Developmental changes and telomere loss in hematopoietic cells. (A) Ontogeny-related differences in the production of CD34⁺ cells in culture. (Reproduced from Lansdorp et al⁴²  with permission.) Candidate stem cells with a CD34⁺CD45RA^lowCD71^low phenotype were purified from fetal liver (wk12), umbilical cord blood, and adult bone marrow and 10⁴ sorted cells were cultured in serum-free culture medium supplemented with IL-6, IL-3, steel factor, and erythropoietin as described.³⁹  At the indicated time interval, the total number of nucleated cells (light blue squares) and CD34⁺ cells (dark blue squares) present in the cultures was calculated from cell counts and the percentage of viable CD34⁺ cells measured by flow cytometry. All CD34⁺ cells from bone marrow cultures and fractions of the CD34⁺ cells from cord blood and fetal liver cultures were sorted and used for continuation of the cultures. (B) Loss of telomeric DNA in human hematopoietic cells upon proliferation in vivo and in vitro (reproduced from Vaziri et al⁴³  with permission from the National Academy of Sciences of the United States). DNA samples from total nucleated cells from 2 different donors for each tissue before and after culture at increasing time points were subjected to terminal restriction fragment (TRF) size Southern blot analysis as described elsewhere.⁴³  Cultures were initiated with highly enriched stem cells as described in panel A. (C) Quantitative analysis of the TRF data shown in panel B. The first time point (blue square with circle) represents the TRF value in nucleated cells from each tissue before purification. The mean and standard error of 2 independent TRF measurements (different gels) for each DNA sample purified from total nucleated cells at each time point are shown. The total number of cells was used to calculate population doublings. The loss of telomeric DNA in these cultures varied between 19 and 54 base pairs per population doubling. For details see Vaziri et al.⁴³