EMT blockage is required for mouse naïve pluripotent stem cell derivation. Mus musculus

Pluripotency is the differentiation capacity of particular cells exhibited in the early embryo in vivo and embryonic stem (ES) cells have been shown to originate from the inner cell mass (ICM) of an E3.5 blastocyst. Although the potential for ES cells to differentiate into the three germ layers is equated to ICM cells, they differ in the ability to maintain the capacity for self-renewal. Despite several studies on the maintenance of ES cells in the ground state of pluripotency, the precise mechanism of conversion from the ICM to the ES cell remains unclear. Here , we have examined the cell characteristics and expression profile within the intermediate stages of ES cell derivation from the ICM. Gene clustering and ontology (GO) analyses showed a significant change in the expression of epigenetic modifiers and DNA methylation-related genes in the intermediate stages. We have proposed that an epithelial-to-mesenchymal transition (EMT) blockage is required during derivation of mouse ES cells from E3.5 blastocysts. This study suggests a novel mechanistic insight into ES cell derivation and provides a time-course transcriptome profiling resource for the dissection of gene regulatory networks that underlie the transition from ICM to ES cells. Overall design: For the first microarray analysis, Blastocyst-outgrowths were collected on days-3, -5, -7, and -9 from seeded E3.5 blastocysts as well as from isolated ICMs and ES cells, in three biological replicates. Between 20 to 30 outgrowths were picked and pooled for each biological replicate. For the second microarray analysis, we collected new samples in a time resolution experiment that included ICM-outgrowths on days-0.5, -1, -2, -3, -5 and ES cells that were in passages -2, -4, and-15. In addition, we collected ICM-outgrowths cultivated in N2B27 supplemented with SB431542 on days -1 (D1-SB) and -3 (D3-SB) as the negative control. Between 30 to 40 ICM-outgrowths were picked and pooled in two biological replicates.