No description available.

Includes bibliographies. Aims to identify homeobox genes which may have a developmental role during early embryogenesis by the characterization of homeobox gene expression in undifferentiated ES cells, and in a range of differentiated ES cell derivatives.

Le développement dune placentation hémochoriale requiert une invasion trophoblastique de lendomètre maternel jusquà ses couches profondes ainsi quune érosion des vaisseaux maternels jusquau niveau du tiers supérieur du myomètre. Les cellules trophoblastiques envahissent et modifient les vaisseaux utérins pour permettre une augmentation du flux sanguin vers lunité foeto-placentaire. Dautre part, lembryon qui simplante va non seulement se connecter aux vaisseaux maternels mais aussi générer des vaisseaux de novo. Le processus dangiogenèse endométriale au niveau du site dimplantation est modulé spatio-temporellement par un dialogue complexe entre des facteurs endocrines, paracrines et autocrines. Parmi ces facteurs, lhCG, sécrétée précocement par le trophoblaste, va tenir un rôle de choix. Outre son rôle lutéotrophique bien décrit, lhCG promeut linvasion trophoblastique et présente une action clé sur la tolérance maternelle de lembryon. Nos travaux ont mis en évidence un rôle direct de lhCG sur langiogène endométriale ainsi quun rôle indirect via une boucle paracrine par activation du récepteur hCG/LH à la surface des cellules endothéliales et épithéliales de lendomètre et une stimulation de lexpression du VEGF dans les cellules épithéliales sous linfluence de lhCG. Grâce à de nombreux modèles dangiogenèse in vitro, in vivo et ex-vivo, nous avons décrypté la signalisation cellulaire mise en uvre lors de la liaison de lhCG à son récepteur. De plus, nos travaux démontrent un rôle artériogène inattendu de cette hormone. Ceci revêt toute son importance dans létude de leffet angiogène de lhCG lié à des processus physiologiques comme langiogenèse au niveau du site dimplantation, pathologiques comme leffet tumorigène de lhCG sécrété en fortes quantités lors de pathologies trophoblastiques.

A Disintegrin-like And Metalloproteinase with Thrombospondin (TSP)-Type I sequence motifs 9 (ADAMTS9) is widely expressed in mouse and human fetal tissues. ADAMTS9 null mice cannot survive beyond E7.5 and its haploinsufficiency is associated with cardiac and aortic anomalies. This project hypothesized that ADAMTS-9 plays an important role during early embryogenesis. By using human embryonic stem cells (hESCs) as a model, the objectives of this study were to study the expression of ADAMTS9 and to determine the effects of ADAMTS9 perturbation on hESC differentiations. The expression of ADAMTS9 was compared between undifferentiated and differentiated hESCs. Its mRNA was maintained at similar levels in different passages of normal hESC lines. Interestingly, significantly lower expression was detected in karyotypically abnormal VAL4A when compared to the normal cells (H9 and VAL3). ADAMTS9 immunoreactivity was detected in cells located at the boundary of hESC colonies. The expression of ADAMTS9 was then studied during differentiation of hESC. ADAMTS9 mRNA and protein expression increased time-dependently during the first 24 days? of embryoid body (EB) formation. The expression pattern was similar to that of mesoderm and endoderm markers. Upon more specific lineage differentiation induced by retinoic acid and bone morphogenesis protein 4, ADAMTS9 mRNA expression was significantly increased. The positive correlation of ADAMTS9 with ESC differentiation was also found in mouse system, in which ADAMTS9 was increased time-dependently during mouse EB formation and down-regulated during reprogramming from somatic cells into induced pluripotent cells. Previous studies have shown that down-regulation of ADAMTS9 in several tumor tissues was attributed to ADAMTS9 hypomethylation. However, the present study demonstrated that the expression of ADAMTS9 was reduced in hESC after treatments with inhibitors of DNA methylation (5-aza-2?deoxycytidine) and histone deacetylase (VPA). The cellular localization of ADAMTS9 during hESC differentiation was further studied by co-localization of ADAMTS9 with several lineage specific markers. It was found that ADAMTS9 co-localized with mesoderm and endoderm markers. The functional role of ADAMTS9 in hESCs was then studied by transient ADAMTS9 knockdown. ADAMTS9 siRNA significantly decreased the expression level of mesoderm marker, REN. Thus, the role of ADAMTS9 during mesoderm differentiation was followed. ADAMTS9 was found to be dramatically increased after mesoderm differentiation of hESCs. In mesodermal cells, ADAMTS9 was co-expressed with vascular endothelial markers, VEGF and CD31, but not with pericyte markers, alpha muscle actin. Lentiviral vector encoding ADMATS9 shRNA was used for long term knockdown of ADAMTS9. ADAMTS9 down-regulation had no effect on the proliferation of hESCs. In agreement with the siRNA study, ADAMTS9 shRNA also significantly reduced the expression of REN. Upon mesoderm differentiation, ADAMTS9 knockdown resulted in a decreasing trend of mesoderm marker, CD34. In conclusion, the present study demonstrated a positive association of ADAMTS9 expression with hESC differentiation. ADAMTS9 was dramatically induced during mesoderm differentiation and its knockdown led to down-regulation of mesoderm markers. Together with the fact that ADAMTS9 expression was associated with endothelial cell markers suggested its possible role during endothelial cells formation. The roles of ADAMTS9 during hESC differentiation and early embryo development warrant further investigation. / published_or_final_version / Obstetrics and Gynaecology / Master / Master of Philosophy

Oxidative stress and reactive oxygen species (ROS) such as hydrogen peroxide (H2O2), which is detoxified by catalase, are implicated in fetal death and birth defects, but embryonic levels of catalase are only about 5% of adult activity, and its protective role is unknown. Our approach involved the use of mice genetically modified to either: (1) express low levels of endogenous catalase (acatalasemic, aCat); or, (2) express human catalase resulting in elevated levels of embryonic catalase activity (hCat). Using these mouse models we investigated the protective importance of constitutive embryonic catalase against endogenous ROS and the ROS-initiating teratogen phenytoin in embryo culture and in vivo. We hypothesized that aCat mice would be more sensitive to endogenous embryonic and phenytoin-enhanced ROS production, while hCat embryos would be less sensitive. aCat and hCat embryos respectively exhibited reduced and enhanced catalase activity compared to wild-type (WT) controls, with conversely enhanced and reduced spontaneous and phenytoin-enhanced embryopathies and DNA oxidation. Among aCat embryos exposed to phenytoin, embryopathies increased with decreasing catalase activity, and were completely blocked by addition of exogenous catalase. The alterations in phenytoin embryopathies were not due to pharmacokinetic differences, as drug concentrations in maternal and fetal tissues were similar among all strains. However, phenytoin concentrations in fetal brain exceeded those in fetal liver or maternal tissues, which may explain the predominance of cognitive deficits over structural birth defects in children exposed in utero to phenytoin. Similarly in untreated aged mice (about 18 months), female aCat mice showed a substantial loss in motor coordination compared to WT controls in the rotarod test. Following in utero exposure to phenytoin, the effect of altered embryonic catalase activity on postnatal neurodevelopment was assessed by several pre- and post-weaning tests. Catalase deficiency (aCat), independent of drug treatment, reduced performance in surface righting, negative geotaxis tests and rotarod tests. Conversely, high catalase expression (hCat) enhanced performance in the surface righting, negative geotaxis, air righting and rotarod tests. Our results provide the first evidence that the quantitatively minor amounts of antioxidative enzymes like catalase in the embryo and fetus provide important protection against the molecular damage and adverse fetal effects caused by developmental and drug-enhanced oxidative stress. Accordingly, interindividual variation in embryonic/fetal activities of catalase, and possibly other antioxidative enzymes, likely constitute an important determinant of risk for adverse developmental outcomes.

Oxidative stress and reactive oxygen species (ROS) such as hydrogen peroxide (H2O2), which is detoxified by catalase, are implicated in fetal death and birth defects, but embryonic levels of catalase are only about 5% of adult activity, and its protective role is unknown. Our approach involved the use of mice genetically modified to either: (1) express low levels of endogenous catalase (acatalasemic, aCat); or, (2) express human catalase resulting in elevated levels of embryonic catalase activity (hCat). Using these mouse models we investigated the protective importance of constitutive embryonic catalase against endogenous ROS and the ROS-initiating teratogen phenytoin in embryo culture and in vivo. We hypothesized that aCat mice would be more sensitive to endogenous embryonic and phenytoin-enhanced ROS production, while hCat embryos would be less sensitive. aCat and hCat embryos respectively exhibited reduced and enhanced catalase activity compared to wild-type (WT) controls, with conversely enhanced and reduced spontaneous and phenytoin-enhanced embryopathies and DNA oxidation. Among aCat embryos exposed to phenytoin, embryopathies increased with decreasing catalase activity, and were completely blocked by addition of exogenous catalase. The alterations in phenytoin embryopathies were not due to pharmacokinetic differences, as drug concentrations in maternal and fetal tissues were similar among all strains. However, phenytoin concentrations in fetal brain exceeded those in fetal liver or maternal tissues, which may explain the predominance of cognitive deficits over structural birth defects in children exposed in utero to phenytoin. Similarly in untreated aged mice (about 18 months), female aCat mice showed a substantial loss in motor coordination compared to WT controls in the rotarod test. Following in utero exposure to phenytoin, the effect of altered embryonic catalase activity on postnatal neurodevelopment was assessed by several pre- and post-weaning tests. Catalase deficiency (aCat), independent of drug treatment, reduced performance in surface righting, negative geotaxis tests and rotarod tests. Conversely, high catalase expression (hCat) enhanced performance in the surface righting, negative geotaxis, air righting and rotarod tests. Our results provide the first evidence that the quantitatively minor amounts of antioxidative enzymes like catalase in the embryo and fetus provide important protection against the molecular damage and adverse fetal effects caused by developmental and drug-enhanced oxidative stress. Accordingly, interindividual variation in embryonic/fetal activities of catalase, and possibly other antioxidative enzymes, likely constitute an important determinant of risk for adverse developmental outcomes.

No description available.

Includes bibliographies. / xi, 127, [90] leaves, [31] leaves of plates : ill. (some col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Aims to identify homeobox genes which may have a developmental role during early embryogenesis by the characterization of homeobox gene expression in undifferentiated ES cells, and in a range of differentiated ES cell derivatives. / Thesis (Ph.D.)--University of Adelaide, Dept. of Biochemistry, 1996

Thesis (Ph. D.)--University of Adelaide, Dept. of Biochemistry, 1996. / Includes bibliographical references.

Dissertation (DPhil)--University of Stellenbosch, 2007. / Bibliography. Also available via the Internet.