Studies on BrdU labeling of hematopoietic cells: Stem cells and cell lines

✍ Scribed by Lizhen Pang; Prem Veer Reddy; Christina I. McAuliffe; Gerald Colvin; Peter J. Quesenberry

• Publisher: John Wiley and Sons
• Year: 2003
• Tongue: English
• Weight: 416 KB
• Volume: 197
• Category: Article
• ISSN: 0021-9541
• DOI: 10.1002/jcp.10357

✦ Synopsis

Abstract

Studies using chronic in vivo BrdU exposure, isolating primitive stem cells, and determining BrdU labeling, indicate that stem cells cycle. BrdU is also incorporated into DNA during damage/repair. DNA, which has incorporated BrdU due to cycle transit is heavier than normal, while the density of DNA with damage/repair incorporation is intermediate. DNA density of purified lineage—rhodamine low (rho^low^) Hoechst low (Ho^low^) stem cells or FDC‐P1 cell line cells—was assessed in vitro, after exposure to cytokines and BrdU (cycling model) or cytokines and BrdU with bleomycin to induce strand breaks and hydroxyurea to halt cycle progression (damage/repair model). We determined DNA density using cesium chloride (CsCl) gradients and either fluorometry or dot blot chemiluminesence. DNA from BrdU labeled cycling Lin‐rho^lo^Ho^lo^ or FDC‐P1 cells was heavier than normal DNA, while damage repair DNA had an intermediate density. We then assessed BrdU labeling of Lin‐rho^lo^Ho^lo^ cells in vivo. We found that 70.9% of lin‐rho^lo^Ho^lo^ cells labeled at 5 weeks. DNA density of these cells was low, in the damage/repair range, but similar results were obtained with stem cells, which had proliferated in vivo. Dilution of BrdU in in vitro culture of proliferating FDC‐P1 cells also resulted in damage/repair density. We conclude that in vitro BrdU labeling models can distinguish between proliferation and damage/repair, but that we cannot obtain high enough in vivo levels to address this issue. All together, while we cannot absolutely exclude damage/repair as contributing to stem cell BrdU labeling, the data indicate that primitive bone marrow stem cells are probably a cycling population. J. Cell. Physiol. 197: 251–260, 2003© 2003 Wiley‐Liss, Inc.