Low proliferation of progenitors in extramedullary tissues compared with BM. (A,D-E) Analysis of 10X scRNA-seq data from 117 200 cells, combining all donors but comparing different tissues. (A) Percentage of cells in S-G₂-M phase (assigned by cell cycle phase scoring as described previously⁷²) in matched BM and spleen (SPL) from the same donor for each indicated progenitor cluster. Two-sided exact binomial test. (B) Representative flow cytometry plots of BM (left), SPL (middle), and PB (right) CD19^–CD34⁺CD38^–CD45RA^– HSCs/MPPs (top row) or CD19^–CD34⁺CD38⁺ progenitor cells (bottom row) in G₀ (Ki-67^–DAPI^–), G₁ (Ki-67⁺DAPI^–), and S-G₂-M (Ki-67⁺DAPI⁺) cell cycle phases. (C) Frequency of phenotypic HSCs/MPPs (top row) or CD19^–CD34⁺CD38⁺ progenitor cells (bottom right) from each tissue in S-G₂-M phase (Ki-67⁺DAPI⁺) assessed by flow cytometry. Median ± 95% confidence interval is shown. A 2-tailed unpaired t test was used to compare BM and SPL (normal distribution), and 2-tailed Mann-Whitney U tests were used to compare BM/SPL with PB (not normally distributed); n = 3 nonmatched BM and SPL tissues, n = 9 PBs measured over 8 experiments. (D-E) Estimated cellular output from early to late progenitors of the MEMB (D) and the My (E) branch calculated from the number of active cells assuming all divisions are symmetric divisions toward differentiation (see Methods). Gray line indicates theoretical exponential expansion; vertical error bars indicate the range observed in the different tissues; and horizontal error bars indicate the standard deviation of the estimated number of divisions for each expansion stage. EoBasoMCP; eosinophil/basophil/mast cell progenitor; EryP, Ery progenitor; ILCp, innate lymphoid cell progenitor; LyP, lymphoid progenitor; MkP, megakaryocyte progenitors; MEP, megakaryocyte-erythroid progenitor; MPP, multipotent progenitor; MyP, myeloid progenitor.