Reactive oxygen species enhance differentiation of human embryonic stem cells into mesendodermal lineage

Recently, reactive oxygen species (ROS) have been studied as a regulator of differentiation into specific cell types in embryonic stem cells (ESCs). However, ROS role in human ESCs (hESCs) is unknown because mouse ESCs have been used mainly for most studies. Herein we suggest that ROS generation may play a critical role in differentiation of hESCs; ROS enhances differentiation of hESCs into bi-potent mesendodermal cell lineage via ROS-involved signaling pathways. In ROS-inducing conditions, expression of pluripotency markers (Oct4, Tra 1-60, Nanog, and Sox2) of hESCs was decreased, while expression of mesodermal and endodermal markers was increased. Moreover, these differentiation events of hESCs in ROS-inducing conditions were decreased by free radical scavenger treatment. hESC-derived embryoid bodies (EBs) also showed similar differentiation patterns by ROS induction. In ROS-related signaling pathway, some of the MAPKs family members in hESCs were also affected by ROS induction. p38 MAPK and AKT (protein kinases B, PKB) were inactivated significantly by buthionine sulfoximine (BSO) treatment. JNK and ERK phosphorylation levels were increased at early time of BSO treatment but not at late time point. Moreover, MAPKs family-specific inhibitors could prevent the mesendodermal differentiation of hESCs by ROS induction. Our results demonstrate that stemness and differentiation of hESCs can be regulated by environmental factors such as ROS.

Neural cell adhesion molecule (NCAM) promotes the differentiation of hippocampal precursor cells to a neuronal lineage, especially to a glutamatergic neural cell type

Rat hippocampal precursor cells isolated from hippocampi of embryonic day 16.5 (E16.5) rat embryos were found to proliferate in the presence of basic fibroblast growth factor. Addition of soluble neural cell adhesion molecule (NCAM) to these precursor cells reduced cell proliferation in a dose dependent manner and enhanced the induction of precursor cells' differentiation to the neuronal lineage. Given these findings that NCAM induces the differentiation of hippocampal precursor cells, we investigated possible effects of NCAM on the expression of basic helix-loop-helix (bHLH) transcription factors during the differentiation. Soluble NCAM upregulated the transcription of bHLH transcription factors, neurogenin1 and NeuroD, but decreased HES5. Western blot analysis showed that NCAM increased the expression levels of CaMKII, p-MAPK, GluR1 and NR1 but decreased p-STAT3. These results support a role for NCAM in the inhibition of proliferation and the induction of neural differentiation of hippocampal neural precursor cells, and act as developmental regulators of the bHLH families, ultimately leading to the generation of glutamatergic neural cell types in the differentiation of hippocampal precursor cells.