Figure 5.

Gradual loss of regenerative capacity of primitive hematopoietic cells involved in the treated leukemic model, caused by excessive proliferation related senescence. a Competitive bone marrow transplantation assays were used to examine the regenerative capacity of hematopoietic cells in vivo. CD45.1⁺ bone marrow cells from wild-type B6.SJL female mice (5 × 10⁵) and CD45.2⁺ bone marrow cells (5 × 10⁵) sorted from the 1-day treated leukemic mice on different days post therapy were co-transplanted into each B6.SJL mice (CD45.1⁺) which have undergone lethal irradiation (9.5 Gy). Donor contribution and lineage differentiation status were examined 1 month later at a frequency of once per month for four consecutive months. b Donor contribution on the fourth month post transplantation was examined and deeply analyzed combined with the frequencies of HSC (CD45.2⁺LK⁺S⁺ cells) and LT-HSC (CD45.2⁺LK⁺S⁺CD34⁻Flk2⁻ cells) in BM CD45.2⁺ hematopoietic cell fractions of the 1-day treated leukemic mice on the indicated days tested by flow cytometry (n = 3–9). c Lineage differentiation status of the 4 months post transplantation (n = 3–9). d, e Higher levels of senescence confirmed in primitive hematopoietic cells post therapy compared to normal control. Senescence status of CD45.2⁺LK⁺S⁺ cells in the BM of normal control and the 1-day treated leukemic mice were tested on the 3rd and 5th day post therapy by C₁₂-FDG staining. The representative flow cytometry plots are shown in d, while the statistic results (n = 7–12) are shown in e. f, g Expression patterns of genes regulating HSC function in primitive hematopoietic cells changed post therapy. CD45.2⁺LK⁺S⁺ cells of normal control and the 1-day treated leukemic mice on the 3rd and 5th day post therapy were sorted out and used for analysis of gene expression patterns, including p16, p21, p53, EGR1 and FOS. All data were presented as mean ± SEM. Statistical significance determined as: *p < 0.05; **p < 0.01; ***p < 0.001; ns not significant.