Skip to main content
. Author manuscript; available in PMC: 2020 Feb 7.
Published in final edited form as: Cell Stem Cell. 2018 Nov 29;24(2):227–239.e8. doi: 10.1016/j.stem.2018.11.007

Figure 6. Multilineage differentiation potential of human gut HSPCs in vitro and in vivo.

Figure 6.

(A) Representative colonies from short-term CFC assay of Lin− CD34-enriched cells from BM or ileum LPL of D#259 (46 y/o male). Scale bar: 100μm. Bar graph shows normalized colony number per 35mm dish. (B) Representative colonies from long-term LTC-IC assay of single cell sorted HSCs from D#305 (28 y/o female) and D#332 (38 y/o male) BM or ileum LPLs co-cultured with MS5-DL1ind100 stromal cells. CD3+ T cells (green), CD20+ B cells (red) and CD14+ myeloid cells (purple) were detectable within the colony on day 42 by fluorescence microscopy. HC human PBMCs were positive staining controls. Scale bar: 20 μm. Bar graph shows the percentage of colonies forming from single-cell sorted BM or ileum LPL HSC. (C) Sublethally irradiated representative NSG mouse (#110) of 3 mice analyzed with human thymus implant (HLA-A3+A2−A9−) received FACS-sorted CD45+/dimCD34+ cells (1 × 106) from D#293 ileum LPLs (53 y/o female, HLA- A3−A2+A9−) and CD34+ magnetic-activated cell sorting (MACS)-isolated cells (1 × 105) from D#291 BM (26 y/o female, HLA- A3− A2− A9+) at a ratio of 10:1. Representative FCM gating shows T/NK/B/Myeloid lineage reconstitution in mouse blood approximately 19 weeks post-adoptive transfer (second row) and in mouse BM at termination at week 24 (third row). Human HC PBMCs (HLA- A3−A2+A9−) serve as positive controls (first row). Human thymus implant (HLA-A3+A2−A9−) harvested from another NSG mouse (#558) at termination at week 24 (fourth row) contains gut-derived CD4+CD8+ T cells and CD34+ progenitors. Mouse #558 received CD45+/dimCD34+ FACS-sorted cells (4.5 × 105) from D#291 ileum LPLs and CD34+ MACS-isolated cells (2.5 × 105) from D#293 BM at a ratio of 1.8:1. (D) Percentage of ileum (red), BM (purple) or thymus (green)-derived cells among hCD45+mCD45− populations in NSG mouse (#110) blood at different times post-adoptive transfer. (E) Percentage of different lineages repopulating in NSG mouse (#110) blood among HLA- A3−A2+A9− compartment originating from D#293 ileum. See also Figure S6 and Table S3, S4.