Daniel J Pearce1, Dominique Bonnet. 1. Hematopoietic Stem Cell Laboratory, Cancer Research UK, London Research Institute, London, UK.
Abstract
OBJECTIVE: In murine hematopoietic tissue, direct repopulation experiments have demonstrated that the side population (SP) represents a remarkable enrichment of hematopoietic stem cells. Human SP has been phenotyped as negative for lineage antigens as well as CD34. However, in the 9 years since the original publication, no long-term hematopoietic reconstitution has been reported for the adult human SP/CD34(-) subset. Elevated levels of aldehyde dehydrogenase (ALDH) have been demonstrated in murine and human progenitor cells when compared to other hematopoietic cells. METHODS: Here, we report the phenotype of human cord blood SP cells. We established the technique of simultaneous phenotyping, Hoechst exclusion, and ALDH labeling on murine tissues. We then performed the simultaneous analysis of phenotype, SP, and ALDH activity on human cord blood and bone marrow cells. Finally, we analyzed the phenotype and functional potential of human cord blood ALDH(+) cells to determine whether Lin(-)/CD34(-) cells are identified via this technique. RESULTS: We demonstrate that human Lin(-)/CD34(-)/ALDH(+) cells are capable of long-term repopulation. Although the SP technique identifies cells that overlap with the ALDH(+) cell population, this is restricted to the CD34(+) cell subset. CONCLUSION: Hoechst exclusion ability does not seem to be the method of choice for the isolation of human hematopoietic stem cells.
OBJECTIVE: In murine hematopoietic tissue, direct repopulation experiments have demonstrated that the side population (SP) represents a remarkable enrichment of hematopoietic stem cells. HumanSP has been phenotyped as negative for lineage antigens as well as CD34. However, in the 9 years since the original publication, no long-term hematopoietic reconstitution has been reported for the adult humanSP/CD34(-) subset. Elevated levels of aldehyde dehydrogenase (ALDH) have been demonstrated in murine and human progenitor cells when compared to other hematopoietic cells. METHODS: Here, we report the phenotype of human cord blood SP cells. We established the technique of simultaneous phenotyping, Hoechst exclusion, and ALDH labeling on murine tissues. We then performed the simultaneous analysis of phenotype, SP, and ALDH activity on human cord blood and bone marrow cells. Finally, we analyzed the phenotype and functional potential of human cord blood ALDH(+) cells to determine whether Lin(-)/CD34(-) cells are identified via this technique. RESULTS: We demonstrate that human Lin(-)/CD34(-)/ALDH(+) cells are capable of long-term repopulation. Although the SP technique identifies cells that overlap with the ALDH(+) cell population, this is restricted to the CD34(+) cell subset. CONCLUSION:Hoechst exclusion ability does not seem to be the method of choice for the isolation of human hematopoietic stem cells.
Authors: Ines A Silva; Shoumei Bai; Karen McLean; Kun Yang; Kent Griffith; Dafydd Thomas; Christophe Ginestier; Carolyn Johnston; Angela Kueck; R Kevin Reynolds; Max S Wicha; Ronald J Buckanovich Journal: Cancer Res Date: 2011-04-15 Impact factor: 12.701
Authors: Jonathan M Gerber; B Douglas Smith; Brownhilda Ngwang; Hao Zhang; Milada S Vala; Laura Morsberger; Steven Galkin; Michael I Collector; Brandy Perkins; Mark J Levis; Constance A Griffin; Saul J Sharkis; Michael J Borowitz; Judith E Karp; Richard J Jones Journal: Blood Date: 2012-01-19 Impact factor: 22.113
Authors: Garrett G Muramoto; J Lauren Russell; Rachid Safi; Alice B Salter; Heather A Himburg; Pamela Daher; Sarah K Meadows; Phuong Doan; Robert W Storms; Nelson J Chao; Donald P McDonnell; John P Chute Journal: Stem Cells Date: 2010-03-31 Impact factor: 6.277