microRNA profiling as tool for developmental neurotoxicity testing (DNT)
microRNAs (miRNAs) are small non-coding RNA molecules functioning as post-transcriptional regulators of gene expression. miRNAs play a significant role in organism development, regulating developmental timing, cell differentiation, and specification. In the developing brain, miRNAs regulate neural stem cell differentiation, lineage specification, synaptogenesis, and brain morphogenesis. Temporal and spatial specificity of miRNA expression make them an attractive marker to study cellular responses to toxicant exposure. Neural differentiation of murine embryonic stem cells (mESCs) has been established as an alternative method to study developmental neurotoxicity (DNT) in vitro. This unit will describe a method for miRNA profiling (miRNomics) as a molecular end point to study developmental neurotoxicity. A protocol for neural differentiation of mESC will be described as a cellular model for DNT testing. The miRNomics protocol is versatile and can be used with other DNT cellular systems such as primary cultures, human embryonic stem cells (hESCs), or induced pluripotent stem cells (iPSCs).