Isolation of Human Umbilical Cord Blood Aldehyde Dehydrogenase–Expressing Progenitor Cells that Modulate Vascular Regenerative Functions In Vitro and In Vivo

This unit describes the isolation and application of human umbilical cord blood progenitor cells to modulate vascular regenerative functions using in vitro co‐culture systems and in vivo transplantation models. Using aldehyde dehydrogenase as a marker of stem cell function, blood‐derived progenitors can be efficiently purified form human umbilical cord blood using flow cytometry. We describe in vitro approaches to measure cell‐mediated effects on the survival, proliferation, and tube‐forming function of endothelial cells using growth‐rate assays and Matrigel tube‐forming assays. Additionally, we provide a detailed protocol for inducing acute unilateral hindlimb ischemia in immune‐deficient mice to assess progenitor cell–modulated effects on vascular regeneration by tracking the recovery of blood flow using noninvasive laser Doppler perfusion imaging. Collectively, we present combined in vitro and in vivo transplantation strategies for the pre‐clinical assessment of human progenitor cell–based therapies to treat ischemic disease. Curr. Protoc. Stem Cell Biol. 25:2A.10.1‐2A.10.19. © 2013 by John Wiley & Sons, Inc.