Fig. 4: Determination of functional Mk-biased HSC immune phenotype.
From: Expansion of human megakaryocyte-biased hematopoietic stem cells by biomimetic Microniche
a–d BD Rhapsody single-cell RNA-seq profiles of CD34+CD38−CD45RA−CD90+ cells sorted after culture in Microniche or control medium. a Four cell clusters were identified by k-means clustering analysis. Each dot represents one cell and colors represent distinct cell clusters (C1–C4), as indicated. Pie chart showing the percentages of each cell cluster in control and Microniche groups. b CD49f expression in each cell cluster. Ab refers to antibody. c Expression of the indicated genes in the four distinct clusters. Bubble color indicates Log relative average expression; bubble size indicates Log relative percentage expression. d Heatmap of scaled expression for top 10 highest differentially expressed genes (DEGs) in the C2 cluster compared to other clusters, including surface marker genes SELL (also known as CD62L) and PROM1 (also known as CD133). e Representative FACS profiles of CD62L−CD133+ distribution within CD34+CD38−CD45RA−CD90+CD49fneg/low/high subpopulations of UCB CD34+ cells before (fresh) and after 7 days of culture in control medium or control medium supplemented with SR1 (1 μM), UM171 (40 nM), PVA (0.1%) or Microniche. f Heatmap of Log percentage (left) and representative FACS profiles (right) of hCD41a+ cells in mCD45−hCD45−mCD41−mCD42d−mCD61− population of recipient BM cells. Positive xenograft rates in groups were listed. g Mk-HSC frequency based on the mCD45−hCD45−mCD41−mCD42d−mCD61−hCD41a+ population 16 weeks post-transplantation. CD62L−CD133+ group n = 54, non-CD62L−CD133+ group n = 50 biologically independent animals. Chi-square test. Solid lines indicate best-fit linear model and dashed lines confidence intervals. Box plots show the median (middle line) with the 25th and 75th percentiles (box). h Heatmap of Log percentage (left) and representative FACS profiles (right) of mCD45−hCD45+ cells in recipient BM cells. Positive xenograft rates in groups were listed. i HSC frequency in xenograft NOG mice 16 weeks post-transplantation. CD62L−CD133+ group n = 54, non-CD62L−CD133+ group n = 50 biologically independent animals. Chi-square test. Solid lines indicate best-fit linear model and dashed lines confidence intervals. Box plots show the median (middle line) with the 25th and 75th percentiles (box).
