Additionally, to further confirm the role of the wnt signalling pathway in NTDs, we performed RT‒qPCR analysis of wnt target genes in both the brains and spines of E9.5 embryos in the NTD model and found that all six tested wnt target genes were upregulated, but that Kdm5a was downregulated in the brains of these embryos (Fig. 4C). Here, KDM5A is linked to neural tube defect.