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- Skin graft with dermis and appendages generated in vivo by cell competition, Nature Communications 2024
- Genome engineering with Cas9 and AAV repair templates generates frequent concatemeric insertions of viral vectors, Nature Biotechnology 2024
- Chemically defined cytokine-free expansion of human haematopoietic stem cells, Nature 2023
- Functional primordial germ cell-like cells from pluripotent stem cells in rats, Science 2022
- Immunological barriers to haematopoietic stem cell gene therapy, Nature Reviews Immunology 2022
- Treatment of a genetic brain disease by CNS-wide microglia replacement, Science Translational Medicine 2022
- In vitro and in vivo functions of T cells produced in complemented thymi of chimeric mice generated by blastocyst complementation, Scientific Reports 2022
- Investigation of Cas9 antibodies in the human eye, Nature Communications 2022
- Generation of Tfap2c-T2A-tdTomato knock-in reporter rats via adeno-associated virus-mediated efficient gene targeting, Molecular Reproduction Development 2022
- Dual-antigen targeted iPSC-derived chimeric antigen receptor-T cell therapy for refractory lymphoma, Molecular Therapy 2022
- Advances in Allogeneic Cancer Cell Therapy and Future Perspectives on "Off-the-Shelf" T Cell Therapy Using iPSC Technology and Gene Editing, Cells. 2022
- Generation of heterozygous PKD1 mutant pigs exhibiting early-onset renal cyst formation, Laboratory Investigation 2022
- Pluripotent stem cells related to embryonic disc exhibit common self-renewal requirements in diverse livestock species, Development 2021
- Tracing the emergence of primordial germ cells from bilaminar disc rabbit embryos and pluripotent stem cells, Cell Reports 2021
- Polyvinyl alcohol hydrolysis rate and molecular weight influence human and murine HSC activity ex vivo, Stem Cell Res. 2021
- iPSC-Derived Neoantigen-Specific CTL Therapy for Ewing Sarcoma, Cancer Immunology Research 2021
- High glucose macrophage exosomes enhance atherosclerosis by driving cellular proliferation & hematopoiesis, iScience 2021
- ISSCR guidelines for the transfer of human pluripotent stem cells and their direct derivatives into animal hosts, Stem Cell Reports 2021
- ISSCR Guidelines for Stem Cell Research and Clinical Translation: The 2021 update, Stem Cell Reports 2021
- Terminal Sequence-Specific Interparticle Attraction between DNA Duplex-Carrying Polystyrene Microparticles in Aqueous Salt Solution Assessed by Optical Tweezers, ACS Publications 2021
- Feasibility of large experimental animal models in testing novel therapeutic strategies for diabetes, World Journal of Diabetes 2021
- DNA Terminal-Specific Dispersion Behavior of Polystyrene Latex Microparticles Densely Covered with Oligo-DNA Strands Under High-Salt Conditions, Analytical Sciences 2021
- Blastocyst complementation using Prdm14-deficient rats enables efficient germline transmission and generation of functional mouse spermatids in rats, Nature Communications 2021
- Cas9-AAV6 gene correction of beta-globin in autologous HSCs improves sickle cell disease erythropoiesis in mice, Nature Communications 2021
- Generation of Functional Organs Using a Cell-Competitive Niche in Intra- and Inter-species Rodent Chimeras, Cell Stem Cell 2021
- Haematopoietic stem cell self-renewal in vivo and ex vivo., Nat Rev Genet. 2020
- Long-term ex vivo haematopoietic-stem-cell expansion allows nonconditioned transplantation, Nature 2019
- Generation of pluripotent stem cell-derived mouse kidneys in Sall1-targeted anephric rats, Nature Communications 2019
- Changing concepts in hematopoietic stem cells, Science 2018
- Intra-embryo Gene Cassette Knockin by CRISPR/Cas9-Mediated Genome Editing with Adeno-Associated Viral Vector, iScience 2018
- Large-Scale Clonal Analysis Resolves Aging of the Mouse Hematopoietic Stem Cell Compartment, Cell Stem Cell 2018
- Interspecies organogenesis generates autologous functional islets, Nature 2017
- Depleting dietary valine permits nonmyeloablative mouse hematopoetic stem cell transplantation, Science 2016
- Inhibition of Apoptosis Overcomes Stage-Related Compatibility Barriers to Chimera Formation i Mouse Embryos, Cell Stem Cell 2016
- Stem cells and interspecies chimaeras, Nature 2016
- Hoxb5 marks long-term haematopoietic stem cells and reveals a homogenous perivascular niche, Nature 2016
- A Safeguard System for Induced Pluripotent Stem Cell-Derived Rejuvenated T Cell Therapy, Stem Cell Reports 2015
- Roles of histone H3K27 trimethylase Ezh2 in retinal proliferation and differentiation, Developmental Neurobiology 2015
- Successful Reprogramming of Epiblast Stem Cells by Blocking Nuclear Localization of β-Catenin, Stem Cell Reports 2015
- Targeted Organ Generation Using Mixl1-Inducible Mouse Pluripotent Stem Cells in Blastocyst Complementation, Stem Cells and Development 2015
- Clonal Analysis Unveils Self-Renewing Lineage-Restricted Progenitors Generated Directly from Hematopoietic Stem Cells, Cell 2013
- Generation of rat pancreas in mouse by interspecific blastocyst injection of pluripotent stem cells, Cell 2010
A new model of self-renewal and lineage potential in hematopoiesis
In the conventional model, LT-HSCs form ST-HSCs, which form MPPs prior to lineage commitment. In the updated model, stem cells can be multipotent (HSCs) or myeloid-restricted (MySCs). HSCs can generate MySCs via the myeloid-bypass pathway, but it is unclear if MySCs can generate HSCs. Loss of self-renewal generates repopulating progenitors (RPs), which can be multipotent (multiRPs) or myeloid-restricted (MyRPs).
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Recent Articles
Stanford Medicine News Center, May 29, 2019
Radiation-free stem cell transplants, gene therapy may be within reach
Researchers at Stanford and the University of Tokyo may have cracked the code to doing stem cell transplants and gene therapy without radiation and chemotherapy.
For decades, researchers have been stymied in their attempts to grow large numbers of hematopoietic stem cells in the laboratory. These rare bone marrow cells are solely responsible for generating all the cells of the blood and immune system. Difficulties in growing the cells have seriously hampered many research efforts, including those aimed at making stem cell transplantation or gene therapy in patients with certain cancers or blood disorders easier and safer.
Now, researchers at the Stanford University School of Medicine and the University of Tokyo have cracked the code. By tinkering with the components of the nutritive broth in which the cells are grown, the specialized molecules used to support their growth and the physical conditions under which the cells are cultivated, the researchers have shown for the first time that it’s possible to coax hematopoietic stem cells from mice to renew themselves hundreds or even thousands of times within a period of just 28 days.
“This has been one of my life goals as a stem cell researcher,” said Hiromitsu Nakauchi, MD, PhD, professor of genetics at Stanford. “For 50 years, researchers from laboratories around the world have been seeking ways to grow these cells to large numbers. Now we’ve identified a set of conditions that allows these cells to expand in number as much as 900-fold in just one month. We believe this approach could transform how hematopoietic stem cell transplants and gene therapy are performed in humans.”
In particular, the researchers have shown it is possible to successfully transplant large numbers of the cells into mice without first eliminating the recipients’ own stem cell population. If the technique also works in humans, it could save thousands of patients with blood or immune disorders from a grueling regimen of radiation or chemotherapy prior to transplant. It could also allow clinicians to use a patient’s own genetically corrected stem cells for gene therapy.
We believe this approach could transform how hematopoietic stem cell transplants and gene therapy are performed in humans.
The study was published online May 29 in Nature. Nakauchi shares senior authorship of the study with Satoshi Yamazaki, PhD, an associate professor of stem cell biology at the University of Tokyo. Postdoctoral scholar Adam Wilkinson, PhD, of Stanford and senior research assistant Reiko Ishida of the University of Tokyo are the lead authors.
Hematopoietic stem cells are rare cells found in the bone marrow. Like other stem cells, they can either divide to make more hematopoietic stem cells — a process called self-renewal — or generate the precursors of all the different types of blood and immune cells in the body — a process called differentiation.
It’s long been known that people with immune or blood disorders such as sickle cell anemia or leukemia can be cured with a transplant of healthy hematopoietic stem cells. But in order for the treatment to work, the recipient’s own hematopoietic stem cells must be killed to eliminate the disease and make space for the healthy cells to settle in the bone marrow.
This elimination step, also called “conditioning,” is accomplished with either chemotherapy or radiation, or a combination of the two. The conditioning, however, can cause life-threatening side effects.
This is particularly true in pediatric patients, who can suffer side effects such as growth retardation, infertility and secondary cancer in later life. Very sick or elderly patients often can’t receive transplants because they are unable to tolerate the conditioning treatment.
‘A Holy Grail in the stem cell field’
But for some time, researchers have wondered whether transplanting large numbers of donor hematopoietic stem cells could circumvent the need to remove the existing cells. Perhaps, they reasoned, swamping the recipient’s bone marrow with a tide of healthy donor cells would allow the newcomers to muscle their way in and set up shop making healthy blood and immune cells.
It’s been difficult to test this theory, however, because hematopoietic stem cells are hard to isolate in large numbers. And although it’s been possible for years to grow human hematopoietic stem cells in the laboratory, the cells never self-renew robustly and instead often abandon their stem cell fate to differentiate into precursor cells. As a result, it’s been difficult to study their biology in depth or to generate enough cells to attempt large-scale transplants.
“Expansion of hematopoietic stem cells has been a Holy Grail in the stem cell field,” Nakauchi said. “When something does not work, people tend to think that something is missing. But we decided to try the opposite approach by eliminating all the impurities in the conventional culture system, while also optimizing other aspects that encourage self-renewal of the hematopoietic stem cells.”
The researchers also replaced a potentially contaminated blood protein called serum albumin with polyvinyl alcohol, a water-soluble synthetic chemical that is frequently used in biomedical research.
These modifications to how the cells were grown allowed the researchers to generate enough hematopoietic stem cells for transplant from just 50 starting cells. Although some of the cells did differentiate in culture, many more than normal maintained their stem cell identity throughout the duration of the culture.
“These original 50 cells increased in number about 8,000-fold over 28 days,” Nakauchi said. “Of these, about one of every 35 cells remained a functional hematopoietic stem cell.”
Planning to test human cells
The researchers estimated that the hematopoietic stem cells in the original sample increased in numbers by between 200- to 900-fold — an unprecedented level of expansion. When they transplanted the newly grown cells into mice that had not undergone a conditioning regimen, the animals developed blood and immune cells derived from both the donors’ hematopoietic stem cells and their own, demonstrating that the donor cells had engrafted and remained functional.
“We also found that, during the culture, we can use CRISPR technology to correct any genetic defects in the original hematopoietic cells,” Nakauchi said. “These gene-corrected cells can then be expanded for transplantation. This should allow us to use a patient’s own cells as gene therapy.”
Nakauchi and his collaborators at Stanford are now testing this approach in mice.
Other Stanford co-authors of the study are graduate student Ralph Crisostomo, research associate Ryo Yamamoto, and assistant professor of developmental biology Kyle Loh, PhD. Researchers from the RIKEN BioResource Center also contributed to the study.
The research was supported by the Japan Agency for Medical Research and Development, the Japan Society for the Promotion of Science, the National Institutes of Health (grants R01DK116944, R01HL147124, and DP5OD024558), the Ludwig Foundation, the California Institute for Regenerative Medicine, the Leukemia and Lymphoma Society, the Siebel Stem Cell Institute, the Baxter Foundation, Bloodwise and anonymous donors.
Nakauchi is a co-founder and shareholder of ReproCELL Inc.
Stanford’s Department of Genetics also supported the work.
By Krista Conger