Thesis (Ph. D.)--University of Adelaide, 1987. / Includes bibliographical references (leaves [11-1]-11-19).

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Made available in DSpace on 2014-10-09T12:54:42Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:08:00Z (GMT). No. of bitstreams: 0 / Até pouco tempo, o conhecimento da contaminação dos ecossistemas aquáticos, estava restrito a metais e agrotóxicos, sendo pouca ênfase dada a produtos farmacêuticos, que, apesar de serem encontrados em concentrações de nanogramas a microgramas, podem causar efeitos letais e induzir malformações nos organismos. Estudos ecotoxicológicos têm sido realizados para identificar os efeitos destas substâncias, contudo, os dados são insuficientes, principalmente no que se refere a efeitos subletais e de toxicidade em espécies de ambientes tropicais e subtropicais. Efeitos letais e subletais, como crescimento e reprodução, são determinados através de ensaios ecotoxicológicos agudos e crônicos, sendo as malformações dificilmente evidenciadas por estes ensaios. Diante disto, pesquisadores vêm desenvolvendo metodologias mais refinadas, como por exemplo, a exposição direta de embriões a produtos químicos em ensaios de teratogênese. Para Cladocera estes ensaios são restritos. O objetivo deste trabalho foi estudar os efeitos letais e subletais de um antimicrobiano, Triclosan, através de ensaios de ecotoxicidade com Daphnia similis, Ceriodaphnia. dubia e C. silvestrii. A fim de se estimar com maior rigor os efeitos em ambientes tropicais e subtropicais, os ensaios foram realizados com água natural como água de diluição, com e sem fotoperíodo. Também foram realizados ensaios em água Milli- Q água reconstituída, sem fotoperíodo de modo a observar a toxicidade do produto sem interferentes externos. Estágio do desenvolvimento embrionário de D. similis, C. dubia e C. silvestrii foram identificado de modo a otimizar os ensaios de teratogênese. Para o estudo da toxicidade, foram realizados ensaios de ecotoxicidade aguda com D. similis, C. dubia e C. silvestrii e ensaios crônicos com D.similis e C. silvestrii. Para a classificação dos estágios do desenvolvimento embrionário, ovos foram cultivados in vivo a temperatura de 20°C (D. similis) e 25°C (C. dubia e silvestrii), observados a cada hora, até o desenvolvimento completo do organismo. Posteriormente, as condições dos ensaios de teratogênese foram estabelecidas sendo realizados com D. similis. A CE50 48H de Triclosan para D.similis e C. silvestrii em água natural reconstituída com fotoperíodo foi de 0,23 e 0,10 mg.L-1, respectivamente, e de 0,13 mg.L-1para C. silvestrii, sem fotoperíodo. Para os ensaios com água Milli-Q reconstituída, sem fotoperíodo, os valores de CE50;48H para D. similis, C. dubia e C. silvestrii foram de 0, 22; 0,09 e 0,08 mg.L-1, respectivamente. O CEO para D. similis foi de 0,1 mg.L-1 e a o valor de IC50 relativo às malformações das neonatas nos testes crônicos foi de 0,057 mg.L-1. Para C.silvestrii o CEO foi de 0,04 mg.L-1, não sendo evidenciadas malformações nas neonatas. Sete estágios foram observados no desenvolvimento embrionário dos cladoceros com duração (horas) de 34 (±3) para C. dubia, 36(±2) para C. silvestrii e de 51 (±5) para D. similis. O valor de IC50 para malformação nos ensaios de teratogênese (exposição direta de embriões) para D. similis foi de 0,075 mg.L-1, próximo ao obtido nos ensaios crônicos / Dissertação (Mestrado) / IPEN/D / Instituto de Pesquisas Energéticas e Nucleares - IPEN/CNEN-SP

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Each year, 7.9 million babies are born with birth defects. Seventy percent of those could be prevented, ameliorated, or repaired; yet 3.2 million children still die by the age of three (March of Dimes Global Report 2006). We have found that non-specific maternal immune stimulation with the cytokine interferon-gamma (IFN-gamma) can successfully ameliorate some of these defects in the C57BL/6N mouse model. We have observed a reduction in the distal limb malformations syndactyly, polydactyly, and webbing by 47%, 100%, and 63% respectively when IFN-gamma is given 2 days prior to MNU administration. We have also observed that IFN-gamma works at the placental level to protect against MNU-induced damage. Trophoblast loss and associated cytokine alterations occur in gestation day (GD) 14 placenta following GD9 MNU exposure, showing that fetal-maternal communication can be hindered due to MNU. In the labyrinthine layer of the placenta, we observed multifocal fibrinous necrosis of endothelial cells due to MNU, however IFN-gamma almost completely protected the trophoblast and endothelial cells when given to the dam as an immune stimulant. To determine the genes participating in these processes, gene microarray studies were conducted. Hepatocyte growth factor (HGF), interleukin 1 beta (IL1Β), and insulin-like growth factor 2 (IGF2) were elucidated as genes that were significantly expressed in GD12 placenta. These genes are similar in that they are all connected to the Jak-Stat signaling pathway. These findings provide a possible mechanism for birth defect reduction by maternal immune stimulation with IFN-gamma in MNU-challenged mice. / Master of Science

<p>Heavy metals polluting our environment cause concern for developing organisms. Among them, cadmium with extremely slow elimination from the body, causes lower birth weight in humans but has not been classify as a human teratogen. Studies in different laboratory animals have shown that cadmium indeed is a potent teratogen. Exposure to cadmium during early mouse embryonic stages (e.g. day 7-8 post-coitus) interferes with the closure of the anterior neural pore producing exencephalic embryos. The underlying mechanisms are not understood, but the heavy accumulation of cadmium in extra- and intraembryonic endoderm and chorioallantoic placenta, however not in the neuroepithelium, suggests that the effects on neural tube closure is due to indirect mechanisms. In this thesis, the disruption in the mouse embryo at the time of neural tube closure of the hierarchies of some signalling pathways and gene regulatory networks that control embryonic development has been studied after cadmium exposure. Cadmium was shown to cause DNA damage as measured by Comet assay, and to activate genes and proteins in the apoptotic pathways (<i>p53, p21, Bcl-2, Bax</i>, and caspase-3), increasing the number of apoptotic cells mostly in areas of physiological cell death, especially in the neuroepithelium. Many of these effects could be reversed by zinc pre-treatment, known to counteract the teratogenic effect of cadmium. Cadmium was also shown to affect Zn-transport and –regulatory proteins in the embryo, but perhaps more importantly in yolk sac placenta, and in the decidua (ZnT-1, MT-I, and ZIP-4). Using gene arrays, cadmium was found to considerably affect gene expression of rather few genes, such as those of metallothioneins and stress-related proteins, supporting in principle an extraembryonic site of action of cadmium. In addition, a number of genes expressed in the anterior visceral endoderm (<i>Hesx1, HNF3β, Cerl, Otx2</i> and <i>Sox2</i>) where cadmium accumulates, and known to signal to the anterior neuroepithelium, was affected by cadmium. This finding may suggest a new principle for chemical teratogenesis.</p>

Heavy metals polluting our environment cause concern for developing organisms. Among them, cadmium with extremely slow elimination from the body, causes lower birth weight in humans but has not been classify as a human teratogen. Studies in different laboratory animals have shown that cadmium indeed is a potent teratogen. Exposure to cadmium during early mouse embryonic stages (e.g. day 7-8 post-coitus) interferes with the closure of the anterior neural pore producing exencephalic embryos. The underlying mechanisms are not understood, but the heavy accumulation of cadmium in extra- and intraembryonic endoderm and chorioallantoic placenta, however not in the neuroepithelium, suggests that the effects on neural tube closure is due to indirect mechanisms. In this thesis, the disruption in the mouse embryo at the time of neural tube closure of the hierarchies of some signalling pathways and gene regulatory networks that control embryonic development has been studied after cadmium exposure. Cadmium was shown to cause DNA damage as measured by Comet assay, and to activate genes and proteins in the apoptotic pathways (p53, p21, Bcl-2, Bax, and caspase-3), increasing the number of apoptotic cells mostly in areas of physiological cell death, especially in the neuroepithelium. Many of these effects could be reversed by zinc pre-treatment, known to counteract the teratogenic effect of cadmium. Cadmium was also shown to affect Zn-transport and –regulatory proteins in the embryo, but perhaps more importantly in yolk sac placenta, and in the decidua (ZnT-1, MT-I, and ZIP-4). Using gene arrays, cadmium was found to considerably affect gene expression of rather few genes, such as those of metallothioneins and stress-related proteins, supporting in principle an extraembryonic site of action of cadmium. In addition, a number of genes expressed in the anterior visceral endoderm (Hesx1, HNF3β, Cerl, Otx2 and Sox2) where cadmium accumulates, and known to signal to the anterior neuroepithelium, was affected by cadmium. This finding may suggest a new principle for chemical teratogenesis.

Exposure to the anticonvulsant valproic acid (VPA) is associated with a 7.5% rate of major malformations and a 1-2% incidence of neural tube defects (NTDs). Although the teratogenic outcomes resulting from VPA use during pregnancy were first identified in the 1980s, the mechanisms by which VPA induces birth defects are not fully elucidated. Based on evidence in the literature, the studies in this thesis examined the role of in utero VPA exposure on oxidative stress and epigenetic alterations in the developing embryo to provide further mechanistic insight into VPA’s teratogenic pathway. The first study investigated the role of oxidative stress in VPA-induced teratogenesis. Using CD-1 mice, catalase was shown to protect against VPA-induced effects on developmental and morphological parameters in both whole embryo culture and in vivo models. Studies in whole embryo culture demonstrated that markers of oxidative damage were not altered by VPA; however, VPA increased apoptosis in the neuroepithelium, which was attenuated by the addition of catalase. The second objective addressed epigenetic modifications induced by VPA in an in vivo mouse model. Maternal administration of VPA resulted in increased acetylation of histones H3 and H4, increased methylation of histone H3K4, and decreased methylation of histone H3K9. Furthermore, these changes were localized to VPA target tissues including the neuroepithelium, heart, and somites. Global DNA methylation in the embryo was not altered by VPA. The final objective was to determine VPA’s effect on a marker of DNA damage, markers of cell cycle proteins, and a marker of apotosis in vivo. Maternal administration of VPA resulted in a rapid increase of γH2A.X, a marker of DNA double strand breaks (DSBs). Increased expression of p27KIP1, a cyclin-dependent kinase inhibitor, and activated caspase-3, a marker of apoptosis, were observed and these changes were localized to the neuroepithelium of developing embryos. In conclusion, this thesis supports the hypothesis that VPA-induced increases in ROS production and HDAC inhibition may lead to altered gene expression patterns and consequently teratogenic effects, namely NTDs. / Thesis (Ph.D, Pharmacology & Toxicology) -- Queen's University, 2011-05-24 13:12:33.778

Includes bibliographic references. / 1 v. (various foliations) : ill. ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Details the results of in vivo and in vitro studies into the effects of zinc deficiency, hypervitaminosis A, alcohol, nicotine and salicyclic acid, at the morphological and ultrastructural levels in the developing embryo. / Thesis (Ph.D.)--University of Adelaide, Dept. of Anatomy and Histology, 1992