Genetic and phenotypic response of neural tube defect mouse mutants to folic acid

Supplementation with folic acid (FA) is a public health policy to reduce the risk of neural tube defects (NTDs). However, compliance with recommended levels is low, and ≈50% of women and their babies are resistant to the beneficial effect of FA. If NTD response to FA could be predicted, compliance might improve and treatment could be targeted directly to responsive women. To test for maternal markers of fetal NTD risk and FA response, mouse models were used. Females of six single gene mouse mutants that are prone to NTD affected pregnancies were studied, among which three had an untested response to FA. We found that the NTD phenotype of these three mutants was resistant to maternal FA supplementation. Unexpectedly, these three mutants showed a substantial FA-induced loss of homozygous and heterozygous pre-implantation embryos. With two responsive and four resistant NTD mutants, whole genome expression and metabolite profiles were then analyzed to identify maternal markers for occurrence of fetal NTDs and FA response. Canonical pathways related to retinoid X receptor (RXR) function were associated with all NTD mutants regardless of FA response. By contrast, no specific pathways distinguished NTD-responsive and -resistant mutants, suggesting that multiple mechanisms control response to FA. Finally, gene expression analysis verified with in vitro assays showed that FA supplementation affected both Wnt/beta-catenin signaling and cell cycle regulation in the Lrp6 mutant. Overall, FA-induced loss of homozygous as well as otherwise healthy and viable heterozygous embryos highlight the need to determine the cause and stage of this early embryonic lethality. Moreover, the association of RXR related pathways with all six NTD mutants suggests a shared mechanism for NTD pathogenesis in which the role of RXR should be investigated. Our studies of the Lrp6 loss-of-function mutant, together with published work of the Lrp6 gain-of-function mutant, suggest that FA supplementation attenuates canonical Wnt signaling to normal levels in the latter mutant thereby restoring normal neural tube development, and to sub-optimal levels in the former mutant thereby compromising embryonic viability. The mechanisms by which FA affects Wnt signaling and the functional dependence of neural tube development on the level of Wnt signaling remain to be determined.