IOX2 There has been controversy regarding whether subsets

There has been controversy regarding whether subsets of premigratory neural crest IOX2 are predetermined to form distinct cell types or multipotent and capable of forming numerous derivatives. Recently, this has been elegantly put to rest by single cell lineage analysis using Confetti transgenic mice. By labeling individual premigratory or migrating neural crest cells, the authors conclusively demonstrate that the majority of both are multipotent (Baggiolini et al., 2015), consistent with previous intracellular dye injections experiments in chick embryos (Bronner-Fraser and Fraser, 1988, 1989). Similarly, in vitro clonal analysis has demonstrated that many individual clones formed from early migrating neural crest cells can form numerous derivatives (Baroffio et al., 1988; Calloni et al., 2009; Sieber-Blum et al., 1993). This suggests that at least some cell fate decisions take place during neural crest migration. Our results support this, as crestospheres initiated from single cells retained expression of FOXD3, reflecting their self-renewal ability as multipotent stem-like cells. Moreover, our clonal analysis shows that individual crestosphere clones can differentiate into neural, melanocytic, and mesenchymal cell types, reflecting their multipotency. Interestingly, our results also show that the expression of neural crest markers is heterogeneous and dynamic in crestospheres, suggesting that perhaps only a subpopulation of crestosphere cells (∼10%) are true stem cells with the ability to self-renew.
Experimental Procedures
Acknowledgments
Introduction The liver is a central organ for metabolism, and the parenchymal cells, or hepatocytes, play key roles for homeostasis by expressing numerous metabolic and synthetic enzymes. As they express a number of cytochrome P450 oxidases (CYP450s) responsible for the oxidative biotransformation of many endogenous compounds as well as drugs, primary cultures of hepatocytes have been used for drug discovery and toxicology. However, primary hepatocytes exhibit low metabolic activity in vitro, and the supply of human hepatocytes is also limited and variable. To overcome these challenges, human embryonic stem cells (hESCs) and human-induced pluripotent stem cells (hiPSCs) have been considered as an alternative cell source for production of human hepatocytes. To date, there are many studies reporting hepatic differentiation of hiPSCs/hESCs (Ogawa et al., 2013; Si-Tayeb et al., 2010; Takayama et al., 2012). However, in most cases, differentiation of hepatocytes from hiPSCs is accomplished by a time-consuming culture protocol with multiple differentiation steps using expensive cytokines. Also, hepatocytes derived from hiPSCs possess a limited capacity for proliferation and functional maturation. Thus, it is beneficial to develop a simplified culture system for large-scale production of mature hepatocytes from hiPSCs. As liver progenitor cells (LPCs) such as hepatoblasts proliferate extensively in vitro, it would be useful if such cells could be derived from hiPSCs. The development of the mouse liver begins with early endoderm development. The cells of the ventral foregut endoderm are induced to the hepatoblast stage by fibroblast growth factor (FGF) and bone morphogenetic protein (BMP) signaling from the heart and septum transversum mesenchyme (STM). Following induction, hepatoblasts proliferate and migrate into the STM to form the liver bud with non-parenchymal cells, such as endothelial progenitor cells and hepatic mesenchymal cells (Zaret and Grompe, 2008). Importantly, hepatoblasts isolated from fetal liver can be cultured long-term while maintaining the potential to differentiate into both hepatocytes and cholangiocytes, two types of liver epithelial cell (Suzuki et al., 2000; Tanimizu et al., 2003). LPCs can also be isolated from normal as well as injured adult livers and maintained in culture for long term, although their role in vivo remains elusive (Miyajima et al., 2014).