A model for p53 gene -teratogen interaction in early mouse embryos
In the United States, approximately 120,000 babies are born each year with a structural birth defect due to abnormal development. Majority of the causes of these defects is unknown although studies suggest that they may be due to a combination of genetic susceptibility and prenatal environmental exposure. To adequately characterize mechanisms of abnormal development, an understanding of normal development is required. Animal models are useful in the functional analyses of genes that regulate normal development and in experimental teratogenesis. During neurulation, mouse embryos are sensitive to 2-chloro-2′-deoxyadenosine (2CdA), a prototypic teratogen that mimics a naturally-occurring metabolic toxin, 2′-deoxyadenosine (dA). Altered sensitivity to teratogens and neural tube defects are associated with inactivation of p53, a gene that controls various cellular changes. Previous studies showed that exposure of gestational day 8 (GD8) mouse embryos to 2CdA and dA induced p53 protein production and programmed cell death (apoptosis). No study has determined the molecular mechanism by which these compounds affect normal development of embryos. The objective of this study was to determine the genetic mechanisms of susceptibility of embryos to 2CdA. ^ Molecular techniques such as immunostaining and polymerase chain reaction were used to investigate the role of p53 in the susceptibility of embryos to 2CdA. Expression of two ocular patterning genes, Pax6 and Siah1b, were analyzed to determine the molecular mechanisms of pathogenesis. Drug analysis of exposure levels in embryos was performed by high pressure liquid chromatography. The genetic basis of pro-drug activation by murine deoxyguanosine kinase (dGK) was determined by molecular cloning. ^ Exposure of GD8 embryos to 2CdA resulted in microphthalmia and anophthalmia within 72h. A dose-response curve showed a threshold between 1.5 and 5.0 mg/kg, embryolethality at 10.0 mg/kg and a linear relationship between exposure and phenotype. Peak levels of 2CdA in embryos were attained within 15 minutes and dGK was expressed on GD8. The earliest effect of exposure to 2CdA was significant reductions in bioenergy substrates ATP and NADH. These energy deficiencies may have resulted in activation of the p53 pathway, which led to reduced Pax6 expression and but had no effect on Siah1b at a critical time in eye development. All of these steps culminated in eye reduction defects, which were p53 gene dose-dependent. In conclusion, disruption of oxidative metabolism, which can be p53-dependent, is a possible mechanism of congenital malformations in mouse embryos. Knowledge of altered signaling in animal models can be extrapolated into predictive tests towards prevention of birth defect in humans. ^
Wubah, Judith Afedua, "A model for p53 gene -teratogen interaction in early mouse embryos" (1998). ETD Collection for Thomas Jefferson University. AAI9915379.