Thesis (Ph. D.)--University of Missouri--Columbia, 1998. / Typescript. Vita. Includes bibliographical references (leaves : 129-152). Also available on the Internet.

The aims of the research were to establish whether diphenylhydantoin and phenobarbitone are teratogenic in mice both in vivo and in an in vitro whole embryo culture system, to investigate possible mechanisms of teratogenicity and to examine whether the methods used in this study, may form a basis for developing systems of more extensive drug teratogenicity screening.

Leung, Chun Yin. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 155-169). / Abstracts in English and Chinese. / Title Page --- p.i / Acknowledgements --- p.ii / Table of Content --- p.iii / List of Figures --- p.viii / List of Graphs --- p.x / List of Tables x --- p.iv / Abbreviations --- p.xvii / Abstract --- p.xviii / Abstract (Chinese) --- p.xx / Chapter Chapter 1: --- General Introduction / Chapter 1.1 --- Introduction to retinoids --- p.2 / Chapter 1.2 --- Role of endogenous retinoic acid in embryonic development --- p.3 / Chapter 1.3 --- Regulation of retinoic acid in embryonic development --- p.5 / Chapter 1.3.1 --- Retinoic acid synthesis and degradation --- p.5 / Chapter 1.3.2 --- Retinoic acid signaling --- p.8 / Chapter 1.4 --- Effect of excess vitamin AJ RA on embryogenesis --- p.8 / Chapter 1.4.1 --- Examples of human animal studies --- p.9 / Chapter 1.4.2 --- Mechanisms of retinoid teratogenesis --- p.11 / Chapter 1.4.2.1 --- Apoptosis --- p.11 / Chapter 1.4.2.2 --- Altered proliferation --- p.12 / Chapter 1.4.2.3 --- Altered cell migration --- p.12 / Chapter 1.4.2.4 --- Altered differentiation --- p.13 / Chapter 1.4.3 --- Critical period of RA administration caused specific Malformations --- p.14 / Chapter 1.5 --- Effect of vitamin A/ RA deficiency on embryogenesis --- p.15 / Chapter 1.6 --- Excess and deficiency of RA cause similar types of malformations --- p.17 / Chapter 1.6.1 --- Retinoic acid-induced renal malformations mouse model --- p.18 / Chapter 1.7 --- Strategy of thesis --- p.19 / Chapter Chapter 2: --- General Materials and Methods / Chapter 2.1 --- Mouse maintenance and mating methods --- p.23 / Chapter 2.2 --- All-trans retinoic acid preparation and injection --- p.23 / Chapter 2.3 --- Whole mount in situ hybridization --- p.24 / Chapter 2.3.1 --- Riboprobe synthesis --- p.24 / Chapter 2.3.1.1 --- Bacterial culture --- p.24 / Chapter 2.3.1.2 --- DNA plasmids extraction --- p.24 / Chapter 2.3.1.3 --- Linearization of plasmid --- p.25 / Chapter 2.3.1.4 --- Purification of linearized plasmid --- p.26 / Chapter 2.3.1.5 --- In vitro transcription and labeling --- p.26 / Chapter 2.3.2 --- Sample collection --- p.27 / Chapter 2.3.3 --- Hybridization --- p.28 / Chapter 2.3.4 --- Post hybridization wash and antibody development --- p.29 / Chapter 2.3.4.1 --- Embryo powder preparation --- p.30 / Chapter 2.3.4.2 --- Pre-absorption of antibody --- p.30 / Chapter 2.3.5 --- Post-antibody and staining --- p.31 / Chapter 2.4 --- Real-time quantitative reverse transcription -polymerase chain reaction (RT-PCR) --- p.32 / Chapter 2.4.1 --- Sample collection --- p.32 / Chapter 2.4.2 --- RNA extraction --- p.32 / Chapter 2.4.3 --- Reverse transcription into cDNA --- p.33 / Chapter 2.4.4 --- Quantitative real-time PCR --- p.33 / Chapter 2.4.5 --- Preparation of cDNA standards --- p.34 / Chapter 2.5 --- High pressure liquid chromatography (HPLC) --- p.35 / Chapter 2.5.1 --- Chromatographic system --- p.35 / Chapter 2.5.2 --- Standards preparation --- p.35 / Chapter 2.5.3 --- Embryo sample collection and preparation --- p.36 / Chapter 2.5.4 --- HPLC conditions --- p.36 / Chapter 2.5.5 --- Sample recovery --- p.37 / Chapter 2.5.6 --- Bradford assay --- p.38 / Chapter 2.6 --- RA-responsive cell line --- p.38 / Chapter 2.6.1 --- Cell culture --- p.39 / Chapter 2.6.2 --- Seeding and loading sample to 96-well plate --- p.40 / Chapter 2.6.3 --- X-gal staining --- p.41 / Chapter Chapter 3: --- Time and Dose Responses to RA / Chapter 3.1 --- Introduction --- p.43 / Chapter 3.1.1 --- Time response to RA --- p.43 / Chapter 3.1.2 --- Dose response to RA --- p.45 / Chapter 3.1.3 --- Other factors affecting susceptibilities to RA --- p.46 / Chapter 3.2 --- Experimental design --- p.48 / Chapter 3.3 --- Materials and methods --- p.50 / Chapter 3.3.1 --- Time response to RA --- p.50 / Chapter 3.3.2 --- Dose response to RA --- p.50 / Chapter 3.3.3 --- Examination of fetuses --- p.51 / Chapter 3.3.4 --- Statistical analysis --- p.51 / Chapter 3.4 --- Results --- p.53 / Chapter 3.4.1 --- Time response --- p.53 / Chapter 3.4.1.1 --- Time response to RA-induced resorption --- p.53 / Chapter 3.4.1.2 --- Time response to RA-induced renal malformations --- p.54 / Chapter 3.4.1.3 --- Time response to RA-induced changes in growth parameters --- p.57 / Chapter 3.4.1.4 --- Time response to RA-induced non-renal malformations --- p.60 / Chapter 3.4.2 --- Dose response --- p.64 / Chapter 3.4.2.1 --- Dose response to RA-induced resorption --- p.64 / Chapter 3.4.2.2 --- Dose response to RA-induced renal malformations --- p.65 / Chapter 3.4.2.3 --- Dose response to RA-induced changes in growth parameters --- p.68 / Chapter 3.4.2.4 --- Dose response to RA-induced non-renal malformations --- p.71 / Chapter 3.5 --- Discussion --- p.74 / Chapter Chapter 4: --- Effect of Teratogenic Dose of RA on RA Synthesis and Endogenous RA Levels in the Embryo / Chapter 4.1 --- Introduction --- p.79 / Chapter 4.1.1 --- RA synthesis in embryo --- p.79 / Chapter 4.1.2 --- Detection of endogenous RA in embryo --- p.81 / Chapter 4.2 --- Experimental design --- p.83 / Chapter 4.3 --- Materials and methods --- p.84 / Chapter 4.3.1 --- Localization of mRNA transcripts in whole embryo by in situ hybridization --- p.84 / Chapter 4.3.2 --- Vibratome sectioning --- p.85 / Chapter 4.3.2.1 --- Preparation of Gloop --- p.85 / Chapter 4.3.2.2 --- Sample preparation and sectioning --- p.85 / Chapter 4.3.3 --- Quantification of mRNA expression levels in whole embryo and in metanephros by real-time RT-PCR --- p.86 / Chapter 4.3.4 --- Detection of RA levels in whole embryo by HPLC --- p.87 / Chapter 4.3.5 --- Detection of RA levels in metanephros by RA-responsive cell line --- p.87 / Chapter 4.3.6 --- Statistical analysis --- p.88 / Chapter 4.4 --- Results --- p.89 / Chapter 4.4.1 --- Comparison of mRNA expression levels of different iso forms of RA synthesizing enzymes Raldh and RA catabolizing enzymes Cyp26 between embryos of RA-treated and vehicle-treated control mice at various time points after treatment --- p.89 / Chapter 4.4.2 --- Comparison of mRNA expression levels of different iso forms of RA synthesizing enzymes Raldh and RA catabolizing enzymes Cyp26 between metanephroi of embryos of RA-treated and vehicle-treated control mice at various time points after treatment --- p.93 / Chapter 4.4.3 --- Comparison of the in situ hybridization pattern of different iso forms of Raldh between embryos of RA-treated and vehicle-treated control mice at different time points after treatment --- p.95 / Chapter 4.4.3.1 --- In situ hybridization pattern of Raldh 1 --- p.96 / Chapter 4.4.3.2 --- In situ hybridization pattern of Raldh2 --- p.97 / Chapter 4.4.3.3 --- In situ hybridization pattern of Raldh3 --- p.100 / Chapter 4.4.4 --- Comparison of the in situ hybridization pattern of Cyp26al and Cyp26bl between embryos of RA-treated and vehicletreated control mice at different time points after treatment --- p.101 / Chapter 4.4.4.1 --- In situ hybridization pattern of Cyp26al --- p.101 / Chapter 4.4.4.2 --- In situ hybridization pattern of Cyp26bl --- p.102 / Chapter 4.4.5 --- Comparison of RA levels between embryos of RA-treated and vehicle-treated control mice at different time points after treatment --- p.103 / Chapter 4.4.6 --- Comparison of RA levels between metanephroi of embryos of RA-treated and vehicle-treated control mice at different time points after treatment --- p.105 / Chapter 4.5 --- Discussion --- p.106 / Chapter Chapter 5: --- Effect of Supplementation with Low Doses of RA on RA Teratogenesis / Chapter 5.1 --- Introduction --- p.111 / Chapter 5.1.1 --- RA supplementation --- p.111 / Chapter 5.1.2 --- Wilms' tumor suppressor gene Wtl --- p.112 / Chapter 5.1.3 --- Apoptosis --- p.113 / Chapter 5.2 --- Experimental design --- p.115 / Chapter 5.3 --- Materials and methods --- p.117 / Chapter 5.3.1 --- Oral gavage of low dose of RA --- p.117 / Chapter 5.3.2 --- Determination of Wtl expression level by real-time quantitative RT-PCR --- p.117 / Chapter 5.3.3 --- Preparation of paraffin sections and TUNEL staining --- p.118 / Chapter 5.3.3.1 --- Sample collection --- p.118 / Chapter 5.3.3.2 --- "Dehydration, embedding and sectioning" --- p.118 / Chapter 5.3.3.3 --- TUNEL staining --- p.119 / Chapter 5.3.4 --- Statistical analysis --- p.121 / Chapter 5.4 --- Results --- p.122 / Chapter 5.4.1 --- Time response to RA supplementation in rescuing kidney development --- p.122 / Chapter 5.4.2 --- Dose response to RA supplementation in rescuing kidney development --- p.127 / Chapter 5.4.3 --- RA supplementation restored various growth parameters --- p.132 / Chapter 5.4.4 --- RA supplementation rescued non-renal malformations --- p.134 / Chapter 5.4.5 --- Wtl expression in the metanephros after RA supplementation --- p.142 / Chapter 5.4.6 --- Apoptotic cell death in the metanephros after RA supplementation --- p.143 / Chapter 5.5 --- Discussion --- p.145 / Chapter Chapter 6: --- Conclusion and Future Perspectives --- p.150 / References --- p.155 / Figures / Graphs

Diss. (sammanfattning) Stockholm : Karol. inst., 2003. / Härtill 7 uppsatser.

Diss. (sammanfattning) Uppsala : Uppsala universitet, 2009.

In two separate experiments, weight adjusted doses of nocodazole and acrylamide were injected intraperitoneally at various time intervals into twelve week old female mice. Within the nocodazole experiment, the doses were injected at varying time intervals before and after mating. On day seventeen of gestation, the female mice were sacrificed and their uterine contents examined. Nocodazole induced a significant increase in reproductive pathology per total implants when administered one hour after mating to the (SECxC57BL)F, stock: 5.00% total deads, 70.23% moles, and 3.41% abnormal fetuses. Acrylamide treatment produced a significant reduction in live births when administered six hours after mating: 50.86% moles and 46.46% living fetuses per total implants.

Retinol (vitamin A) and its active metabolite, all-trans retinoic acid, signal through nuclear retinoic acid and retinoid X receptor (RAR/RXR) heterodimers. These complexes regulate the expression of genes involved in developmental processes such as limb development. In excess, retinoids are potent teratogens and cause marked reductive effects on the developing limb. The goal of this thesis was to elucidate the molecular mechanisms underlying retinoid-induced limb dysmorphogenesis. Specifically, using an in vitro limb culture system, I examined the involvement of one RAR isoform, RARgamma, in mediating retinoid insult. / My first objective was to examine how limbs deficient in RARgamma responded to exogenous retinoid exposure. I showed that RARgamma-null limbs (on an RARalpha1-null background) exhibited less severe limb defects following retinoid insult when compared to their wild-type counterparts. Additionally, the absence of RARgamma abolished the retinoid-induced misregulation of genes important for chondrogenesis (Sox9 and Col2a1 ) and limb outgrowth (Meis-1 and -2). / The next objective set out to determine how pharmacological activation of RARgamma affected limb development. The RARgamma-selective agonist (BMS-189961) caused limb dysmorphology (namely, effects on cartilage) that was comparable to pan-RAR activation with all-trans retinoic acid. A chondrogenesis-focused gene array analysis identified Mgp and Gdf10 as two RARgamma-responsive genes that may mediate retinoid-induced limb insult. / Subsequently, I assessed the functional involvement of Mgp in mediating retinoid teratogenicity. Limbs were treated with all- trans retinoic acid and warfarin (an inhibitor of MGP); warfarin co-treatment rescued limbs from retinoid-induced insult. / My final objective was to determine the importance of Gdf10 in mediating limb development. Recombinant human Gdf10-soaked beads were implanted into distal limb structures; ectopic overexpression of Gdf10 in the web (but not the digital ray) resulted in marked proximal limb malformations. / Collectively, these studies have illustrated the importance of RARgamma in retinoid teratology and have identified several potential mechanisms by which retinoids cause limb defects.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2010.

No description available.

L’asthme est l’une des maladies chroniques les plus fréquentes durant la grossesse, affectant environ 4% à 12% des femmes enceintes et ayant une prévalence qui a augmenté au cours des dernières décennies. Plusieurs études ont identifié l'asthme comme un facteur de risque pour plusieurs enjeux de santé défavorables chez le fœtus et la mère. Les lignes directrices de traitement recommandent l’utilisation de médicaments antiasthmatiques pendant la grossesse afin de contrôler l’asthme et d’éviter les problèmes de santé maternels et fœtaux. L’évaluation de la littérature sur l'utilisation maternelle de médicaments antiasthmatiques et le risque de malformations congénitales majeures a relevé plusieurs études sur l’innocuité des bêta2-agonistes inhalés à courte durée d’action (BACA) et des corticostéroïdes inhalés (CSI) pendant la grossesse, mais peu de données sur les bêta2-agonistes à longue durée d’action (BALA) ainsi que sur les thérapies combinées (BALA-CSI). Un programme de recherche en trois volets a été développé pour combler ces lacunes. Dans le premier volet, nous avons entrepris une revue systématique de la littérature sur l'impact de l'utilisation de BACA et de BALA pendant la grossesse sur le risque de différents problèmes périnataux. Vingt et une études originales ont été identifiées. Quatre études ont rapporté une augmentation significative du risque de malformations congénitales avec BACA, une étude a rapporté une augmentation significative du risque de malformations congénitales avec BALA et quatre études ont rapporté un risque significatif accru de malformations congénitales avec bêta2-agonistes (BACA et/ou BALA). Toutefois, aucun risque majeur n’a été trouvé pour les autres complications périnatales. Fait important, la plupart des études récupérées ont subi plusieurs limitations méthodologiques, y compris l'utilisation des femmes non-asthmatiques comme groupe de référence et la faible puissance statistique. De plus, les résultats qui en découlent doivent être interprétés avec prudence. Dans le deuxième volet, nous avons utilisé la base de données Québec Asthma and Pregnancy Database qui comprend toutes les grossesses de femmes asthmatiques et un échantillon aléatoire de femmes non-asthmatiques ayant accouchées entre 1990 et 2010 pour effectuer deux études. La première était une étude comparant la prévalence des malformations congénitales majeures entre les femmes enceintes asthmatiques traitées avec une combinaison de BALA-CSI et celles traitées avec une dose plus élevée de CSI en monothérapie. Dans une sous-cohorte, il y’avait 643 femmes qui utilisaient un BALA plus CSI à dose faible et 305 qui ont utilisé une dose moyenne de CSI ; l'autre sous-cohorte comprenait 198 utilisatrices de BALA plus CSI à dose moyenne et 156 utilisatrices de CSI à dose élevée. La prévalence de malformations majeures a été 6,9% et 7,2%, respectivement. Le risque de malformations congénitales majeures était similaire entre ces deux groupes de femmes avec un odds ratio ajusté (OR) de 1,1 (IC 95%: 0,6-1,9) pour les femmes souffrant d’asthme modéré et un OR ajusté de 1,2 (IC 95%: 0,5-2,7) pour les femmes souffrant d’asthme sévère. La seconde était une étude méthodologique visant à étudier l’impact de six différentes définitions opérationnelles de malformations congénitales qui varient selon la source des données et la méthode de classification sur l’estimation de la prévalence des malformations et de l'association entre l'asthme maternel et les malformations majeures. Sur 467,946 grossesses, 12,3% étaient de femmes enceintes souffrant d’asthme actif. Nous avons démontré que la source des données et la méthode de classification ont eu un impact considérable sur la prévalence des malformations congénitales majeures (augmentation entre 10,0% et 50,4%), alors qu’elles ont eu peu d’influence sur l’association entre l’asthme maternel et les malformations congénitales. Dans le troisième volet du programme de recherche, nous avons développé une procédure systématique pour la classification des médicaments utilisés au cours du premier trimestre de grossesse en agents tératogènes et potentiellement tératogènes dans un contexte de recherche. Nous avons développé une procédure systématique qui s’actualise facilement, avec des composantes objectives dans la plupart de ses processus. Nous avons établi une liste comprenant 91 médicaments tératogènes, et une autre liste comprenant 81 médicaments potentiellement tératogènes. Les résultats présentés dans cette thèse ont fourni des données importantes sur l’innocuité des traitements de l'asthme pendant la grossesse, aidant les cliniciens et les femmes enceintes à choisir un traitement pharmacologique sécuritaire pour maintenir l’asthme sous contrôle. De plus, les données présentées dans cette thèse sur la minimisation du biais d'indication, les définitions opérationnelles de malformations congénitales et l’identification des médicaments tératogènes pourront aisément être utilisées par les chercheurs en pharmacoépidémiologie, en tératologie et en épidémiologie périnatale. / Asthma is one of the most prevalent chronic diseases during pregnancy, affecting about 4% to 12% of pregnant women and shows an increasing prevalence over time. In the past decades, several studies have identified asthma as a risk factor for several poor fetal and maternal outcomes. A consensus exists on favoring the use of asthma medications during pregnancy to maintain asthma under control to prevent adverse maternal and fetal outcomes. An assessment of the published literature on maternal asthma medications and the risk of major congenital malformations revealed more data on the safety of short-acting beta2-agonists (SABA) and inhaled corticosteroids (ICS) during pregnancy compared to long-acting beta2-agonists (LABA), as well as a paucity of data on the fetal safety of combination therapies (e.g. LABA-ICS). A three-part research program was developed to fill this knowledge gap and answer other intriguing questions we faced, adding necessary evidence in this field. In the first part, we summarized the published evidence on the impact of maternal use of SABA and LABA during pregnancy and different perinatal outcomes in a comprehensive systematic review. Twenty-one original studies were identified. Four studies reported a significant increased risk of congenital malformations with SABA, one study reported a significant increased risk of congenital malformations with LABA and four studies reported a significant increased risk of congenital malformations with beta2-agonists (SABA and/or LABA). However, no major increased risk was found for the other perinatal outcomes. Importantly, most of the retrieved studies suffered several methodologic limitations, including using non-asthmatic women as the reference group and low statistical power. Moreover, the non-significant results reported should be interpreted with caution. In the second part, we used the Quebec Asthma and Pregnancy Database – which includes all pregnancies in asthmatic women and a random sample in nonasthmatic women between 1990 and 2010 – to conduct two studies. The first was a comparative safety study examining the prevalence of major congenital malformations in pregnant asthmatic women treated with a combination of LABA-ICS compared to those treated with a higher dose of ICS monotherapy. In one subcohort there were 643 women who used a LABA plus low-dose ICS and 305 women who used a medium-dose ICS; the other subcohort included 198 users of a LABA plus a medium dose ICS and 156 users of a high-dose ICS. The prevalence of major malformations was 6.9% and 7.2%, respectively. The risk of major malformations did not differ when a combination therapy was used among both moderate and severe asthmatic women (aOR: 1.1; 95% CI: 0.6–1.9 and aOR: 1.2; 95% CI: 0.5–2.7 respectively). The second was a methodological study aiming to compare the prevalence of major malformations using six different case ascertainment definitions that vary by the source of data and the classification method, as well as to evaluate the impact of these definitions on the association between maternal asthma and major malformations. From the 467,946 pregnancies, 12.3% were with active asthma. We demonstrated that the source of data and the classification method had a considerable impact on the prevalence of major malformations (increases between 10.0% and 50.4%), but only a small influence on the measure of association. In the third part of the research program, we aimed at constructing a systematic procedure for the classification of proven and potential teratogenic medications during the first trimester of pregnancy to be used for research. We structured a procedure that is both systematic and updatable, with objective components in most of its processes. We identified a substantial list of teratogenic medications, including 91 medications, and an extensive list of potentially teratogenic medications, including 81 medications. The results presented in the current thesis provided essential evidence on the safety of asthma treatments during pregnancy, helping clinicians and mothers to choose the optimal therapeutic regimen to keep asthma under control. The added knowledge on indication bias minimization, congenital malformations ascertainment and teratogenic medications are directly transferable to researchers in pharmacoepidemiology, teratology and other related research fields.