Hoxb5 marks the long-term hematopoietic stem cell and reveals a near homogenous perivascular niche and prospective isolation and differentiation of the mouse skeletal stem cell
Abstract/Contents
- Abstract
- The hematopoietic stem cell (HSC) is arguably the most extensively characterized tissue stem cell and has served as a model for understanding numerous aspects of stem cell biology. Since its identification by prospective isolation, complex multi-parameter flow cytometry has facilitated studies on many aspects of HSC biology, including, self-renewal, differentiation, aging, niche, and diversity. However, due in part to its rarity, complexity of isolation, and lack of HSC-specific lineage tracing, many findings still remain controversial. Here we demonstrate by unbiased multiple-step screening, identification of a single gene, Hoxb5 (homeobox B5 also known as Hox-2.1), whose expression is limited to the long-term HSC (LT-HSC) in mice; linking of Hoxb5 to fluorescent proteins allows identification, isolation, and localization of the LT-HSC. In a single-color tri-mCherry reporter mouse driven by endogenous Hoxb5 regulation, only 20% of total immunophenotypic HSCs are labelled by the fluorescent protein. Critically, only the Hoxb5-positive HSCs show long-term reconstitution capacity after transplantation—i.e., > 16 week hematopoietic reconstitution in primary transplant recipients, and reconstitution in secondary recipients. Bone marrow (BM) analysis of recipient mice also reveals that Hoxb5-positive HSCs can give rise to Hoxb5-negative multipotent and oligopotent hematopoietic progenitors, but not vice versa (Hoxb5-negative cells do not give rise to Hoxb5-positive hematopoietic cells), indicating Hoxb5-positive LT-HSC are at the top of the hematopoietic hierarchy. Only 7--35% of various previously defined immunophenotypic HSCs are LT-HSCs. Finally, by in situ imaging of mouse bone marrow, we show that > 94% of LT-HSCs (Hoxb5+) are directly attached to VE-cadherin+ cells, implicating the perivascular space as a near-homogenous location of LT-HSCs. Prospective isolation and transplantation has been critical to the identification of the the hematopoietic stem cell and its downstream progenitors. Here we utilize this same approach to help identify the stem and progenitors of the skeletal lineage. Transcriptional profiling of these candidate stem and progenitor fractions revealed differential expression of CD24a and CD200, two cell surface molecules that we demonstrate can be utilized to separate chondrocyte progenitors from bone progenitors. Using modulation of the BMP2 and VEGF signalling pathways allowed us to direct differentiation of these stem and progenitor fractions to the formation of bone or cartilage. Lastly, we compared the transcriptome of our stem and progenitor fractions to the lineage tracing experiments using Mx-1, Sp7 (Osterix), and Bglap.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2016 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Chen, James Yuhtyng |
|---|---|
| Associated with | Stanford University, Institute for Stem Cell biology and Regenerative Medicine. |
| Primary advisor | Weissman, Irving L |
| Thesis advisor | Weissman, Irving L |
| Thesis advisor | Kim, Peter, 1958- |
| Thesis advisor | Nakauchi, Hiromitsu, 1952- |
| Thesis advisor | Shizuru, Judith Anne |
| Advisor | Kim, Peter, 1958- |
| Advisor | Nakauchi, Hiromitsu, 1952- |
| Advisor | Shizuru, Judith Anne |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | James Yuhtyng Chen. |
|---|---|
| Note | Submitted to the Institute for Stem Cell Biology and Regenerative Medicine. |
| Thesis | Thesis (Ph.D.)--Stanford University, 2016. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2016 by James Yuhtyng Chen
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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