Functional characterization of individual human hematopoietic stem cells cultured at limiting dilution on supportive marrow stromal layers.

作者：Herbert J. Sutherland, Peter M. Lansdorp, D Henkelman, AC Eaves, Connie J. Eaves · 发表于：Proceedings of the National Academy of Sciences · 年份：1990 · DOI：10.1073/pnas.87.9.3584 · 被引用次数：682 · 研究领域：Hematopoietic Stem Cell Transplantation、Acute Myeloid Leukemia Research、Mesenchymal stem cell research

A major goal of current hematopoiesis research is to develop in vitro methods suitable for the measurement and characterization of stem cells with long-term in vivo repopulating potential. Previous studies from several centers have suggested the presence in normal human or murine marrow of a population of very primitive cells that are biologically, physically, and pharmacologically different from cells detectable by short-term colony assays and that can give rise to the latter in long-term cultures (LTCs) containing a competent stromal cell layer. In this report, we show that such cultures can be used to provide a quantitative assay for human "LTC-initiating cells" based on an assessment of the number of clonogenic cells present after 5-8 weeks. Production of derivative clonogenic cells is shown to be absolutely dependent on the presence of a stromal cell feeder. When this requirement is met, the clonogenic cell output (determined by assessment of 5-week-old cultures) is linearly related to the input cell number over a wide range of cell concentrations. Using limiting dilution analysis techniques, we have established the frequency of LTC-initiating cells in normal human marrow to be approximately 1 per 2 X 10(4) cells and in a highly purified CD34-positive subpopulation to be approximately 1 per 50-100 cells. The proliferative capacity exhibited by individual LTC-initiating cells cultured under apparently identical culture conditions was found to be highly variable. Values fo...