No description available.

At embryonic day 3.5 (E3.5), inner cell mass (ICM) cells co-express the transcription factors NANOG and GATA6. Between E3.5 and E4.5, cells of the ICM differentiate into epiblast (Epi) and primitive endoderm (PrE). These two lineages are distinguished by the differential expression of the previously coexpressed transcription factors; Epi cells express NANOG while PrE cells express GATA6. FGF/ERK signalling is responsible for Epi and PrE differentiation but it does not explain the initial co-expression of both factors and how the mutually exclusive expression arises. β-catenin is the downstream effector of Wnt signalling, and it is also found in the membrane forming a complex with E-cadherin. Depending on it subcellular location, β-catenin has been associated with pluripotency and differentiation of mESCs, whose origin is the mouse embryo. My hypothesis was that changes in both cellular pools of β-catenin are involved in ICM differentiation. To characterize Wnt/β-catenin role during preimplantation development, I applied quantitative immunofluorescence analysis (QIF) together with chemical and classical genetics in in vitro and in vivo models. I found that high membrane β-catenin levels are associated with Epi cells from E4.0 stage, while nuclear β-catenin levels are higher in co-expressing cells at E3.5 and PrE precursors at E4.0. My results indicate that increases in nuclear β-catenin levels allow the ICM cells to be specified earlier, determined by an earlier appearance of mutually exclusive expression of GATA6 and NANOG in vitro and ex vivo. Moreover, increased β-catenin levels promote specification towards PrE fate, observed by the presence of higher percentages of PrE cells. Conversely, a decrease in β-catenin levels result in slower ICM specification into Epi and PrE. Finally, modulation of FGF/ERK signalling in mouse embryos, which is the main pathway in this cell fate choice, led to changes in β-catenin subcellular location and levels. Altogether, my results are consistent with a role for Wnt/β-catenin signalling facilitating PrE fate acquisition concomitantly with FGF/ERK signalling.

The purpose of this study was to investigate the relationships between adult children of alcoholics (ACOA's) recollections of their family of origin home environments, with present day levels of ACOA differentiation of self, and trait anxiety. The author explored the influence of ACOA gender differences and a variety of demographic information.;The ACOA volunteers for this study were recruited from The College of William and Mary in Virginia. to be included in the study, subjects must have met the following criteria: (a) the volunteer's age must have been 18 years or above; (b) subjects met the ACOA criteria by scoring a 6 or above on the Children of Alcoholics Screening Test.;Volunteers completed paper and pencil test packages as the data gathering technique for this study. The following instruments were included in the test package: (1) State-Trait Anxiety Inventory; (2) Differentiation of Self Scale; (3) Children of Alcoholics Screening Test; (4) Family Environment Scale; and (5) a Personal History Questionnaire.;It was hypothesized that: (1) ACOA's would demonstrate a significant relationship between perceived past family environments and levels of differentiation of self; (2) ACOA's would demonstrate a significant relationship between perceived past family environments and levels of trait anxiety; (3) an inverse relationship would exist between differentiation of self and trait anxiety; (4) ACOA's were expected to have lower levels of cohesion and expressiveness, and higher levels of conflict; (5) ACOA's were expected to be less independent, achievement orientated, intellectual, and show less interest in recreational activities; and (6) ACOA's were expected to have low degrees of organization and high degrees of control.;It was concluded that all six significant relationships did exist.;Future studies are needed to: improve on sampling techniques; account for distorted perceptions of the past and personality changes due to the passage of time; and explore the influence of the alcoholic parent's gender on the ACOA's subsequent development.

No description available.

Prevalence of diabetes, especially type 2 diabetes mellitus (T2DM) is increasing worldwide. Millions of people are already affected by T2DM and estimates predict over half a billion people will likely be suffering from the disease by 2030. T2DM is associated with an increased risk of developing cardiovascular disease. Cardiovascular dysfunction is the leading cause of mortality among type 2 diabetics. Treatment for T2DM has changed over time. Though it was once known as insulin independent, a large portion of type 2 diabetics are now treated with insulin injections. However, type 2 diabetics treated with insulin are more likely to suffer from heart complications. Due to this, we sought to determine the specific effect of insulin and insulin-induced ceramide accrual on heart mitochondrial bioenergetics. To do so we used both in vitro and in vivo models. H9c2 cardiomyocytes and adult male mice were treated with insulin with or without the ceramide biosynthesis inhibitor myriocin. Mitochondrial bioenergetics were determined in permeabilized cardiomyocytes and myocardium. In this study we demonstrate that insulin induced ceramide accrual in both isolated cardiomyocytes and whole murine myocardium. We further found that insulin treatment is sufficient to disrupt mitochondrial respiration in both models. Inhibition of the ceramide accrual rescued mitochondrial respiration, indicating that ceramide is necessary for the insulin-induced alterations in heart mitochondrial respiration. These results suggest that insulin has a role in the development of heart complications associated with T2DM due to cardiomyocyte mitochondrial disruption. They also implicate ceramide as a possible mediator in the development of insulin-related heart disorders.

More conservation programs directed towards insects are needed in order to prevent the continue decline in which many insect species in the world are heading to. New ways to use the remaining microhabitats are needed to mitigate the fragmentation and degradation that urbanization and modern agriculture have contributed to in the southern Swedish landscapes. This study focusses on how microhabitats like road-verges can help the conservation projects of pollinating insects. To increase the knowledge about three groups of pollinators (Apidae, Rhopalocera and Syrphidae) and their distribution between two different microhabitats, a malaise trap was set up in road verges and in sample locations further away from the road-verges. The result from this study shows that road-verges are a microhabitat that could work as mitigation to the decline of some pollinating species, if right actions are taken. This study shows that Syrphidae was most acceptant to the hostile environment close to the roads. If these actions are going to help the most treated species is still unknown.

Variously aged (C57BL X C3H)F1 hybrid mice were used as recipients in experiments to determine if spleen or bone marrow cells from normal C57BL or C3H donors, spleen or bone marrow cells from C57BL donors previously treated with cortisone, or C57BL bone marrow cells previously incubated with serum from X-irradiated C57BL mice could induce a graft-versus-host reaction (GVHR). The mice were assayed for GVHR by mortality or splenomegaly as detected by the discriminant spleen assay developed by Simonsen. Results show that spleen cells from normal C57BL, normal C3H, or cortisone-treated C57BL mice are capable of inducing a GVHR in F1 hybrids, resulting in death or splenomegaly, whereas bone marrow cells from normal C3H, normal C57BL, or cortisone-treated C57BL mice were unable to induce a GVHR in F1 hybrids. It was also found that sera from lethally irradiated C57BL mice did not cause C57BL bone marrow to induce a GVHR in F1 hybrids.

Coronary heart disease is a prevalent issue in developed countries throughout the world. It can have crippling effects on the quality of life and even lead to mortality, in the case of myocardial infarction. Part of the problem is the lack of a robust regenerative response in mammals after injury. Zebrafish have an amazing ability to regenerate after injury, and studies have demonstrated that the regenerative response recapitulates embryonic development. Our lab previously reported the first analysis of coronary vessel development in zebrafish and demonstrated that coronary endothelial cells undergo angiogenesis to form a vascular network. The roles of perivascular cells in this process have not been examined in zebrafish. Using a transgenic reporter line marking pdgfrb expression, I found that pdgfrb is first observed in epicardium at the AV canal. At later stages of coronary vessel development, pdgfrb positive cells become localized to the perivascular region of mature vessels. I also observe that early in development, Tcf21 and pdgfrb co-express, which suggests a close relationship between the epicardium and pdgfrb+ cells. Previous findings from our lab revealed that cxcl12b+ cells localize to large coronary vessels during development. My findings reveal that pdgfrb+ marks perivascular cells of both capillaries and large coronary vessels. Lineage tracing analysis revealed that a subset of pdgfrb+ perivascular cells derive from tcf21 labeled epicardial cells. To see if disruption of Pdgfrb signaling impacts coronary development, I examined pdgfrb mutant hearts. In the Pdgfrb mutant, a mature coronary vessel network fails to form, and instead we observe isolated endothelial cell islands. Lastly, I characterized a transgenic line that expresses a dominant negative form of Pdgfrb (dnpdgfrb) and can be potentially used for later developmental and/or regenerative studies. My findings indicate strong dnpdgfrb induction can be achieved at adult stages. My studies will greatly enhance our current understanding of coronary vessel development, and can be used as the basis for studying perivascular cells and their interactions with endothelial cells after cardiac injury in regeneration.

Though it is well known that tobacco related products can cause prenatal maldevelopment, very little is known on how tobacco products affect bone tissue as it develops in the embryo. Identifying which chemicals can induce the greatest harm to the prenatal skeletal system is an improbable task as there are over 7,000 chemicals in tobacco smoke alone. We hypothesized that the Toxicological Priority Index (ToxPI) program can be used to rank osteogenic cytotoxicity potential to aid in the assessment of what chemicals out of the thousands can cause osteogenic differentiation inhibition. ToxPI aggregates information from various assays and incorporates them into visual “pie charts” which allow chemicals to be ranked against each other by given parameters. The larger the pie chart the greater likelihood of potential effects and vice versa. Seventeen tobacco chemical constituents were ranked using ToxPI and those chemicals with pie charts (0 To assess the ability of ToxPI to correctly predict maldevelopment in silico eight compounds were then tested in vitro: four of them being ToxPi positive and the other four having null predicted effects. To verify the predictions, human embryonic stem cells were differentiated into osteoblasts and exposed to various concentrations of each compound. Cell viability was measured via MTT assay in conjunction with a calcium assay to measure osteogenic differentiation. In addition, adult human feeder fibroblasts cell viability in response to exposure was measured. ToxPI positive predictions (xin vitro, caused differentiation inhibition. Together our data suggests that ToxPi might be useful to identify strongly inhibitory chemicals based on their cytotoxicity but might also give false negative results for chemicals that cause differentiation inhibition at sub-toxic levels.

Development in the annual killifish Austrofundulus limnaeus can follow two distinct developmental trajectories. Typical development includes the entrance of embryos into a state of metabolic and developmental arrest termed diapause. Alternately, embryos can escape diapause and develop directly without pause. These two trajectories are characterized by differences in the rate and timing of developmental, morphological, and physiological traits. Insulin and Insulin-like growth factor (IGF) signaling (IIS) is known to regulate entrance into diapause in a variety of invertebrates. In this thesis I explore the possible role of IGFs in the regulation of development and diapause in embryos of A. limnaeus. Here I report stage-specific expression of IGF-I and II proteins and their associated mRNA transcripts. Patterns of IGF-I protein expression are consistent with IGF signaling playing a major role in supporting the escape trajectory. In addition, treatment of embryos with a potent inhibitor of the IGF-I receptor (IGF1R) mimics the diapause developmental pattern even under conditions that should favor direct development. Evaluation of mRNA gene expression patterns in the two developmental trajectories suggests a role for IGF-I signaling through the RAS-MAPK-ERK pathway, which may be promoting the escape phenotype. Additionally, IGF-I activity may be enhanced in escape trajectory embryos though upregulation of IGF binding protein 2 (IGFBP-2) mRNA. These data suggest a major role for IGF signaling in the promotion of the escape trajectory, and thus we predict that specific mechanisms are in place in diapause-bound embryos that block IGF signaling and thus promote entrance into diapause. The data presented here suggest that blocking IGF signaling is critical for induction of diapause, but also suggests that other signaling pathways are likely also at play. Other pathways such at the TGF-beta signaling molecules and SMAD pathway, may also be involved in the direct regulation of the diapause phenotype, as has been shown for other animal models of developmental arrest.