Dissertations / Theses on the topic 'Center for Perinatal Biology'

The studies presented in this dissertation establish the dynamics of Eleven nineteen Lysine-rich leukemia (ELL) family of elongation factors during B cell differentiation and provide a description of ELL3 function in B cells. The transition from a mature naïve B cells into an activated B cell is dependent on a large increase in transcriptional output, which is followed by focused expression on secreted immunoglobulin upon terminal differentiation into plasma cell. While ELL family members have previously been implicated in alternative splicing at the immunoglobulin heavy chain locus in plasma cells, their presence and function prior to differentiation is currently not known. However, the use of elongation factors has been implied by the finding of mostly paused RNA polymerase II in the genome of naïve B cells. In the first study, the expression of transcriptional elongation factor ELL3 is shown to be restricted to activated B cells and B cell lymphomas. All three family members were characterized in B cell lymphoma cell lines, genome wide expression, microarray analysis and primary B cell stimulus. The expression of ELL3 was induced upon activation of B cells concurrently with family member ELL. In addition, the abundant expression of ELL3 was restricted to GC derived B cell lymphoma cell lines. While the expression of ELL is maintained, the expression of ELL3 is diminished and ELL2 is up-regulated in terminally differentiated plasma cells. The expression of master regulator of terminal plasma cell differentiation PRDM1 was inverse correlated with that of ELL3. To further establish PRDM1s role in regulating the ELL family member dynamics, global binding was assessed in plasma cell lines. Chromatin immunoprecipitation followed by quantitative PCR was utilized to identify direct association of PRDM1 at exclusively the ELL3 loci. Ectopic expression of PRDM1 in B cells down regulated the expression of ELL3. Furthermore, two consensus PRDM1 binding sites were defined at the ELL3 loci, which mediate significant repression of the promoter activity. Collectively, these experiments indicate that PRDM1 mediates the switch from ELL3 in B cells to ELL2 in plasma cells. The data presented in the final chapter aimed at defining a function for ELL3 in the cells that express it most abundantly, which are B cell lymphoma cell lines. Transient depletion of ELL3 in a Burkitt’s lymphoma cell line resulted in a diminished proliferation rate due to a severe disruption of DNA replication and its regulators minichromosome maintenance proteins. Additionally, compromised cell division and mitotic regulators were observed along with increased DNA damage and cell death. The data presented here demonstrate a key role for ELL3 in the proliferation and survival of B cell lymphomas and positions ELL3 as an attractive therapeutic target against B cell lymphoma’s with a germinal center origin.

Asthma is a debilitating disease of the lungs affecting 235 million people worldwide. Fungus-associated asthma leads to a particularly severe type of disease, and exposure to environmental fungi and their products is unavoidable due to the ubiquitous nature of fungal species. Besides being allergenic, fungi are opportunistic pathogens, and anti-fungal and/or allergic pathways may be modified through repeated inhalation of immunomodulatory agents, affecting the outcome of fungus-induced asthma.

Our aim in this project was to investigate the extent to which repeated inhalation of immunomodulatory agents influence the lung mucosal responses in a naïve murine host or in one that had been sensitized to fungal proteins (allergic). The immunomodulatory substances chosen hold relevance to human inhalational exposure, and included live or irradiation-killed Aspergillus fumigatus (a fungi) spores, deoxyxnivalenol (a mycotoxin), and fluticasone propionate (an inhalationally administered corticosteroid, commonly prescribed for allergic asthma). In a naïve host, inhalation of live A. fumigatus spores showed pathological features of fungal asthma. However, in an allergen-sensitized lung, both dead and live A. fumigatus spores established fungal airway disease, albeit to different extents. Next, we tested the effect of deoxynivalenol in an allergic host and found that its repeated inhalation did not affect pulmonary disease pathology, but did lead to a dose- and time- dependent increase in mucosal and systemic total IgA. Finally, we tested the effect of fluticasone propionate, and found that it did not influence the development of fungal airway disease, but did induce dynamic changes in lung physiology and antibody titers.

Besides mimicking human inhalational exposures, inhalation ensures direct interaction of the inhaled substances with airway epithelium, which plays an important role in defense against inhaled substances and in asthma pathophysiology. By analyzing various mechanisms involved in murine lung-mucosal response to the inhaled substances, a critical involvement of airway epithelium as an orchestrator of immune responses is highlighted, and this would inform mechanism-based future studies. In conclusion, this project is likely to aid in establishing evidence based standards for fungus-related exposures and in making informed therapeutic decisions for fungus-associated diseases.

Master of Science in Biomedical Sciences
Department of Diagnostic Medicine/Pathobiology
Alison P. Adams
The animal health industry is a growing industry. Owners of pets and other animals want to ensure their animals are healthy. To do this, the animal health industry markets a variety of products from pharmaceutical products, such as antibiotics, to biological products, such as vaccines. These products are developed and marketed after the company provides regulators the necessary information as guided by a set of regulations. Pharmaceutical products follow Title 21 of the Code of Federal Regulations, while biological products follow Title 9 of the Code of Federal Regulations. During the product development process as well as after marketing, regardless of the regulations to follow, each product must go through testing for efficacy, safety, potency, and stability. The regulatory guidelines provide direction to companies on expectations of the testing requirements for each type of product. Different analytical techniques are used to provide the necessary data in support of product development. Discussed in this report, two analytical techniques are well known in the industry, and one is quickly becoming a technique of great value. Mass spectrometry, coupled with liquid chromatography, is an industry standard for testing product potency and purity as well as pharmacokinetics. The enzyme-linked immunosorbent assay (ELISA) is also used to measure potency of products as well as product stability. The newest technique is flow cytometry that characterizes cells within a suspension, most often with the use of cellular biomarkers as targets. By understanding the application of each technique as well as how it relates to regulatory requirements, the industry can provide assurances to regulators that their products are safe and efficacious for the treatment and/or prevention of animal diseases. This report outlines the history, theory, and use of three different analytical techniques currently used for pharmaceutical and biological products in animal health.

Wastewater application to land can be a useful tool for mitigating impacts of nutrient enrichment on aquatic systems. A land application treatment system at VCU’s Rice Rivers Center in Charles City County, VA provided an opportunity to study the impact of wastewater addition on the biogeochemistry of forests representative of the Mid-Atlantic Coastal Plain. Nutrient concentrations in throughfall and leachate were measured at Treatment and Control sites to assess differences in nutrient deposition and retention. Wastewater amended plots from the Walter L. Rice education building received 20-fold (N) and 6-fold (P) higher inputs relative to Control plots and plots located at the Virginia Department of Game and Inland Fisheries building. Despite higher inputs, leaching losses of P from the Rice Treatment plots were comparable to Control plots, indicating that the land-based application system effectively mitigated wastewater loads. Leaching losses of nitrate were two-fold higher from Treatment plots relative to Controls, suggesting a potential N saturation effect and a reduction in N retention capacity of the treated forest. Nitrogen effects on vegetation were indicated by lower root biomass and greater root N content among Treatment plots relative to Control plots. Overall, these results suggest that wastewater–amended plots near the Rice education building receive appreciably higher nutrient inputs and could, therefore, serve as a model system for assessing effects on forest ecosystems.

Neutral two- and three-pion productions in proton-proton collisions at a center of mass energy of 2.465 GeV have been studied using the WASA detector and an internal pellet target at the CELSIUS storage ring in Uppsala. An important part of the detector for the measurments was a central electromagnetic calorimeter composed of 1012 CsI crystals, which measured the photons originating from neutral pion decays. Test measurements and calibration procedures for this detector part were carried out. An important part of the analysis was the identification of the neutral pions from the invariant mass of the decay gammas and the use of Monte Carlo simulations to understand the detector responds. Total cross sections for the pp→ppπ0π0 and pp→ppπ0π0π0 reactions are presented as well as distributions of relevant kinematical variables for the pp→ppπ0π0 reaction. The distributions show significant deviations from phase space predictions. These deviations are typical for resonance production. The excitation of two simultaneous Δ resonances seems to be the main reaction mechanism.

Med den här uppsatsen har jag velat diskutera relationen mellan utomhusdidaktik respektive inomhusdidaktik som utgångspunkt för undervisningen i den svenska skolan. Ett annat syfte är att med utgångspunkt i ovan nämnda didaktiker fördjupa kunskaperna i utbildningsmetodik för lärare i allmänhet och biologilärare i synnerhet.

Studien utgår från två frågeställningar som jag genom intervjuer med två representanter för utomhusdidaktik och två representanter för inomhusdidaktik har försökt besvara.

Resultatet av denna studie visar att utomhusdidaktik och inomhusdidaktik kan vara mycket lika i sina fokus på de didaktiska frågorna. Ett undantag är var-frågan som är av större betydelse för utomhusdidaktiken. Studien visar även att utomhusdidaktik berör många fler ämnen än biologiämnet.

En viktig slutsats är att undervisningen i skolan ofta saknar verklighetsförankring. Utomhusdidaktik och science-center-didaktik, som representerar inomhudidaktik i denna studie kan båda hjälpa skolan att konkretisera den ofta teoretiska undervisningen och bidra till en ökad förståelse hos eleverna.

Diffuse large B cell lymphoma (DLBCL) is the most common non-Hodgkin’s lymphoma. Patients who fail conventional therapy (~50%) have a poor prognosis and few treatment options. It is essential to understand the underlying biological processes, the progression of the disease, and utilize this information to develop new therapeutics. DLBCL patients with high C-MYC expression have a poor prognosis and new therapeutics for these patients are needed. This thesis describes work testing the hypothesis that JQ1, which can indirectly inhibit C-MYC in some tumors, can be used as an effective treatment for DLBCL. Some tumors have an unknown mechanism causing high C-MYC expression, leading me to investigate the underlying mechanisms. YY1 is a transcriptional regulator of c- Myc and has been implicated in DLBCL and as a potential regulator of the germinal center (GC) reaction. DLBCL arises from GC cells or post-GC cells. I tested the hypothesis that YY1 regulates the GC reaction. SMURF2 is an E3-ubiquitin ligase for YY1 and a tumor suppressor for DLBCL. I was interested in examining the mechanism underlying the suppression of DLBCL by SMURF2 leading to the hypothesis that SMURF2 regulates the GC. This thesis shows JQ1 leads to cell death and cellular senescence in human DLBCL cells. I conclude that BRD4 inhibition by JQ1 or derivatives could provide a new therapeutic avenue for DLBCL patients. I also show loss of YY1 perturbs the GC by decreasing the dark zone and increasing apoptosis. Finally I show modulation of SMURF2 does not affect the GC, suggesting SMURF2 utilizes a different mechanism to act as a tumor suppressor and may not modulate YY1 in the context of the GC.

Tooth development is precisely controlled by epithelium-mesenchyme interactions, coordinated signaling pathways and associated transcription factors. Although the processes involved in tooth development are well established, details of the cellular and molecular mechanisms that control tooth development are not fully understood. One of the primary unknown mechanisms is the regulation of dental epithelial stem cells (DESCs), including DESC specification, proliferation and differentiation. In this dissertation, I have addressed this gap in knowledge by studying the role of Pituitary homeobox 2 (Pitx2) and Iroquois 1 (Irx1) in teeth at the cellular and molecular level in mice. PITX2 contains mutations of which are associated for Axenfeld-Rieger syndrome (ARS) in humans and is also required for early tooth development. All the background knowledge is included in Chapter I. In Chapter II, I describe the conditional ablation of Pitx2 in the dental epithelium using a Krt14Cre driver line (Pitx2cKO mice). Knocking out Pitx2 in teeth led to delayed epithelial invagination at bud stage and disruption of tooth morphogenesis at cap stage. At the cellular level, Pitx2 mediates DESC differentiation, daughter cell proliferation in bud stage tooth and regulates enamel knot formation in cap stage tooth. At the molecular level, Pitx2 acts as an upstream regulator of the sonic hedgehog (Shh) signaling pathway by regulating the expression of Shh in the dental epithelial signaling center during early tooth development. In addition, I demonstrated that Pitx2 directly controls the transcription of Irx1. In Chapter III, I determined the cellular and molecular mechanisms of Irx1 in mice. Irx1 general knockout mice were generated by replacing the entire Irx1 gene body with a LacZ reporter gene. Irx1 null mice are neonatal lethal and this lethality is due to pulmonary immaturity with defective surfactant protein secretion. In teeth, Irx1 is expressed in the outer enamel epithelium (OEE) and stratum reticulum (SR) and mediates DESC to OEE and SR differentiation through regulation of Forkhead box protein J1 (Foxj1) and Sex determining region Y-box9 (Sox9). In summary, I identified a Pitx2-Irx1 regulatory network that controls DESC differentiation in teeth, which provided the field with a better understanding of tooth development and tooth regeneration.

Neutral two- and three-pion productions in proton-proton collisions at a center of mass energy of 2.465 GeV have been studied using the WASA detector and an internal pellet target at the CELSIUS storage ring in Uppsala. An important part of the detector for the measurments was a central electromagnetic calorimeter composed of 1012 CsI crystals, which measured the photons originating from neutral pion decays. Test measurements and calibration procedures for this detector part were carried out. An important part of the analysis was the identification of the neutral pions from the invariant mass of the decay gammas and the use of Monte Carlo simulations to understand the detector responds.

Total cross sections for the pp→ppπ⁰π⁰ and pp→ppπ⁰π⁰π⁰ reactions are presented as well as distributions of relevant kinematical variables for the pp→ppπ⁰π⁰ reaction.

The distributions show significant deviations from phase space predictions. These deviations are typical for resonance production. The excitation of two simultaneous Δ resonances seems to be the main reaction mechanism.

Amphibians and aquatic invertebrates have complex life histories that link aquatic and terrestrial food webs. It has been suggested that amphibian reproduction is an important source of carbon to some aquatic systems. This process of energy flow may be shaped by shifts in habitat selection in response to predators. We hypothesized that predators decrease colonization and oviposition of prey, reducing active inputs. Thus predation risk is expected to shift the relative amounts of active and passive subsidies. We manipulated the presence of fish predators in aquatic mesocosms. Results suggest hylid treefrog eggs and hydrophilid beetles were less abundant in predator treatments. This difference in oviposition and colonization translated into small reductions in calories and ash free dry mass of active inputs. However, passive allochthonous inputs were more than double active amounts and variable, therefore relative amounts of active and passive inputs did not differ across the levels of predation risk.

Populations of largemouth bass, Micropterus salmoides, and bluegill, Lepomis macrochirus, were evaluated from five pit-lakes in Muhlenberg County, Kentucky, to determine if accurate proportional stock density (PSD) data can be obtained from a mandatory creel survey. It was hypothesized that the proportion of stock-to-quality (300-400mm) and quality (+400mm) largemouth bass from four years (2007-2010) of creel survey data would be statistically similar to those generated through on-site sampling in 2011. Fish were collected via a combination of gill netting, seining, hook-and-line fishing, and boat-mounted electro-fishing. In two of the pit-lakes, the sampling-generated length frequency data was not significantly different from the creel survey data (Pump Gadj[1]=0.03, P=0.8629, Goose Gadj[1]=0.76, P=0.3850). There were significant differences between creel and sampling data for the other pit-lakes (Big Reno Gadj[1]=5.74 P=0.0166, Airstrip Gadj[1]=14.3 P=0.0002, Lime Gadj[1]=9.81 P=0.0017). At least one of the lakes likely demonstrated significances because of low sample size (Airstrip and/or Lime). Changes in population structure due to modified harvest regulations may be responsible for the significant differences (Big Reno and Lime). Population structures were verified with relative weight, length-at-age, and an assessment of five years of largemouth bass and bluegill PSD data. It appears that creel survey data does accurately reflect that of simple sampling techniques and can help guide management decisions.

Rubella virus (RUBV) is a positive-strand RNA virus and the causative agent of rubella and congenital rubella syndrome in humans. To replicate its RNA, RUBV forms membrane-associated spherules, called replication complexes (RCs), the induction of which requires the two virus nonstructural proteins (NSPs), P150 and P90. Interestingly, late in infection the NSPs form a unique cytoplasmic fiber network, similar in appearance to microtubules, the function of which is unknown. Little is known about the roles of the RUBV NSPs in forming these structures and, to this end, we scrutinized the behavior and biochemical properties of the NSPs, both after expression from plasmids and during RUBV infection, using mutagenic, biochemical and pharmacological approaches. The following findings were made: First, the precursor from which P150 and P90 are produced via an embedded protease at the C-terminus of P150, called P200, was required for initial targeting to cytoplasmic foci. P150 was the determinant of fiber formation and while P90 had no specific targeting sequences on its own, P90 sequences within P200 were required for correct targeting of P200. An alpha-helix at the N-terminus of P150 was also important for correct targeting of P200, putatively by mediating the interaction between P150 and P90 within the precursor. Second, the membrane binding domain within the NSPs was within the N-terminal ~450 amino acids of P150. P150 is in an exceptionally tight association with membranes. Third, both the N- and C-terminal regions of P150, and specifically long alpha-helices within these regions, are necessary for fiber formation. Fiber formation relied on an intact microtubule network, but neither microtubule repositioning nor dynamic movement along microtubules was required. Additionally, it was shown that microtubules were not necessary in RUBV replication. Finally, P150 fibers were not required for RUBV replication; however, it was shown that the fibers are likely important in formation of cytoplasmic extensions through which a novel system of cell-to-cell transport of viral RNA in the absence of virus particles appears to occur.

Aufgrund der rasanten technischen Entwicklung im Bereich der pränatalen Diagnostik in den letzten Jahren werden mehr angeborene Fehlbildungen zu einem früheren Zeitpunkt der Schwangerschaft und detaillierter erkannt. Das führt dazu, dass die zur Autopsie gelangenden Feten zunehmend kleiner sind und komplexere Befunde aufweisen. Gleichzeitig werden die genetischen Ursachen von mehr Erkrankungen, Fehlbildungen und Syndromen aufgedeckt, so dass die DNA-Diagnostik, Chromosomenanlyse u.a. molekulargenetische und biochemische Erkenntnisse die klinisch-genetische Diagnostik zunehmend ergänzen. Durch Festlegung einer gezielten Autopsiestrategie und Nutzung moderner Kommunikationsmittel kann der Pathologe die in den letzten Jahren gestiegenen klinischen Anforderungen erfüllen. Ein Problem ist es jedoch, dass nicht genügend Pathologen auf dem Gebiet der Fetalpathologie spezialisiert sind. Das trifft nicht nur für Deutschland zu, sondern das ist mehr oder weniger eine weltweite Situation. Ein möglicher Lösungsansatz dieses Problems wäre die Nutzung von Softwareprogrammen, die dazu dienen, unerfahrene Pathologen moderne Anleitungen während der Autopsie von Feten zu geben. Das kann mit einer telepathologischen Betreung durch einen Experten auf diesem Gebiet kombiniert werden. Darüberhinaus erlaubt die derzeitige digitalisierte Befunddokumentation den Vergleich aktueller mit gespeicherten Fällen sowie Recherchen und stellt eine gute Basis zur Erstellung von Lehrmaterial dar.
Due to the enormous technical developments in recent years in the area of prenatal ultrasound more malformations are detected in greater detail and at an earlier point in time during pregnancy than in the past. As a result, fetuses of increasingly smaller size and with complex diagnoses are presented for autopsy. At the same time, the genetic causes of ever more diseases, malformations and syndromes are being discovered, so that DNA diagnostics, chromosome analysis and, among others, molecular-genetic and biochemical investigations have come increasingly to complement clinical-genetic diagnostics. Through establishment of a clinically oriented autopsy strategy and the use of modern communication media, pathologists can satisfy the increased clinical demands of recent years. The remaining problem is that we do not have enough pathologists experienced in fetal pathology. This is not only true for Germany, it is more less the situation worldwide. One means of dealing with this problem may be the use of software programs to guide inexperienced pathologists through the modern autopsy procedure. This can be combined with the use of telepathological advise given by an expert in the field. Over and above this, digitized documentation of findings can be used in research, which allows the comparison of current with previously saved cases and presents a good basis for the development of education material.

Epithelial cells are necessary building blocks of the organs they line. Their apicalbasolateral polarity, characterized by an asymmetric distribution of cell components along their apical-basal axis, is a requirement for normal organ function. Although the centrosome, also known as the microtubule organizing center, is important in establishing cell polarity the mechanisms through which it achieves this remain unclear. It has been suggested that the centrosome influences cell polarity through microtubule cytoskeleton organization and endosome trafficking. In the first chapter of this thesis, I summarize the current understanding of the mechanisms regulating cell polarity and review evidence for the role of centrosomes in this process. In the second chapter, I examine the roles of the mother centriole appendages in cell polarity during cell migration and cell division. Interestingly, the subdistal appendages, but not the distal appendages, are essential in both processes, a role they achieve through organizing centrosomal microtubules. Depletion of subdistal appendages disrupts microtubule organization at the centrosome and hence, affects microtubule stability. These microtubule defects affect centrosome reorientation and spindle orientation during cell migration and division, respectively. In addition, depletion of subdistal appendages affects the localization and dynamics of apical polarity proteins in relation to microtubule stability and endosome recycling. Taken together, our results suggest the mother centriole subdistal appendages play an essential role in regulating cell polarity. A discussion of the significance of these results is included in chapter three.

Epithelial cells are necessary building blocks of the organs they line. Their apicalbasolateral polarity, characterized by an asymmetric distribution of cell components along their apical-basal axis, is a requirement for normal organ function. Although the centrosome, also known as the microtubule organizing center, is important in establishing cell polarity the mechanisms through which it achieves this remain unclear. It has been suggested that the centrosome influences cell polarity through microtubule cytoskeleton organization and endosome trafficking. In the first chapter of this thesis, I summarize the current understanding of the mechanisms regulating cell polarity and review evidence for the role of centrosomes in this process. In the second chapter, I examine the roles of the mother centriole appendages in cell polarity during cell migration and cell division. Interestingly, the subdistal appendages, but not the distal appendages, are essential in both processes, a role they achieve through organizing centrosomal microtubules. Depletion of subdistal appendages disrupts microtubule organization at the centrosome and hence, affects microtubule stability. These microtubule defects affect centrosome reorientation and spindle orientation during cell migration and division, respectively. In addition, depletion of subdistal appendages affects the localization and dynamics of apical polarity proteins in relation to microtubule stability and endosome recycling. Taken together, our results suggest the mother centriole subdistal appendages play an essential role in regulating cell polarity. A discussion of the significance of these results is included in chapter three.

Aufgrund der rasanten technischen Entwicklung im Bereich der pränatalen Diagnostik in den letzten Jahren werden mehr angeborene Fehlbildungen zu einem früheren Zeitpunkt der Schwangerschaft und detaillierter erkannt. Das führt dazu, dass die zur Autopsie gelangenden Feten zunehmend kleiner sind und komplexere Befunde aufweisen. Gleichzeitig werden die genetischen Ursachen von mehr Erkrankungen, Fehlbildungen und Syndromen aufgedeckt, so dass die DNA-Diagnostik, Chromosomenanlyse u.a. molekulargenetische und biochemische Erkenntnisse die klinisch-genetische Diagnostik zunehmend ergänzen. Durch Festlegung einer gezielten Autopsiestrategie und Nutzung moderner Kommunikationsmittel kann der Pathologe die in den letzten Jahren gestiegenen klinischen Anforderungen erfüllen. Ein Problem ist es jedoch, dass nicht genügend Pathologen auf dem Gebiet der Fetalpathologie spezialisiert sind. Das trifft nicht nur für Deutschland zu, sondern das ist mehr oder weniger eine weltweite Situation. Ein möglicher Lösungsansatz dieses Problems wäre die Nutzung von Softwareprogrammen, die dazu dienen, unerfahrene Pathologen moderne Anleitungen während der Autopsie von Feten zu geben. Das kann mit einer telepathologischen Betreung durch einen Experten auf diesem Gebiet kombiniert werden. Darüberhinaus erlaubt die derzeitige digitalisierte Befunddokumentation den Vergleich aktueller mit gespeicherten Fällen sowie Recherchen und stellt eine gute Basis zur Erstellung von Lehrmaterial dar.
Due to the enormous technical developments in recent years in the area of prenatal ultrasound more malformations are detected in greater detail and at an earlier point in time during pregnancy than in the past. As a result, fetuses of increasingly smaller size and with complex diagnoses are presented for autopsy. At the same time, the genetic causes of ever more diseases, malformations and syndromes are being discovered, so that DNA diagnostics, chromosome analysis and, among others, molecular-genetic and biochemical investigations have come increasingly to complement clinical-genetic diagnostics. Through establishment of a clinically oriented autopsy strategy and the use of modern communication media, pathologists can satisfy the increased clinical demands of recent years. The remaining problem is that we do not have enough pathologists experienced in fetal pathology. This is not only true for Germany, it is more less the situation worldwide. One means of dealing with this problem may be the use of software programs to guide inexperienced pathologists through the modern autopsy procedure. This can be combined with the use of telepathological advise given by an expert in the field. Over and above this, digitized documentation of findings can be used in research, which allows the comparison of current with previously saved cases and presents a good basis for the development of education material.

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Esta dissertação tem como proposta discutir o papel da universidade pública brasileira na preparação do acadêmico para o mercado de trabalho. Elegeu-se o Programa de Treinamento Profissional da Universidade Federal de Juiz de Fora como objeto de análise, tendo-se restringido o estudo aos projetos de Treinamento Profissional desenvolvidos no Centro de Biologia da Reprodução. Partiu-se da questão de que o trabalho de orientação de bolsistas de Treinamento Profissional realizado no CBR proporciona, além do objetivo básico do programa, o desenvolvimento de competências profissionais de relevância para a formação do graduando. Contribuiria, dessa forma, para minimizar a tensão oriunda da transição do ambiente acadêmico para o mercado de trabalho, favorecendo a adaptação do graduado ao novo contexto de atividade. Pretendeu-se também constatar se, existindo a presença de atividades envolvendo competências profissionais, a mesma adviria de uma proposta consciente dos orientadores dos projetos, ou se ocorreria fortuitamente, fruto de ações individualizadas. As constatações apontaram para o segundo caso, ou seja, a inexistência de um trabalho unificado. Em face dessa percepção propõe-se a institucionalização, no CBR, do trabalho voltado para competências profissionais, assim como o desenvolvimento de um programa de orientação aos bolsistas. Acredita-se que o aprofundamento da questão é relevante por contribuir para a formação profissional do acadêmico e pode ser ampliada para os demais projetos de Treinamento Profissional.
This paper aims to discuss the role of the Brazilian public university in the preparation of the academic for the job market. It was elected the Professional Training Program at the Federal University of Juiz de Fora as the analysis object, having been restricted the study to the projects of Professional Training developed at the Center for Biology of Reproduction. We started from the question that the work of guidance of Professional Training collegers conducted at the CBR provides, in addition to the basic objective of the program, the development of professional skills of the utmost importance for the formation of the student. Thereby, It could contribute to minimize the stress of the transition from the academic environment to the labor market, favoring adaptation of the academic to the new context of activity. The study was also intended to see, if confirmed the presence of activities involving professional skills, if it would come from a conscious motion of the project guiders, or if occurs randomly, as the result of individual actions. The findings pointed to the second case, ie, the absence of a unified work. Given this perception, it is proposed the institutionalization, at CBR, of the work focused on professional skills, as well as the development of an orientation program for the students. It is believed that the development of this issue is relevant because it contributes to the professional training of the academic and can be extended to other projects of Professional Training.

The Cal Poly Pier (Pier) Master/Facility Plan (FP) document provides the vision of the future for the Pier, a marine science research facility. The Plan facilitates project development and management of the Pier while meeting university and department research goals. Specifically, the FP document establishes goals and strategies to direct long-term development of the Pier, streamlines agency approval and permit requirements, provides context for pier management, and assists the permitting process for future development as it relates to regulatory permits and programmatic growth on the Cal Poly Pier to help meet goals of the Center for Coastal Marine Sciences (CCMS). The Cal Poly Pier is the marine field station for the California Polytechnic State University San Luis Obispo (Cal Poly) CCMS and is one of several facilities that supports research and educational activities. The CCMS is a CSU Campus Center research organization that provides research and education activities as a part of Cal Poly’s overall mission while offering opportunities to interested parties beyond Cal Poly, such as private and public entities. The 3,057-foot long pier provides students, faculty, researchers, and other users unrivaled access to the marine environment of the Central Coast and fosters hands-on learning opportunities to progress marine research and science. The Master Plan name was changed to Facility Plan to streamline the plan approval process and to minimize the potential for errors.

This study evaluated a novel perinatal health education program, entitled Preparing for Caring, teaching infant touch and handling skills for those who care for babies. Sensitive, attuned, and responsive interactions between caregivers and infants are crucial for the healthy development of foundational brain architecture. Research has confirmed that evidence-based perinatal health programs have the potential to increase the caregiver’s satisfaction and self-efficacy, to reduce anxiety about caring for a newborn, to increase the caregiver’s capacity to form an attachment bond with the infant, and to promote the infant’s health and development. The present study built upon evidence that nurturing touch is positively associated with brain development which positively impacts behavior, cognition, and the health trajectories of children from low-income urban and minority families who are more likely to experience disparities in lifespan health, including increased infant and maternal mortality. Caring and stimulus-rich environments, especially those promoted in intergenerational programs such as Early Head Start, offer “the most compelling evidence” for producing positive changes in both parents and children. The current study, providing perinatal parenting education within an urban Early Head Start (EHS) setting, was hypothesized to effect changes in parents that will positively influence their parenting skills during a critical period of neural and emotional growth and thereby positively influence their children’s development. Mixed-methods data were collected from EHS early childcare educators, parents and community caregivers, administrators, and the program developers. Analyses evaluated program fidelity, specifically in its translation to a new population and setting; identified barriers to and facilitators of implementation of the program; identified which program components were most likely to be accepted and incorporated into daily use by participants; and conducted and shared the results of a pilot study on what an impact evaluation of what participating in this program might look like for key outcomes (including caregiver self-efficacy and maternal self-esteem).

The roles of interleukin-1$\beta$ (IL-1$\beta$) after prenatal trauma in the perinatal brain were studied by lesioning fetal mice in utero at embryonic day 18 (E18) and examining them at postnatal days zero and six (P0 and P6), corresponding to 2 and 9 days postlesion, respectively The regulation of glia limitans reformation by IL-1$\beta$ following neural trauma in the prenatal mouse brain was examined by immunocytochemistry and computer-assisted image analysis. Studies using the human recombinant IL-1 receptor antagonist (IL-1ra), a competitive inhibitor of the IL-1 neural receptor, revealed that IL-1$\beta$ binding to neural IL-1 receptors was necessary for astrocytes to form the new glia limitans that ultimately closed off exposed neural tissue Significantly increased IL-1$\beta$ immunoreactivity was measured in vehicle-injected or nontreated prenatally lesioned animals at the lesion site and in neural parenchyma near the lesion. IL-1$\beta$ immunoreactivity significantly decreased in animals treated with IL-1ra. Both human recombinant IL-1$\beta$ and IL-1ra + IL-1$\beta$ combination injections restored IL-1$\beta$ immunoreactivity upregulation in perilesional tissue Upregulation of IL-1$\beta$ immunoreactivity was due, at least in part, to local IL-1$\beta$ mRNA production, as shown by in situ hybridization. IL-1$\beta$ mRNA was significantly upregulated in cells of varying morphologies near the lesion site. IL-1ra and IL-1$\beta$ injections significantly increased IL-1$\beta$ mRNA in cells near the lesion site as compared to vehicle-injected animals. When IL-1ra and IL-1$\beta$ were injected simultaneously, IL-1$\beta$ mRNA levels were not significantly different from vehicle-injected animals Finally, the role IL-1$\beta$ might play in cell death following traumatic perinatal brain injury was examined. Animals treated with IL-1ra showed significantly increased areas of total cell death and apoptosis when compared to vehicle. In contrast, animals treated with IL-1$\beta$ showed significantly decreased areas of total cell death and apoptosis as compared to IL-1ra-treated animals. The addition of IL-1$\beta$ to IL-1ra treatment at the time of injection attenuated the increase in both types of cell death induced in IL-1ra-treated animals Viewed together, these studies provide evidence that IL-1$\beta$ binding to the neural IL-1 receptor is an important signal to activate cellular survival and activity culminating in successful healing and repair in the damaged perinatal mouse brain
acase@tulane.edu

The prairie and bordering woodlands of the Armand Bayou Nature Center, Harris County, Texas were sampled as an example of Texas Upper Coastal Prairie. The prairie is homogeneous and species-rich but shows very low dominance. Paspalum plicatulum, Dichanthelium spp., Carex cherokeensis, Andropogon virginicus, and Schizachyrium scoparium are the dominant graminoids. This prairie resembles the typical Upper Coastal Prairie of Texas but contains a greater number of eastern species with few southern or southwestern influences. Baccharis halimifolia has invaded the prairie and prairie climax species are no longer dominant. There is little evidence of a reduction in diversity due to brush encroachment. The woodlands are an oak and elm dominated riparian forest of the Upper Coastal Prairie with an important shrub component. Quercus phellos, Q. falcata, Ulmus americana, U. alata and U. crassifolia are common canopy species. Ilex vomitoria, Viburnum dentata, Callicarpa americana and Ligustrum sinense are the dominant shrubs.

The relationship between gene copy number and protein expression levels has not thoroughly been examined in humans or mouse models of Down syndrome (DS) in relationship to developmental changes in the trisomic brain. Found on human chromosome 21 (Hsa21) and triplicated in DS, Dual-specificity tyrosine-phosphorylated regulated kinase 1A (DYRK1A) has been linked in DS to neurological deficits by restricting cell growth and proliferation. Little information exists regarding DYRK1A during perinatal development and how its expression may lead to cognitive deficits, and none exists that explores the gene-to-protein relationship during these critical time periods. This study aims to 1) Quantify variable DYRK1A expression across development as a function of age, sex, and brain region in trisomic Ts65Dn mice compared to euploid counterparts and 2) establish that the spatiotemporal pattern of developmental DYRK1A in the brain is not influenced solely by gene copy number, and that reduction of Dyrk1a in euploid and trisomic mice does not result in a corresponding global reduction of DYRK1A expression. DYRK1A was quantified in three areas of the postnatal brain at seven ages using the Ts65Dn mouse, the most studied model of DS, and found that trisomic expression is significantly increased on postnatal day ([P]6), declining by the third week to near euploid levels. We also uncovered a sexual dimorphic expression of DYRK1A when comparing animals of different sexes within the same genotype. Data from Dyrk1a knockdown mice indicated that reducing only Dyrk1a in euploid and in otherwise trisomic animals yields highly variable levels of DYRK1A, dependent on sex and tissue type, supporting the non-intuitive relationship between gene dosage and protein expression. These data emphasize the need to understand the age-dependent regulation of antecedent conditions that are causing changes in Dyrk1a expression in the brain.

The field of "perinatal programming" has increasingly implicated an adverse early-life environment in the etiology of many chronic health problems and mental disorders. The following dissertation research is based on the hypothesis that the programming of brain and behavior by an altered early-life environment is propagated by inflammatory mechanisms in the placenta and developing brain. Offspring outcomes of two different maternal environmental exposures--air pollution and a "Western diet" (both highly relevant for the modern world)--were assessed in a mouse model in order to identify mechanisms common to developmental programming more generally.

The first set of experiments characterized the long-term behavioral and metabolic consequences of prenatal air pollution exposure in adult offspring. The male offspring of diesel exhaust particle (DEP)-exposed dams were predisposed to obesity, insulin resistance, and increased anxiety following placement on a high-fat diet (HFD) in adulthood. Furthermore, DEP/HFD male offspring exhibited evidence of macrophage priming, both in microglia and peripheral macrophages. The next experiment examined whether prenatal air pollution exposure could also synergize with a simultaneous "second hit" (i.e., maternal stress) during gestation. The offspring of mothers exposed to both air pollution and stress during gestation were more anxious as adults, but only the male offspring of this group also exhibited impaired cognition, in conjunction with neuroinflammatory changes. A further experiment revealed that prenatal air pollution exposure altered microglial maturation in a TLR4- and sex-dependent manner, consistent with the previous results. However, we found limited evidence of a placental immune response to DEP, potentially due to analysis too late in gestation.

The second set of experiments characterized the enduring behavioral and metabolic consequences of maternal consumption of a "Western diet" (HFD in combination with BCAA supplementation) prior to and during gestation and lactation. The adult offspring of HFD-fed dams were more anxious in adulthood, despite being placed on a low-fat diet at weaning. Male HFD offspring were also hyperactive, whereas female HFD offspring exhibited more severe metabolic disturbances. Furthermore, there was evidence of microglial priming and peripheral macrophage priming in male HFD offspring, similar to the prenatal air pollution model. The next experiment also found evidence of altered microglial development due to maternal HFD, in conjunction with widespread, sex-specific immune gene regulation in the placenta in response to maternal diet. Moreover, maternal HFD decreased placental serotonin production, and also programmed long-term alterations in serotonergic function in the prefrontal cortex of adult HFD offspring. Taken together, these experiments define sexually dimorphic innate immune mechanisms in the placenta and developing brain that may underlie the long-term metabolic and behavioral consequences of maternal environmental exposures.

Dissertation

Dynamic interactions between the amino acid acceptor end of tRNA and the ribosome underlie the synthesis of successive peptide bonds at the peptidyl transferase center (PTC) of the 50S ribosomal subunit. Photo-crosslinking of the 3′-terminal nucleotide of tRNA, which is adjacent to the attached amino acid or peptide, to components of the 50S subunit has proven to be a sensitive means for identifying specific protein and RNA segments in close proximity to the site of peptide bond formation. I have used this approach to follow changes in the position of the tRNA during peptide bond formation using several photoreactive tRNA-derived ligands. Three new photoreactive tRNA derivatives have been synthesized for use as probes of the PTC of the ribosome. In two of these derivatives, the 3′ adenosine in position 76 of yeast tRNAPhe has been replaced by either 2-azidodeoxyadenosine or 2-azido-2′-O-methyladenosine, while in a third the 3′-terminal 2-azidodeoxyadenosine of the tRNA is joined to puromycin via a phosphoramidate linkage to generate a photoreactive transition-state analog. All three derivatives bind to the P site of 70S ribosomes with affinities similar to that of unmodified tRNA Phe and can be crosslinked to components of the 50S ribosomal subunit by irradiation with near UV light. Yeast tRNAPhe containing 2-azidoadenosine, [2N3A76]tRNAPhe, typically crosslinks to the N-terminal sequence of protein L27 as well as to nucleotides U2506 and U2585 of the 23S rRNA. While the photoreactive transition-state analog, [2N3dA76]tRNAPhe-p-Puro, crosslinked the same components as [2N3A76]tRNAPhe, the distribution of crosslinks is altered significantly. The crosslinking to nucleotide U2506 is strongly reduced, and two new crosslinked nucleotides A2450 and A2602 were detected. Characteristic differences in the crosslinking patterns suggest that these tRNA derivatives can be used to follow subtle changes in the position of the tRNA relative to the components of the PTC.

Eukaryotic cells contain scores of small nucleolar RNAs (snoRNAs) that are complexed with proteins in particles known as snoRNPs. SnoRNPs function in processing of ribosomal RNA (rRNA) and nucleotide modification of ribosomal and other RNAs that pass through the nucleolus. The modifications involved are: (1) isomerization of uridine to pseudouridine (Ψ), and; (2) methylation of the 2′-hydroxyl of ribose moieties (Nm). The Ψ and Nm modified nucleotides are the most abundant in rRNA. The purpose of the thesis work is to obtain a better understanding of the synthesis of snoRNPs and the function(s) of the modifications they form. To this end, three lines of research were carried out. Two focus on different snoRNA-associated proteins. The third examines the effect of snoRNP-mediated pseudouridylation of rRNA on ribosome structure and function. One of the snoRNA-associated proteins studied is a conserved helicase previously isolated from a mouse nuclear extract on the basis of association with a conserved snoRNA structure known as the box C/D motif. Here, it was shown that the yeast ortholog of this protein (Rvb2p) is required for production of snoRNAs (both major families) and for localization of snoRNP proteins. The findings are consistent with a role for Rvb2p in the assembly or trafficking of snoRNPs. A second project was aimed at determining if a particular H/ACA snoRNP protein is the enzyme that forms Ψ in rRNA. The protein, Cbf5p, was previously shown to have limited sequence homology with known Ψ synthases (96). Here, in collaboration with the John Carbon laboratory, it was determined that point mutations in Cbf5p in this region of homology can abolish in vivo pseudouridylation of rRNA, arguing that Cbf5p is indeed the global Ψ synthase. Finally, the function of Ψ in the peptidyl transferase center (PTC) was investigated by depleting cells of snoRNPs that modify this domain, and evaluating the under-modified ribosomes. Interestingly, cells lacking Ψs in the PTC are impaired in growth and protein synthesis. Chemical probing revealed changes in the structure of the mutant ribosomes. Enhanced reactivity with dimethylsulfate was observed for nucleotides predicted to influence binding of elongation and initiation factors. The results demonstrate that the structure and activity of yeast ribosomes depend on pseudouridylation of the PTC. The possibility that guide snoRNPs have function(s) other than modification is discussed.