Ciliated Cells Differentiated from Mouse Embryonic Stem Cells

¹ Department of Biological Science, Graduate School of Science, The University of Tokyo, Tokyo, Japan
² Department of Tissue Regeneration, Research Institute, International Medical Center of Japan, Tokyo, Japan
³ Department of Anatomy, Saitama Medical School, Saitama, Japan
⁴ Department of Life Sciences (Biology), Graduate School of Arts and Sciences, The University of Tokyo, Tokyo, Japan

^* To whom correspondence should be addressed. E-mail: hamazaki{at}ri.imcj.go.jp.

	Abstract

In the present study, we demonstrated that the mouse embryonic stem (ES) cells were differentiated into ciliated epithelial cells, with characteristics of normal ciliated cells. These cells expressed ciliary marker proteins, such as -tubulin IV and hepatocyte nuclear factor-3/forkhead homologue 4 (HFH-4), and processed microtubules arranged in the 9+2 structure, specific alignment observed in normal ciliary microtubles. The cilia of these cells were beating at a frequency of 17-20 Hz. Gene expression analysis revealed that the differentiated embryoid bodies (EBs) containing these ciliated cells expressed respiratory marker genes such as thyroid transcription factor-1 and surfactant protein-C. For the induction of ciliated cells, culture of EBs in serum-free medium during the initial 2 days of the attachment was indispensable. When EBs were treated with bone morphogenetic proteins, the expression of HFH-4 was decreased, and the ciliated cells were scarcely differentiated. Previous methods for inducing ciliated cells in vitro from embryonic or adult tissues involved an air-liquid interface. The system used in this study more closely mimics the normal development of ciliated cells, thus an added advantage of the system is as a tool for studying the differentiation mechanism of normal ciliated epithelial cells.