Human pluripotent stem cells (hPSCs) can be differentiated into any type of cell, however, storing them long-term without contamination, which compromises their clinical use, is a great challenge. One strategy is to differentiate hPSCs into multipotent progenitors, which cannot be differentiated into all cell types, but still can be used as source for a select group. Mesenchymal stem cells (MSCs) are one option, but their limited proliferative ability call for an intermediate between them and PSCs. Cranial neural crest cells (NCC) have the potential to be differentiated into cells responsible for the craniofacial skeleton, cornea, peripheral nervous system, and skin pigmentation, and are therefore an ideal choice as a multipotent progenitor. "We know that PSCs make tumours, and MSCs from adults do not. So, in theory, NCCs make a safer intermediate," explains associate professor Makoto Ikeya, a lead researcher on NCC reprogramming. 

An important component to minimize contaminations is the use of a chemically-defined medium and feeder-free cultures. Such culturing protocols for NCCs are few and can be improved both in terms of efficiency and simplicity. Ikeya and other CiRA members have recently reported a new protocol in PLOS ONE that does both. The method uses small molecules to inhibit GSK3β and TGFβ and successfully induces 70-80% of hPSCs from various cell lines into NCCs.

Importantly, freezing the NCCs did not compromise their growth or differentiation ability, which means they can be kept long term. Moreover, the NCCs were differentiated into an assortment of cell lineages, including neurons, cornea, and MSCs, which were further differentiated into osteogenic, chondrogenic and adipogenic lineages.

Interestingly, there was no need to inhibit BMP during the NCC induction. BMP is considered an important signal in NCC development, but past research has been controversial regarding whether it must be controlled. Although the findings here suggest BMP is not essential, Ikeya feels that the present results may argue for BMP temporal dynamics, where BMP is active in the first few days of NCC induction, but inhibited thereafter. "It is controversial". Regardless, Ikeya is optimistic about how this simple protocol will invite more opportunities for cell therapies.

Image: NCCs differentiated into peripheral neurons. Each image shows staining with a different antibody.

Article Details

Title: Derivation of Mesenchymal stromal cells form Pluripotent Stem Cells through a Neural Crest Lineage using Small Molecule Compounds with Defined Media

Authors: Makoto Fukuta, Yoshinori Nakai, Kosuke Kirino, Masato Nakagawa, Kazuya Sekiguchi, Sanae Nagata, Yoshihisa Matsumoto, Takuya Yamamoto, Katsutsugu Umeda, Toshio Heike, Naoki Okumura, Noriko Koizumi, Takahiko Sato, Tatsutoshi Nakahata, Megumu Saito, Takanobu Otsuka, Shigeru Kinoshita, Morio Ueno, Makoto Ikeya, Junya Toguchida

Journal: PLOS ONE 9(12):e112291. DOI:10.1371/journal.pone.0112291