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Vitamin A and its association with Congenital Diaphragmatic Hernia

  • Author / Creator
    Rocke, Ayanna W
  • Introduction: Congenital Diaphragmatic Hernia (CDH), is a birth defect that occurs in approximately 1 in every 3,000 births. It arises when the diaphragm fails to form properly during development leaving a hole in the muscle. In utero, the abdominal contents protrude through hole, taking up intrathoracic space, and, in so doing, impeding growth of the lungs. The consequent lung hypoplasia is the cause of significant perinatal morbidity and mortality. While the cause of CDH is poorly understood, the Retinoid Hypothesis has been suggested to explain the etiology and has been widely used. The hypothesis states that abnormal retinoic acid signaling, an active metabolite of dietary vitamin A, plays a role in the development of CDH. Two teratogens, Nitrofen (2,4-Dichlorophenyl 4-nitrophenyl ether) and Bisdiamine (N,N’-bis (dichloroacetyl)-1,8-octamethylenediamine) have been shown to induce CDH in offspring when given to pregnant dams. The two goals of this thesis were 1) to test the hypothesis that maternal Vitamin A status influences the development of teratogen induced CDH in a mouse model, and 2) to investigate whether CDH inducing teratogens alter the expression of retinoid metabolism-related genes in the developing embryo. Methods: To achieve the first goal, a teratogenic mouse model of CDH was established by titrating the dose of Nitrofen and Bisdiamine until an adequate number of affected fetuses (with CDH) was observed without an excessive number of intrauterine deaths. Then, maternal vitamin A status was manipulated by feeding mice diets with deficient (0 IU vitamin A/g), sufficient (4 IU vitamin A/g) or excess (25 IU vitamin A/g) vitamin A. Vitamin A status was confirmed in the maternal and fetal tissues by HPLC. Lastly, we induced CDH in the offspring by treating timed pregnant mice on varied vitamin A diets with a combination of Nitrofen and Bisdiamine. Offspring were collected via dissection and the effect of the teratogen on the incidence and severity of CDH were recorded. For the second goal, offspring of timed pregnant mice exposed to Nitrofen and Bisdiamine were collected 24 hours after administration of teratogen. Offspring expression of specific genes involved in retinoid metabolism and CDH were then measured using qPCR and compared to controls. Results: We established a teratogenic mouse model of CDH by administering 0.5 mg/kg of Nitrofen and 0.125 mg/kg of Bisdiamine to pregnant dams. Also, we showed that manipulating dietary vitamin A content of female mice, changes their vitamin A status, and the status of their offspring. Mice fed a vitamin A deficient diet and their offspring had less vitamin A in the livers and plasma and those fed a vitamin A excess diet and their offspring had more vitamin A detected in their livers and plasma. Finally, for our first research goal, we demonstrated that marginal maternal vitamin A status, and, by extension, marginal fetal vitamin A status, is more susceptible to teratogen-induced fetal CDH, with increased incidence of CDH when compared to offspring with sufficient vitamin A status. Excess vitamin A status decreased the offspring’s susceptibility to insult from teratogen exposure even further with an even lower incidence of CDH. With investigating whether CDH inducing teratogens alter the expression of retinoid metabolism-related genes in the developing embryo, we observed a 50% decrease in gene expression of Stra6 and Rarb. This showed that the teratogen affected the expression of these two genes which have been implicated in the formation of CDH previously in both human and animal studies. Conclusion: This research helped support the Retinoid Hypothesis and highlighted the need for future studies on the role of vitamin A on diaphragm development.

  • Subjects / Keywords
  • Graduation date
    Fall 2019
  • Type of Item
    Thesis
  • Degree
    Master of Science
  • DOI
    https://doi.org/10.7939/r3-s5a7-fq92
  • License
    Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of these terms. The author reserves all other publication and other rights in association with the copyright in the thesis and, except as herein before provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.