Single cell profiling of human hematopoietic cells

We demonstrated that the induction of BAZ2B expression in human CD34+ cells or Lineage-CD34+CD38+ cells, enhanced the stemness of clonogenicity of the cells in in vitro assays. In order to confirm the stemness mRNA expression signature of the progenitors we performed single-cell sequencing of the BAZ2B-treated CD34+ or Lineage-CD34+CD38+ progenitors and used the luciferase-treated progenitors as a control. To assess the stemness of the reprogrammed progenitor population at the molecular level, we proceeded to perform transcriptional profiling of single cells before and after treatment with the BAZ2B. To establish a positive control for the stemness signature, we sorted hematopoietic multipotent stem fractions of HSCs (Lin-CD34+CD38-CD45RA-CD90+), MPPs (Lin-CD34+CD38-CD45RA-CD90-), MLPs (Lin-CD34+CD38-CD45RA+) and the lineage-committed progenitor populations (Lin-CD34+CD38+) and performed single-cell RNA sequencing. Then, to investigate BAZ2B-mediated induction toward an HSC-like state we induced Luciferase or BAZ2B cDNA ectopic expression in CD34+ cells for 14 days and sorted Lineage-GFP+ cells for single cell expression profiling. To specifically test reprogramming of lineage committed hematopoietic progenitors we induced BAZ2B/Luciferase expression in Lineage-CD34+CD38+ cells for 14 days and sequenced the Lineage-GFP+ single cells.