**6. Conclusion**

**Figure 7.** Histologic findings, modified from Mimura et al. (2005b) with permission. In the cryo group (A), HCEC group (B), and sphere eyes-up group (C), stromal edema is prominent and no cells are present on Descemet's membrane at the central cornea. In contrast, a monolayer of cells can be detected on Descemet's membrane in the sphere eyesdown group (D). There is no mononuclear cell infiltration near Descemet's membrane, suggesting no rejection of the

**Figure 8.** Flat mount preparations with phase-contrast (A–D) and fluorescence (E–H) microscopy, modified from Mim‐ ura et al. (2005b) with permission. At the central cornea, there are no cells on Descemet's membrane in the cryo group (A, E), HCEC group (B, F), and sphere eyes-up group (C, G). In contrast, HCEC-like hexagonal cells are present in the

For regenerative medicine, amplification of stem cells is required to treat each tissue or organ. Although much attention has been paid to maintaining the undifferentiated nature

sphere eyes-down group (D). These cells are also DiI-positive (H). Scale bar=100 μm.

**5.4. Advantages of transplanting CE precursors**

xenogeneic cells (D). Scale bar=100 μm, hematoxylin & eosin stain.

444 Regenerative Medicine and Tissue Engineering

We demonstrated that the endothelium from the peripheral region of the human cornea contains a higher density of precursors with strong proliferative capacity compared to the central endothelium. These HCEC precursors are able to differentiate into both mesenchy‐ mal and neural cells. We have also established a method for mass production by isola‐ tion of precursors from cultured HCECs using the sphere-forming assay. Transplantation of spheres into the anterior chamber and short-term maintenance of the eyes-down position was shown to be a simple and effective treatment strategy in our rabbit model of bul‐ lous keratopathy. This method of managing corneal endothelial defects may have the potential to replace conventional full-thickness corneal grafting and compensate for the worldwide shortage of donor corneas.
