**1. Introduction**

[49] Sparrow MP, Lamb JP. Ontogeny of airway smooth muscle: structure, innervation, myogenesis and function in the fetal lung. Respir Physiol Neurobiol. 2003;137(2-3):

[50] Suarez-Rodriguez R, Belkind-Gerson J. Cultured nestin-positive cells from postnatal mouse small bowel differentiate ex vivo into neurons, glia, and smooth muscle. Stem

[51] Tollet J, Everett AW, Sparrow MP. Development of neural tissue and airway smooth muscle in fetal mouse lung explants: a role for glial-derived neurotrophic factor in lung innervation. Am J Respir Cell Mol Biol. 2002;26(4):420-9. Epub 2002/03/29. [52] Bratincsak A, Lonyai A, Shahar T, Hansen A, Toth ZE, Mezey E. Using brain slice cultures of mouse brain to assess the effect of growth factors on differentiation of bone marrow derived stem cells. Ideggyogy Sz. 2007;60(3-4):124-9. Epub 2007/04/25.

[53] Nishida T, Tsuji S, Tsujii M, Ishii S, Yoshio T, Shinzaki S, et al. Cultured bone mar‐ row cell local implantation accelerates healing of ulcers in mice. J Gastroenterol.

[54] Ohnishi S, Yasuda T, Kitamura S, Nagaya N. Effect of hypoxia on gene expression of bone marrow-derived mesenchymal stem cells and mononuclear cells. Stem Cells.

[55] Jerareungrattan A, Sila-asna M, Bunyaratvej A. Increased smooth muscle actin ex‐ pression from bone marrow stromal cells under retinoic acid treatment: an attempt for autologous blood vessel tissue engineering. Asian Pac J Allergy Immunol.

[56] Tamama K, Sen CK, Wells A. Differentiation of bone marrow mesenchymal stem cells into the smooth muscle lineage by blocking ERK/MAPK signaling pathway.

361-72. Epub 2003/10/01.

428 Regenerative Medicine and Tissue Engineering

Cells. 2004;22(7):1373-85. Epub 2004/12/08.

2008;43(2):124-35. Epub 2008/03/01.

2007;25(5):1166-77. Epub 2007/02/10.

2005;23(2-3):107-13. Epub 2005/10/29.

Stem Cells Dev. 2008. Epub 2008/06/20.

The cornea is composed of a multilayered epithelium, Bowman's membrane, stroma, Descemet's membrane, and endothelium. The corneal endothelium (CE) is a single layer of hexagonal cells that separates the corneal stroma from the aqueous humor of the anterior chamber. Transparency of the cornea is maintained by regulation of stromal hydration through the barrier and pump functions of the CE, and corneal transplantation has long been used to treat corneal endothelial defects. In fact, more than half of the patients who undergo full-thickness corneal transplantation have impairment of visual acuity due to corneal endothelial problems alone and have a normal corneal epithelium (Cosar et al., 2002; Mannis et al., 1981; Rapuano et al., 1990). Corneal transplantation requires a fresh human cornea, but there is a worldwide shortage of donors (Barboza et al., 2007; Cao et al., 2006; Shimazaki et al., 2004; Tuppin et al., 2007).

Stem cells or progenitor cells are defined by a capacity for self-renewal and the ability to generate different types of cells (multipotentiality) that are involved in the formation of mature tissues. In contrast, precursor cells are unipotential cells with limited proliferative capacity. Regenerative stem cells or precursors can be detected by the sphere-forming assay in various adult tissues, including the central nervous system (Nunes et al., 2003), bone marrow (Krause et al., 2001), skin (Kawase et al., 2004; Toma et al., 2001), retina (Coles et al., 2004), corneal epithelium (Mimura et al., 2010a; Yokoo et al., 2008), corneal stroma (Amano et al., 2006; Mimura, 2008a, 2008b; Uchida et al., 2005; Yamagami et al., 2007), and corneal endothelium (Amano et al., 2006; Mimura, 2005a, 2005b, 2005c, 2007, 2010b; Yamagami, 2006, 2007; Yokoo et al., 2005).

© 2013 Mimura et al.; licensee InTech. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2013 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Despite the successful isolation and characterization of stem cells from various tissues, relatively few animal studies have been done to investigate the efficacy of stem cell transplan‐ tation. A three-dimensional carrier that maintains cell-to-cell interactions is indispensable for tissue engineering using stem cells, but the resulting structural complexity does not allow us to easily perform investigations of stem cell transplantation.

We have isolated precursors with the propensity to develop into corneal endothelial-like cells from the CE of human donor corneas (Yokoo et al., 2005). We have also demonstrat‐ ed that cultured human corneal endothelial cells (HCECs) and rabbit CE-derived precur‐ sors are an effective cell source for treating corneal endothelial defects in a rabbit model (Mimura 2005a, 2005b). Because the number of corneal endothelial cell (CEC) precursors that can be isolated from a native cornea is insufficient for corneal transplantation, establishment of a method for the mass production of precursor cells is required before CEC transplantation can be employed clinically.

In this chapter, we introduce our recent work in the fields of regenerative medicine and tissue engineering for the CE using bipotential precursor cells. We isolated precursors with the propensity to develop into CECs from human CE, and we investigated the distribution and proliferative capacity of precursor cells derived from the central and peripheral regions of the cornea by the sphere-forming assay. We also tested the effect of injecting human corneal endothelial spheres anterior chamber (instead of full-thickness corneal transplantation) in a rabbit model of bullous keratopathy, a condition associated with corneal endothelial defects.
