Fig 2: A Representative FACS Plot Following CultureA total of 400 HSCs (CD150+CD48-EPCR+LSK cells) were sorted and cultured for 6 days with the indicated cytokine concentration.(A) Shown are the total cell number, the number of CD150+CD48-LSK cells, and the frequency of CD150+CD48-LSK cells among total cells.(B) A representative FACS plot of cells cultured with a condition of SCF 1.5ng/mL and TPO 1.0ng/mL.

Fig 3: Features of Cultured HSCs(A) EdU incorporation assay to determine the cell cycle status either under quiescent culture condition (SCF 3 ng/mL and TPO 0.1 ng/mL) or under the high cytokine concentration (SCF 100 ng/mL and TPO 100 ng/mL). Shown are representative plots (n=4, mean ±SD).(B) Bar graphs of the frequency of EdU+ cells as determined in (A).(C) The frequency of donor-derived cells in peripheral blood transplanted with cultured HSCs following 14-day (left) or 27-day (right) culture under indicated cytokine concentrations (S: SCF, T: TPO). A total of 500 freshly isolated HSCs is transplanted as control (n=6 for each group, mean ±SD).(D) The total cell number of a surface marker phenotype of CD150+CD48- LSK cells after 1, 2, 3, and 4 week-culture in the indicated conditions (n=4 for each group, mean ±SD).

Fig 4: Long-term donor chimerism is established by M-HSCT, related to Figure 1(A) Schematic of the M-HSCT strategy, illustrating the proposed exchange between mobilized recipient CD45.2 (in gray) and donor CD45.1 cells (in purple).(B) Schematic of the interaction between mobilization agents and their therapeutic targets within the BM microenvironment.(C) Representative plots showing the gating strategy used to characterize LSK and SLAM HSC circulating in the peripheral blood, stained for lineage markers, SCA1, KIT, CD48, and CD150.(D) Representative plots showing the gating strategy used to characterize the donor cells, extracted from the BM, stained for lineage markers, SCA1, KIT, pre-, and post-purification of lineage negative cells. In all the analyses, p less than 0.05 were considered significant (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. “ns” means non-significance).

Fig 5: Systemic hypoxia alters BM hematopoiesis toward increased erythropoiesis.a, Representative dot plots gated on live CD45+Lin-(CD3/CD19/Ly6G)CD115+Ly6Chi cells and proportion of BrdU+ monocytes in naive or LPS-challenged mice housed in normoxia or hypoxia for 24 h and pulsed with BrdU for the last 12 h (naive, n = 5–6; LPS treated, n = 8 in). b, Absolute numbers of BM LSK cells, CD48+CD150- HPC-1, CD48+CD150+ HPC-2 (n = 6 per group) and Lin-cKit+Sca1-CD127-CD16/32-CD34+ CMPs (n = 5–6 group) in mice as in a. c, Proportion of BM pre-GMs, pre-Meg-E, pre-CFU-E and CFU-E CD41-CD32/16- cells in mice as in a (n = 6 per group). d,e, Representative UMAP analysis of BM cells gated on live CD45+Lineage-Sca1-C-Kit+CD41-CD32/16- cells (d) and summary data of proportions of pre-GM and CFU-E cells (e) measured in the BM of mice treated with LPS and housed in normoxia (N) or hypoxia (H) for 5 d (n = 6 per group). Data are shown as the mean ± s.e.m. Each datapoint represents an individual mouse. Data in a–e are pooled from two independent experiments. Statistical testing: for a–c is by one-way ANOVA with Tukey’s multiple comparison test and for e by unpaired, two-tailed Student’s t-test.