Spatially Assembled Bilayer Cell Sheets of Stem Cells and Endothelial Cells Using Thermosensitive Hydrogels for Therapeutic Angiogenesis

Abstract

Although the coculture of multiple cell types has been widely employed in regenerative medicine, in vivo transplantation of cocultured cells while maintaining the hierarchical structure remains challenging. Here, a spatially assembled bilayer cell sheet of human mesenchymal stem cells and human umbilical vein endothelial cells on a thermally expandable hydrogel containing fibronectin is prepared and its effect on in vitro proangiogenic functions and in vivo ischemic injury is investigated. The expansion of hydrogels in response to a temperature change from 37 to 4 °C allows rapid harvest and delivery of the bilayer cell sheet to two different targets (an in vitro model glass surface and in vivo tissue). The in vitro study confirms that the bilayer sheet significantly increases proangiogenic functions such as the release of nitric oxide and expression of vascular endothelial cell genes. In addition, transplantation of the cell sheet from the hydrogels into a hindlimb ischemia mice model demonstrates significant retardation of necrosis particularly in the group transplated with the bilayer sheet. Collectively, the bilayer cell sheet is readily transferrable from the thermally expandable hydrogel and represents an alternative approach for recovery from ischemic injury, potentially via improved cell-cell communication.

Keywords: cell sheet engineering; cell-cell interaction; endothelial cells; stem cells; thermosensitive hydrogels.