogenous and heterogeneous populations, has been conceived, designed, and implemented. The framework integrates a stochastic computational model of interactions to simulate horizontal disease transmission. The intent of the computational modeling has been the integration of stochasticity during simulation of the disease progression while reducing the number of necessary interactions to simulate a disease outbreak. While there are improvements in the computational time reducing the number of interactions needed for simulating disease dynamics, the realization of interactions can remain computationally expensive. Using multi-threading technology to improve performance upon the original computational model, multi-threading experimental results have been tested and reported. In addition, to the contact model, the modeling of biological processes specific to the corresponding pathogen-carrier vector to increase the specificity of the vector-borne disease has been integrated. Last, automation for requesting, retrieving, parsing, and storing specific weather data and geospatial information from federal agencies to study the differences between homogenous and heterogeneous populations has been implemented.

Infectious disease transmission is driven by a complex suite of drivers with behavior and landscape dynamics contributing to epidemics across host-pathogen systems. However, our understanding of the interaction between landscape, behavior, and infectious disease remains limited. In the banded mongoose (Mungos mungo), a novel tuberculosis pathogen, Mycobacterium mungi, has emerged in Northern Botswana that is transmitted through olfactory communication behaviors. Using this host-pathogen system, this thesis explores the influence of various land use areas along the human-wildlife interface on animal behavior, and ultimately, pathogen transmission potential. Using behavior data from remote sensing camera traps, a generalized linear mixed model identified vigilance behavior, land use, and their interaction as important factors in predicting olfactory behavior. Cluster and Classification and Regression Tree (CART) analysis of active den sites (n= 308, across 23 troops) identified the important characteristics of dens across land use areas. In human-modified environments, man-made den sites persisted longer than did natural dens which became unsuitable through environmental processes (e.g., collapse). We also document the occurrence of nighttime activity for this species, perceived to be strictly diurnal. These data provide information critical to the development of robust computational models and underscore the importance of both landscape and behavior in accurately predicting and managing infectious disease outbreaks.
M. S.

Organisms are continuously exposed to heterogeneous micro- and macro-parasitic species, hence simultaneous infections often occur in wild and farm environments. This joint project aimed to develop a co-infection model between chronic and acute infections, evaluating their impact on the fish immune system. Proliferative Kidney Disease was studied on farmed rainbow and brown trout during natural seasonal outbreaks, using a parasite gene (Tetracapsuloides bryosalmonae RPL18) as a proxy for assessment of parasite burden. In hosts with elevated susceptibility PKD pathogenesis was shaped by an anti-inflammatory phenotype, a profound B cell/antibody response and dysregulated TH cell-like activity. Pathogen-free brown trout were exposed to Viral Haemorrhagic Septicaemia Virus (comparatively using European VHSV-Ia and North American VHSV-IVb strains) or to the bacterium Yersinia ruckeri. This European native species was highly resistant to the VHSV-IVb strain, which was undetectable in internal organs despite raising a strong antiviral and mucosal immune response. Following VHS and Yersiniosis infection, haemo-lymphopoietic organs were screened by RT-qPCR to assess the specific pathogen burdens and characterise the immune responses elicited. Transcription patterns were analysed for Interferons, CXC chemokines, SOCS (potential disease resistance biomarkers) and genes of the PACAP system. Lastly, PKD-infected brown trout were co-infected with VHSV-Ia, resulting in typical lesions while showing reduced and delayed mortality. PKD+/VHS+ fish were identified by RT-qPCR and histopathology screening. Pro-inflammatory and antimicrobial peptide genes were modulated following virus co-infection when compared to fish with single infection, with an earlier activation of cellular and humoral responses, and a stronger up-regulation of TH1 and antiviral genes. Oligonucleotide microarrays were used to assess the broader immune gene transcription modulation between single- and co-infected fish. Overall, the results suggest that the immune response of brown trout might be enhanced during the PKD/VHS co-infection.

Master of Science
Department of Electrical and Computer Engineering
Faryad Darabi Sahneh
Recent experience of Ebola outbreak of 2014 highlighted the importance of immediate response to impede Ebola transmission at its very early stage. To this aim, efficient and effective allocation of limited resources is crucial. Among standard interventions is the practice of following up with physical contacts of individuals diagnosed with Ebola virus disease -- known as contact tracing. In an effort to objectively understand the effect of possible contact tracing protocols, we explicitly develop a model of Ebola transmission incorporating contact tracing. Our modeling framework has several features to suit early–stage Ebola transmission: 1) the network model is patient–centric because when number of infected cases are small only the myopic networks of infected individuals matter and the rest of possible social contacts are irrelevant, 2) the Ebola disease model is individual–based and stochastic because at the early stages of spread, random fluctuations are significant and must be captured appropriately, 3) the contact tracing model is parameterizable to analyze the impact of critical aspects of contact tracing protocols. Notably, we propose an activity driven network approach to contact tracing, and develop a Monte-Carlo method to compute the basic reproductive number of the disease spread in different scenarios. Exhaustive simulation experiments suggest that while contact tracing is important in stopping the Ebola spread, it does not need to be done too urgently. This result is due to rather long incubation period of Ebola disease infection. However, immediate hospitalization of infected cases is crucial and requires the most attention and resource allocation. Moreover, to investigate the impact of mitigation strategies in the 2014 Ebola outbreak, we consider reported data in Guinea, one the three West Africa countries that had experienced the Ebola virus disease outbreak. We formulate a multivariate sequential Monte Carlo filter that utilizes mechanistic models for Ebola virus propagation to simultaneously estimate the disease progression states and the model parameters according to reported incidence data streams. This method has the advantage of performing the inference online as the new data becomes available and estimating the evolution of the basic reproductive ratio R₀(t) throughout the Ebola outbreak. Our analysis identifies a peak in the basic reproductive ratio close to the time of Ebola cases reports in Europe and the USA.

Rupture of atherosclerotic plaque is one of the primary causes of death due to cardiovascular disease. The factors directing plaque progression to instability are poorly understood. It is well-known that arteries respond to changes in mechanical stress by remodeling, and that remodeling is mediated by the inflammatory response. Studies have shown that both mechanical stress and markers of inflammation are increased in the fibrous cap and shoulder regions of plaque, where rupture most often occurs. In this study we hypothesized that there are spatial relationships between the local mechanical environment and expression of markers of inflammation in atherosclerosis, and that these relationships are plaque-progression dependent. To test these hypotheses, we analyzed cross-sections at intervals along the length of human coronary atherosclerotic arteries. For each cross-section, a heterogeneous finite element model was developed to determine the spatial distribution of stress. In addition, novel techniques for quantifying inflammatory markers at high spatial resolution were used to determine the distributions of inflammatory markers. The distributions of stress and five markers of inflammation activated NF-kB, macrophages, MMP-1, nitrotyrosine, and microvessels - were then compared to determine whether spatial relationships exists. We demonstrated that the probability of activated NF-kB expression increases monotonically with increasing stress in all stages of plaque progression. This indicates that the relationship between mechanical stress and NF-kB activation is a player throughout the disease process. We found that the relationship between mechanical stress and macrophages is highly dependent on the state of plaque progression. In intermediate stages of progression macrophages increase with moderate stress but drop off again at very high stresses, while in the advanced stage macrophages continue to increase monotonically with stress. We found that MMP1 increases with stress in stages of progression where active remodeling is occurring, but decreases with stress in mature stable plaque. We found no relationship between mechanical stress and nitrotyrosine expression or microvessels. Taken together, these results support the role of mechanical stress in instigating and maintaining the inflammatory response, and help explain how mechanical input is able to direct the complex biological changes involved in remodeling.

L’objectif de ma thèse est de proposer des solutions contre la propagation des maladies infectieuses dans des cas précis, en tenant compte de l'évolution des contacts entre les hôtes. Ce travail porte en particulier sur la détermination du seuil épidémique, un indicateur clé du risque épidémique. Il exploite et étend un formalisme mathématique issu de la théorie des réseaux, qui permet de déterminer le seuil épidémique dans des situations réelles, pour en dégager des mesures de santé publique. Un premier projet met en lumière des facteurs à l'origine de la persistance de la brucellose bovine en Italie en dépit des mesures d'éradication en place. L'approche théorique permet de calculer le seuil épidémique dans chaque région du pays à l'aide de données exhaustives sur les déplacements de bovins entre les exploitations italiennes sur plusieurs années, ainsi que des relevés datés de flambées épidémiques. Est ensuite présentée une extension du formalisme qui prend en compte différentes durées moyennes d’infection dans le calcul du seuil épidémique. Ce travail montre dans différents contextes épidémiologiques comment l’hypothèse classique selon laquelle la durée moyenne d’infection est homogène peut biaiser l’estimation du risque épidémique. Cette méthode permet également d'identifier les hôtes d'une population qui sont principalement responsables du risque épidémique global
My doctoral thesis aims to propose solutions against the spread of infectious diseases in specific contexts, taking into account how host contacts evolve in time using a temporal network representation. It focuses on the determination of the epidemic threshold, a key indicator of the epidemic risk. By leveraging and extending a mathematical formalism from network theory, this work enables the computation of the epidemic threshold in real situations in order to identify public health measures. A first project addresses the persistence of bovine brucellosis in Italy despite the existing eradication measures. Using comprehensive data on cattle movements between Italian farms over several years, as well as time-stamped outbreak records, the epidemic threshold computation in each region of the country provides information on regions vulnerability and proposes factors that may explain disease persistence. An extension of the formalism is then presented, including heterogeneous average infectious periods in the epidemic threshold computation. This work shows in different epidemiological contexts how the classical assumption that the average infectious period is the same for all hosts in a population may bias epidemic risk assessments. This method also identifies the hosts in a population that are primarily responsible for the global epidemic risk

The spread of infectious diseases has been a public concern throughout human history. Historic recorded data has reported the severity of infectious disease epidemics in different ages. Ancient Greek physician Hippocrates was the first to analyze the correlation between diseases and their environment. Nowadays, health authorities are in charge of planning strategies that guarantee the welfare of citizens. The simulation of contagion scenarios contributes to the understanding of the epidemic behavior of diseases. Computational models facilitate the study of epidemics by integrating disease and population data to the simulation. The use of detailed demographic and geographic characteristics allows researchers to construct complex models that better resemble reality and the integration of these attributes permits us to understand the rules of interaction. The interaction of individuals with similar characteristics forms synthetic structures that depict clusters of interaction. The synthetic environments facilitate the study of the spread of infectious diseases in diverse scenarios. The characteristics of the population and the disease concurrently affect the local and global epidemic progression. Every cluster’ epidemic behavior constitutes the global epidemic for a clustered population. By understanding the correlation between structured populations and the spread of a disease, current dissertation research makes possible to identify risk groups of specific characteristics and devise containment strategies that facilitate health authorities to improve mitigation strategies.

La thèse est composée de trois articles: le premier constate pour la Colombie, que la quantité des travailleurs qualifiés et le commerce international entraînent un changement technique biaisé vers les qualifiés, augmentant l’inégalité des salaires, bien que cet effet soit compensé par l’emploi temporaire. Le deuxième analyse un modèle des entreprises hétérogènes formelles et informelles, avec plein emploi, montrant qu’une politique d’ouverture augmente la quantité des entreprises informelles et réduit sa productivité moyenne, diminuant le bien-être. Cependant, forcer les informelles à devenir formelles augmente les salaires moyens et le bien-être. Les estimations Diff in Diff dans le troisième article, présentent l'impact du boom pétrolier 2003-2013 sur les pays touchés et non touchés par la maladie hollandaise. Pour les premières, les flux commerciaux internationaux augmentent bien que l'agriculture dans une moindre mesure, alors que le chômage et le travail informel diminuent
I study the relationship between international trade and labor markets in three papers. In the first one, I find for the Colombian case, that together, the sector skill intensity and the international trade bring about more skill-biased technical change, increasing wage inequality, though such an effect is offset using temporary workers. In the second one, the analysis of a trade model with formal and informal heterogeneous firms, under full employment, shows that an openness policy decreases the average productivity of informal firms while makes formal to become informal, worsening welfare. However, forcing informal firms to become formal, increases average wages and raises welfare. In the third one, Diff in Diff estimates presents the impact of the 2003-2013 oil prices boom, on countries affected and not affected by the Dutch disease. In the former group, international trade flow increases although agriculture at a lower magnitude, while unemployment and informal labor decrease

La gestion des maladies infectieuses dans la faune sauvage se heurte à de nombreuses limites, et le développement de stratégies efficaces représente un défi de taille. Pour atteindre cet objectif, une compréhension fine des facteurs influençant la transmission et la persistance de l’infection est nécessaire. Parmi ces facteurs, l’hétérogénéité de transmission est une caractéristique importante des populations sauvages. En effet, la diversité des comportements, des structures sociales et spatiales, ou encore des espèces peut conduire à des contributions très variables au nombre de nouvelles infections. Par conséquent, quantifier l’hétérogénéité de transmission pourrait permettre d’améliorer l’efficacité des mesures de gestion sanitaire dans la faune sauvage, en ciblant les individus ou les unités de population qui sont responsables de la majorité des évènements de transmission. L’objectif de cette thèse était d’améliorer les connaissances sur la gestion des maladies infectieuses dans des populations sauvages hétérogènes, en utilisant la brucellose à Brucella melitensis dans une population de bouquetin des Alpes (Capra ibex) comme modèle d’étude. En effet, la biologie de la brucellose et l’écologie de l’espèce hôte se prêtent bien à l’existence et donc à l’étude d’une hétérogénéité de transmission à différentes échelles. A l’aide de cultures bactériennes, nous avons tout d’abord montré que seulement 58 % des individus séropositifs sont à risque d’excréter la brucellose, et que ce risque diminue avec l’âge. Ensuite, mettant à profit l’existence d’informations détaillées sur la dynamique de population et le comportement du bouquetin, et de données épidémiologiques dans la population d’étude, nous avons développé un modèle individu-centré afin de quantifier l’hétérogénéité individuelle et spatiale de la transmission. Nous avons démontré que la transmission de la brucellose était hétérogène entre individus, les femelles provoquant environ 90% des nouvelles infections, et entre unités spatiales, plus de 80% des cas de transmission ayant lieu dans les trois sous-unités socio-spatiales qui forment la zone cœur du massif. Nous avons également estimé l’évolution temporelle de la séroprévalence et de la force d’infection, en utilisant différents modèles statistiques. Les résultats suggèrent que l’importante opération de capture menée en 2015, avec test systématique et élimination des individus séropositifs, a permis de diminuer la transmission de la brucellose dans la population. Sur la base de l’ensemble de ces résultats, nous avons évalués une série de stratégies de gestion sanitaire qui pourraient être utilisées à l’avenir dans la population. Les résultats, issus du modèle individu-centré, confirment que la stratégie prioritaire devrait être d’éliminer le plus d’individus séropositifs, et que cibler les femelles et/ou la zone cœur permet d’améliorer l’efficacité des mesures. Bien qu’il n’y ait pas de solution évidente pour la gestion de la brucellose dans notre cas d’étude, les stratégies de gestion ciblées sont très prometteuses et permettent de raffiner les mesures sanitaires classiquement utilisées. Il est donc primordial de bien comprendre l’hétérogénéité de transmission dans les populations sauvages infectées, et de rechercher des stratégies ciblées qui peuvent permettre d’améliorer la gestion en termes d’efficacité et d’acceptabilité
The management of infectious diseases in wildlife reservoirs is particularly challenging and faces several limitations. The development of appropriate management strategies requires a detailed understanding of the factors affecting the transmission and persistence of the infectious agent in the population. Among these factors, heterogeneity of transmission is a common characteristic in natural host-pathogen systems. Indeed, wild animals express a broad range of behaviours, are organised in a variety of social and spatial structures, occupy many areas with very different characteristics and belong to a large diversity of species. Such heterogeneities, from between-individuals to between-species, may result in different contributions to the overall number of new cases of infections. Thus, understanding transmission heterogeneity could provide valuable insights on how to effectively manage these systems, by targeting the individuals or areas that are responsible for most transmissions. The aim of this thesis was to provide insights on the monitoring and management of infectious diseases in heterogeneous wild populations, using Brucella melitensis infection in a French population of wild Alpine ibex (Capra ibex) as a case study. The biology of brucellosis and the ecology of Alpine ibex makes this case study a good candidate for transmission heterogeneity at several levels. Using bacterial examinations, we first established that only 58% of seropositive individuals were at risk to excrete Brucella, and that this risk decreased with increasing age. Then, we took advantage of detailed information available on ibex population dynamics, behaviour, and habitat use, and on epidemiological surveys, to build an individual-based model in order to quantify heterogeneity at the individual and spatial levels. The transmission is extremely heterogeneous between individuals, with females generating around 90% of the new cases of brucellosis infection, and between spatial units, three of the five socio-spatial units (the core area) accounting for more than 80% of brucellosis transmission. Using statistical models to estimate the temporal dynamics of the seroprevalence and of the force of infection in the population, we found evidence that the massive captures with test-and-remove operations that were conducted in 2015 managed to reduce brucellosis transmission in the population. Based on these results, we evaluated several predictive disease management strategies in the individual-based model. Our results confirmed that the primary strategy should be to remove as many seropositive individuals as possible, and that strategies targeting females and/or the core area are more effective than untargeted management. Although there is no silver bullet for the management of brucellosis in the population of study, targeted strategies offer a wide range of promising refinements to classical sanitary measures. We therefore encourage to look for heterogeneity in other infection-wildlife systems and to evaluate potential targeted strategies for improving management schemes in terms of efficiency and acceptability

For the past 45 years, QKI has been studied for its role in the processes of development and central nervous system myelination using the qkv mouse. The presence of a single KH domain and the recent identification of a high-affinity binding site in mRNAs, suggests that it can bind to and regulate mRNAs through processes such as stability, splicing and transport. As a member of the STAR RNA binding family of proteins the QKI isoforms may also be involved in cell signaling pathways.
QKI's involvement in all of these processes, lead us to examine both the protein partners and the mRNA targets of the QKI complex in order to identify potentially new pathways regulated by QKI. In doing so, we identified a novel direct protein-protein interaction with PABP and for the first time described the relocalization of QKI to cytoplasmic granules following oxidative stress. In addition, in vivo mRNA interaction studies were performed and allowed the identification of approximately 100 new mRNA targets in human glioblastoma cells. One of the targets identified was VEGF mRNA.
Another QKI target mRNA is MBP, a major protein component of the myelin sheath and the candidate auto-antigen in multiple sclerosis (MS). In vivo MBP is symmetrically dimethylated on a single arginine residue. To further establish the role of the methylation of MBP in myelination, a methyl-specific antibody and an adenovirus expressing a recombinant protein arginine methyltransferase 5 (PRMT5) was generated. We show that methylated MBP is found in areas of mature myelin and that overexpression of the PRTM5 blocked the differentiation of oligodendrocytes.
Taken together these datas implicate QKI for the first time in the process of human cancer angiogenesis and could explain the vascularization defects observed in some of the qkI mutant mice. In addition, arginine methylation of MBP may prove to have an important role in the process of myelination and in the pathogenesis of demyelination and the autoimmune reaction in diseases such as MS.

Nesta tese apresenta-se a síntese e caraterização de novos derivados de isoquinolinonas, dibenzodiazepinas (DBDA), e acridinas obtidos por acoplamentos usando catalisadores comerciais com metais de transição, com o objetivo principal de funcionarem como inibidores dos recetores de acetilcolinesterase e de MAO-B. Duas novas famílias de isoquinolinonas foram conseguidas a partir de aril-ácido borónicoaldeído obtidas por borilação e desproteção de 2-haloarilamidas acetais, através de arilação usando catalisadores de Rh, Pd e Cu e a partir de ariliminas por reação de borilação e arilação/ciclização one-pot catalisado por Pd, obtendo-se álcoois ou aminas exo-cíclicas quirais. Com o objetivo de síntese de alto rendimento, as arilamidas acetais foram ligadas à resina de Wang por reação de substituição nucleofíla, seguido de borilação, desproteção, e ciclização na fase heterogénea. A resina foi clivada por reação de hidrogenólise obtendo os correspondendes alcanois exo-ciclicos quirais. As reações na fase sólida são até agora desconhecidas. As DBDA foram sintetizadas por acoplamento de Buchwald-Hartwig a partir de derivados de 2-bromoaldiminas, obtidas por condensação dos derivados de 2-bromoaldeídos com Ts, Ms e Ns aminas, e de derivados de 2-bromoanilinas. As acridinas foram obtidas por acoplamento C-N e arilação/ciclização one-pot por reação de derivados de 2-bromoaldiminas e ácido 2-aminofenilboronico pinacol éster; Abstract: Intramolecular Catalytic Arylation of Imines and Analogues - Potential Drugs for Neurodegenerative Diseases This thesis presents the synthesis and characterization of new isoquinolines derivatives, dibenzodiazepine (DBDA) and acridines obtained via transition metal coupling procedures using commercial catalysts. The main objective was the synthesis of cholinesterase and MAO-B inhibitors. Two new families of isoquinolines were obtained from aldehyde-aryl-boronic acids that were accessed from 2-haloarylamide acetals, via catalytic borylation and deprotection. They could also be obtained from arylimines by one-pot borylation / cyclization catalyzed by Pd. This afforded chiral cycloalkanols and exo-cyclic amines. For the purpose of High-Through-Put (HTP) synthesis, the acetals arylamides were immobilized to a Wang resin, by nucleofílica substitution, and then subjects to borylation, deprotection and cyclization on the solid-phase. These types of reactions on the solid-phase are hitherto unknown. The DBDAs were synthesized by Buchwald-Hartwig coupling using 2-bromoaldimines - obtained by condensation of 2-bromoaldehyde derivatives with tosyl, mesyl and nosyl amines -, and from 2-bromoaniline derivatives. The acridines were obtained via one-pot C-N coupling starting with 2-bromoaldimine derivatives and 2-aminophenylboronic acid pinacol ester.

O Vírus da Imunodeficiência Humana (HIV) é o agente etiológico da Síndrome da Imunodeficiência Adquirida (AIDS). A AIDS é uma doença de distribuição mundial, e estima-se que existam atualmente pelo menos 36,9 milhões de pessoas infectadas com o vírus. Durante o seu ciclo replicativo, o HIV promove diversas alterações na fisiologia da célula hospedeira a fim de promover sua sobrevivência e potencializar a replicação. A rápida progressão da infecção pelo HIV-1 em humanos e em modelos animais está intimamente ligada à função da proteína acessória Nef. Dentre as diversas ações de Nef está a regulação negativa de proteínas importantes na resposta imunológica, como o receptor CD4. Sabe-se que esta ação resulta da indução da degradação de CD4 em lisossomos, mas os mecanismos moleculares envolvidos ainda são totalmente elucidados. Nef forma um complexo tripartite com a cauda citosólica de CD4 e a proteína adaptadora 2 (AP-2), em vesículas revestidas por clatrina nascentes, induzindo a internalização e degradação lisossomal de CD4. Pesquisas anteriores demonstraram que o direcionamento de CD4 aos lisossomos por Nef envolve a entrada do receptor na via dos corpos multivesiculares (MVBs), por um mecanismo atípico, pois, embora não necessite da ubiquitinação de carga, depende da ação de proteínas que compõem os ESCRTs (Endosomal Sorting Complexes Required for Transport) e da ação de Alix, uma proteína acessória da maquinaria ESCRT. Já foi reportado que Nef interage com subunidades dos complexos AP-1, AP-2, AP-3 e Nef não parece interagir com subunidades de AP-4 e AP-5. Entretanto, o papel da interação de Nef com AP-1 e AP-3 na regulação negativa de CD4 ainda não está totalmente elucidado. Ademais, AP-1, AP-2 e AP-3 são potencialmente heterogêneos devido à existência de isoformas múltiplas das subunidades codificadas por diferentes genes. Todavia, existem poucos estudos para demonstrar se as diferentes combinações de isoformas dos APs são formadas e se possuem propriedades funcionais distintas. O presente trabalho procurou identificar e caracterizar fatores celulares envolvidos na regulação do tráfego intracelular de proteínas no processo de regulação negativa de CD4 induzido por Nef. Mais especificamente, este estudo buscou caracterizar a participação do complexo AP-1 na modulação negativa de CD4 por Nef de HIV-1, através do estudo funcional das duas isoformas de ?-adaptina, subunidades de AP-1. Utilizando a técnica de Pull-down demonstramos que Nef é capaz de interagir com ?2. Além disso, nossos dados de Imunoblot indicaram que a proteína ?2-adaptina, e não ?1-adaptina, é necessária no processo de degradação lisossomal de CD4 por Nef e que esta participação é conservada para degradação de CD4 por Nef de diferentes cepas virais. Ademais, por citometria de fluxo, o silenciamento de ?2, e não de ?1, compromete a diminuição dos níveis de CD4 por Nef da membrana plasmática. A análise por imunofluorêsncia indireta também revelou que a diminuição dos níveis de ?2 impede a redistribuição de CD4 por Nef para regiões perinucleares, acarretando no acúmulo de CD4, retirados por Nef da membrana plasmática, em endossomos primários. A depleção de ?1A, outra subunidade de AP-1, acarretou na diminuição dos níveis celulares de ?2 e ?1, bem como, no comprometimento da eficiente degradação de CD4 por Nef. Além disso, foi possível observar que, ao perturbar a maquinaria ESCRT via super-expressão de HRS (uma subunidade do complexo ESCRT-0), ocorreu um acumulo de ?2 em endossomos dilatados contendo HRS-GFP, nos quais também detectou-se CD4 que foi internalizado por Nef. Em conjunto, os resultados indicam que ?2-adaptina é uma importante molécula para o direcionamento de CD4 por Nef para a via ESCRT/MVB, mostrando ser uma proteína relevante no sistema endo-lisossomal. Ademais, os resultados indicaram que as isoformas ?-adaptinas não só possuem funções distintas, mas também parecem compor complexos AP-1 com diferentes funções celulares, já que apenas a variante AP-1 contendo ?2, mas não ?1, participa da regulação negativa de CD4 por Nef. Estes estudos contribuem para o melhor entendimento dos mecanismos moleculares envolvidos na atividade de Nef, que poderão também ajudar na melhor compreensão da patogênese do HIV e da síndrome relacionada. Em adição, este trabalho contribui para o entendimento de processos fundamentais da regulação do tráfego de proteínas transmembrana no sistema endo-lisossomal.
The Human Immunodeficiency Virus (HIV) is the etiologic agent of Acquired Immunodeficiency Syndrome (AIDS). AIDS is a disease which has a global distribution, and it is estimated that there are currently at least 36.9 million people infected with the virus. During the replication cycle, HIV promotes several changes in the physiology of the host cell to promote their survival and enhance replication. The fast progression of HIV-1 in humans and animal models is closely linked to the function of an accessory protein Nef. Among several actions of Nef, one is the most important is the down-regulation of proteins from the immune response, such as the CD4 receptor. It is known that this action causes CD4 degradation in lysosome, but the molecular mechanisms are still incompletely understood. Nef forms a tripartite complex with the cytosolic tail of the CD4 and adapter protein 2 (AP-2) in clathrin-coated vesicles, inducing CD4 internalization and lysosome degradation. Previous research has demonstrated that CD4 target to lysosomes by Nef involves targeting of this receptor to multivesicular bodies (MVBs) pathway by an atypical mechanism because, although not need charging ubiquitination, depends on the proteins from ESCRTs (Endosomal Sorting Complexes Required for Transport) machinery and the action of Alix, an accessory protein ESCRT machinery. It has been reported that Nef interacts with subunits of AP- 1, AP-2, AP-3 complexes and Nef does not appear to interact with AP-4 and AP-5 subunits. However, the role of Nef interaction with AP-1 or AP-3 in CD4 down-regulation is poorly understood. Furthermore, AP-1, AP-2 and AP-3 are potentially heterogeneous due to the existence of multiple subunits isoforms encoded by different genes. However, there are few studies to demonstrate if the different combinations of APs isoforms are form and if they have distinct functional properties. This study aim to identify and characterize cellular factors involved on CD4 down-modulation induced by Nef from HIV-1. More specifically, this study aimed to characterize the involvement of AP-1 complex in the down-regulation of CD4 by Nef HIV-1 through the functional study of the two isoforms of ?-adaptins, AP-1 subunits. By pull-down technique, we showed that Nef is able to interact with ?2. In addition, our data from immunoblots indicated that ?2- adaptin, not ?1-adaptin, is required in Nef-mediated targeting of CD4 to lysosomes and the ?2 participation in this process is conserved by Nef from different viral strains. Furthermore, by flow cytometry assay, ?2 depletion, but not ?1 depletion, compromises the reduction of surface CD4 levels induced by Nef. Immunofluorescence microscopy analysis also revealed that ?2 depletion impairs the redistribution of CD4 by Nef to juxtanuclear region, resulting in CD4 accumulation in primary endosomes. Knockdown of ?1A, another subunit of AP-1, resulted in decreased cellular levels of ?1 and ?2 and, compromising the efficient CD4 degradation by Nef. Moreover, upon artificially stabilizing ESCRT-I in early endosomes, via overexpression of HRS, internalized CD4 accumulates in enlarged HRS-GFP positive endosomes, where co-localize with ?2. Together, the results indicate that ?2-adaptin is a molecule that is essential for CD4 targeting by Nef to ESCRT/MVB pathway, being an important protein in the endo-lysosomal system. Furthermore, the results indicate that ?-adaptins isoforms not only have different functions, but also seem to compose AP-1 complex with distinct cell functions, and only the AP-1 variant comprising ?2, but not ?1, acts in the CD4 down-regulation induced by Nef. These studies contribute to a better understanding on the molecular mechanisms involved in Nef activities, which may also help to improve the understanding of the HIV pathogenesis and the related syndrome. In addition, this work contributes with the understanding of primordial process regulation on intracellular trafficking of transmembrane proteins.

碩士
國立中興大學
資訊科學與工程學系所
101
A new computer-aided diagnosis method is proposed to diagnose gastroesophageal reflux disease (GERD) from endoscopic images of the esophageal-gastric junction. To avoid the inferences of endoscope devices and automatic camera white balance adjustment, multiple color invariant models are used to represent endoscopic images. Then, visual vocabularies are built from each color model to describe the mucosa of the esophageal-gastric junction for support vector machine training. To simultaneously consider the prediction results of each color model, a hierarchical support vector machine scheme is proposed. During validation, visual vocabularies extracted from the test endoscopic image are used as the input of the hierarchical support vector machine to diagnose GERD. As shown in the experiments, our method can automatically diagnose GERD without any manual selection of region of interest and achieve better accuracy compared to methods using only one color invariant model.

碩士
國立清華大學
資訊系統與應用研究所
101
During the last few years, the knowledge of drug, disease phenotype and protein has been rapidly accumulated and more and more scientists have been draw attention to inferring drug-disease associations by computational method. Development of an integrated approach for systematic discovering drug-disease associations by those informational data is an important issue. We combine three weighted networks of drug, genomic and disease phenotype data from available experimental data and knowledge then infer drug-disease associations by a hetero-network propagation approach. In the experiments, we adopt prostate cancer and colorectal cancer as our test data. We select the manually curated associations from comparative toxicogenomics database as our benchmark. The ranked results show that our proposed method obtains high specificity and sensitivity and clearly outperforms previous methods. We clearly demonstrate the feasibility and benefits of using network-based analyses of chemical, genomic and phenotype data to reveal drug-disease associations. The potential associations which were inferred by our method drew the biologists’ attention and provide new perspectives for toxicogenomics and drug reposition evaluation.

碩士
國立臺灣大學
流行病學研究所
97
Gene-expression has been a popular research topic in recent years. Student t-test is commonly adopted to screen disease-related genes. However, when the researches are focused on heterogeneous disease, the means of gene-expression levels between case group and control group may be similar, and thus, the difference would be difficult to be detected by conventional Student t-test. This study proposed half Student t-test to examine heterogeneous disease. Test statistics of half Student t-test only considers sample standard deviation of control group, without considering the sample standard deviation of case group. This study applied Monte Carlo simulation and real gene-expression data of colon cancer to compare the power performance of half Student t-test and conventional Student t-test. Under the simulated scenario, this study found that half Student t-test could have 35% higher statistical power than conventional Student t-test. In addition, after false discovery rate (cut-off point set at 0.05) control of colon cancer gene-expression data, half Student t-test could detect 279 more significant genes than conventional Student t-test. Half Student t-test is easy to execute with good statistical power, and is worth to be recommended as a method of detecting heterogeneous disease gene-expression difference.

博士
國立中央大學
化學工程與材料工程研究所
95
The protein VP2, matured from the polyprotein, which was encoded by the genome of infectious bursal disease virus (IBDV), is the primary host-protective immunogen of IBDV. According to VP2-452H subviral particle (SVP) analysis, the determination crystal structure and enzyme-linked immunosorbent assay (ELSIA), showed thar His-tag was not exposed on the surface of VP2-452H SVPs. Thus illustrate that the His-tag apparently did not attribute to the effective purification of this protein by IMAC. An affinity must have existed between the protein VP2 and the immobilized metal ions to achieve SVP binding with Ni-NTA resin. Accordingly, the IBDV generated from DF-1 cell culture and non-tagged VP2-441 SVP generated from a baculovirus-insect cell expression system were purified by IMAC. The purification of IBDV viron through IMAC obtained a 60.5% recovery, and the IMAC-purified IBDV has a similar morphology to the wild-type IBDV with a diameter of 65 nm through electron microscope observation. For SVP formed by VP2-441 purified by IMAC a recovery 92% and a purity of also 92% of mature VP2 were obtained. SVP formed by VP2-441 exhibited a diameter approximately 25 nm. These results obtained from the above experiments can demonstrate 1) the protein VP2 does have interaction with immobilized nickel ion; and 2) the protein VP2 can assist both IBDV viron and SVPs to have the affinity with Ni-NTA resin. The recombinant protein VP2-441, i.e., a structural protein VP2 of infectious bursal disease (IBD) virus, can self-assemble into T=1 subviral particles (SVPs) in baculovirus expression system. These SVPs are not to have multiple his-tags on the surface which result in an efficient purification by immobilized metal-ion affinity chromatography (IMAC). This study aimed at getting more insight into the interaction between VP2-441 SVPs and immobilized metal (Ni2+) ions at molecular level. First of all, large quantity of highly purified VP2-441 SVPs obtained by a one-step purification process allowed the performance of equilibrium adsorption measurements and subsequent determinations of binding constants by fitting two isotherm models, i.e., Temkin and Langmuir-Freundlich. Two general conclusion are obtained, first, the maximum bound VP2-441 SVPs per volume resin is limited because the pore size of IMA gel (ca. 24 nm in diameter) is similar to that of SVPs (20 – 25 nm) and the diffusion of the latter into the pores is hindered. The other is that SVPs have an extremely high affinity to the immobilized Ni+2 ions because the dissociate constants obtained from different models are in the scale of 10-9 M, which suggested the interaction mimicking that between an antigen and its antibody. The high binding strength is derived from a multiple-site binding between VP2-441 SVPs and Ni2+ ions as demonstrated by a concave-up Scatchard plot. Finally, we found that the adsorption of SVPs can be well described by Temkin model. A detail understanding of SVP-immobilized metal ion interactions can provide useful strategies for conducting preparative-scale separations of SVPs or even a real virus using IMAC.

abstract: In this dissertation the potential impact of some social, cultural and economic factors on Ebola Virus Disease (EVD) dynamics and control are studied. In Chapter two, the inability to detect and isolate a large fraction of EVD-infected individuals before symptoms onset is addressed. A mathematical model, calibrated with data from the 2014 West African outbreak, is used to show the dynamics of EVD control under various quarantine and isolation effectiveness regimes. It is shown that in order to make a difference it must reach a high proportion of the infected population. The effect of EVD-dead bodies has been incorporated in the quarantine effectiveness. In Chapter four, the potential impact of differential risk is assessed. A two-patch model without explicitly incorporate quarantine is used to assess the impact of mobility on communities at risk of EVD. It is shown that the overall EVD burden may lessen when mobility in this artificial high-low risk society is allowed. The cost that individuals in the low-risk patch must pay, as measured by secondary cases is highlighted. In Chapter five a model explicitly incorporating patch-specific quarantine levels is used to show that quarantine a large enough proportion of the population under effective isolation leads to a measurable reduction of secondary cases in the presence of mobility. It is shown that sharing limited resources can improve the effectiveness of EVD effective control in the two-patch high-low risk system. Identifying the conditions under which the low-risk community would be willing to accept the increases in EVD risk, needed to reduce the total number of secondary cases in a community composed of two patches with highly differentiated risks has not been addressed. In summary, this dissertation looks at EVD dynamics within an idealized highly polarized world where resources are primarily in the hands of a low-risk community – a community of lower density, higher levels of education and reasonable health services – that shares a “border” with a high-risk community that lacks minimal resources to survive an EVD outbreak.
Dissertation/Thesis
Doctoral Dissertation Applied Mathematics 2018

The highly optimized hierarchical structure of trabecular bone is a major contributor to its remarkable mechanical properties. At the micro-scale level, individual plate-like and rod-like trabeculae are interconnected, forming a complex trabecular architecture. It is widely believed that bone strength, an important mechanical characteristic that describes the capability of bone to resist fracture, is largely determined by the tissue-level material properties of these microscopic trabecular elements. However, due to the complicated microstructure and irregular morphology of trabecular bone, a link between the tissue-level and the apparent level mechanics in trabecular bone has never been established. Thus, the goal of this thesis is to examine the tissue-level material properties of trabecular bone and their contribution to apparent-level bone mechanics, and ultimately to improve our fundamental understanding and assessment of bone strength in diseased and healthy patients. At the micro-scale level, plate-like and rod-like trabeculae are distinctly aligned along different orientations on the anatomical axis of the skeleton. Also, the highly organized underlying ultrastructure of bone tissue suggests trabecular bone might possess an anisotropic tissue modulus, i.e. different modulus in the axial and lateral cross-section of a trabecula. In this thesis, we studied this tissue-level anisotropy by examining mechanical properties of individual trabecular plates and rods aligned longitudinally, obliquely, and transversely on the anatomical axis using micro-indentation. We discovered that, despite the different orientations of trabeculae, tissue moduli are higher in the axial direction than in the lateral direction for both plates and rods. We also discovered that plates have a higher tissue modulus than rods, suggesting different degrees of mineralization. Furthermore, the tissue mineral density correlated strongly but distinctly with tissue modulus in the axial and lateral directions, providing descriptions on how spatially heterogeneous mineralization at the tissue level affects the tissue modulus. After characterization of the anisotropic and heterogeneous modulus of trabecular bone at the tissue level, we then sought to investigate its contribution to apparent-level mechanical properties, including apparent Young’s modulus and yield strength. Non-linear FE voxel models incorporating experimentally determined anisotropy and heterogeneity were created from micro-computed tomography (µCT) images of healthy trabecular bone samples. Apparent Young's modulus and yield strength predicted by the models were compared to and correlated with gold standard mechanical testing measurements, as well as to the same FE models without incorporation of anisotropy and/or heterogeneity. We discovered that the anisotropic model prediction was highly correlated and indistinguishable from mechanical testing measurements. However, the prediction power of the model was not enhanced by incorporating anisotropy and heterogeneity (compared to a homogeneous and isotropic model), suggesting that variances in tissue-level material properties contribute minimally to the apparent level bone behaviors in healthy bone. However, the possibility remained that a more substantial contribution could arise in diseased bone, particularly diseases in which tissue-level properties are compromised. Therefore, we studied trabecular bone in two diseased conditions – subchondral bone in human knees affected by osteoarthritis and pelvic bone affected by adolescent idiopathic sclerosis – to see how disease can alter the tissue-level and, consequently, apparent-level bone mechanics. In OA bone, we found a significant decrease in tissue modulus in the subchondral bone under severely damaged cartilage compared to control, which provides an explanation for a minimal increase in apparent stiffness with an almost doubled bone volume fraction. In AIS bone, no differences were found in tissue-level or apparent level Young’s modulus compared to control. However, the mineral density was found to play a distinct role in the modulus of growing bone tissue compared to mature bone.

abstract: This dissertation explores the impact of environmental dependent risk on disease dynamics within a Lagrangian modeling perspective; where the identity (defined by place of residency) of individuals is preserved throughout the epidemic process. In Chapter Three, the impact of individuals who refuse to be vaccinated is explored. MMR vaccination and birth rate data from the State of California are used to determine the impact of the anti-vaccine movement on the dynamics of growth of the anti-vaccine sub-population. Dissertation results suggest that under realistic California social dynamics scenarios, it is not possible to revert the influence of anti-vaccine contagion. In Chapter Four, the dynamics of Zika virus are explored in two highly distinct idealized environments defined by a parameter that models highly distinctive levels of risk, the result of vector and host density and vector control measures. The underlying assumption is that these two communities are intimately connected due to economics with the impact of various patterns of mobility being incorporated via the use of residency times. In short, a highly heterogeneous community is defined by its risk of acquiring a Zika infection within one of two "spaces," one lacking access to health services or effective vector control policies (lack of resources or ignored due to high levels of crime, or poverty, or both). Low risk regions are defined as those with access to solid health facilities and where vector control measures are implemented routinely. It was found that the better connected these communities are, the existence of communities where mobility between risk regions is not hampered, lower the overall, two patch Zika prevalence. Chapter Five focuses on the dynamics of tuberculosis (TB), a communicable disease, also on an idealized high-low risk set up. The impact of mobility within these two highly distinct TB-risk environments on the dynamics and control of this disease is systematically explored. It is found that collaboration and mobility, under some circumstances, can reduce the overall TB burden.
Dissertation/Thesis
Doctoral Dissertation Applied Mathematics for the Life and Social Sciences 2018