Dunlevy, L.P.E.; (2006) The role of the folate and methylation cycles in neural tube closure. Doctoral thesis , University of London.

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Abstract

Neural tube defects (NTD) are congenital malformations caused by abnormalities in the developmental process of neurulation. Folate metabolism appears to be a determinant of risk of NTD since periconceptional supplementation with folic acid has been shown to reduce the frequency of NTD in humans while sub-optimal folate levels are a risk factor. The mechanisms underlying prevention of NTD by folic acid or susceptibility owing to reduced levels are not known. The aims of this thesis were to understand the role of the folate and, the closely linked, methylation cycle in the cause and prevention of NTD. The effect of methylation cycle intermediates, homocysteine and methionine, on cranial neural tube closure were investigated in cultured mouse embryos. Homocysteine exposure was embryotoxic but did not increase the incidence of NTD, which suggests that increased levels of homocysteine are not a direct cause of cranial NTD. Embryos cultured with high levels of methionine or methylation cycle inhibitors specifically developed cranial NTD in the absence of other developmental defects. These results suggest that the integrity of the methylation cycle is essential for cranial neural tube closure to occur. Mouse embryos that are homozygous for the Splotch211 mutation exhibit NTD that are preventable by folic acid. In Splotch mice, increased apoptosis has been suggested to be responsible for the production of NTD in homozygous embryos. However, in this study immunohistochemical measurement of apoptosis and proliferation in the neuroepithelium in the cranial region of neurulation-stage embryos suggest that the Splotch mutation does not result in increased apoptosis. Finally, in order to test whether there is an underlying defect in folate metabolism in human NTD fetuses, a series of human embryonic cell lines were analysed. The deoxyuridine monophosphate (dUMP) suppression test was modified for use with mammalian fibroblast cell lines and the efficiency and sensitivity of the modified test were analysed by the use of inhibitors of one-carbon metabolism. The test was then applied to human NTD and control cell lines and the results indicate that a subset of the NTD cases have a diminished response, suggestive of an abnormality of folate metabolism.

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