Chemically Defined Cytokine-free Expansion of Human Haematopoietic Stem Cells

Overview

Journal Nature

Specialty Science

Date 2023 Feb 22

PMID 36813966

Authors

Masatoshi Sakurai

Kantaro Ishitsuka

Ryoji Ito

Adam C Wilkinson

Takaharu Kimura

Eiji Mizutani

Hidekazu Nishikii

Kazuhiro Sudo

Hans Jiro Becker

Hiroshi Takemoto

Tsubasa Sano

Keisuke Kataoka

Satoshi Takahashi

Yukio Nakamura

David G Kent

Atsushi Iwama

Shigeru Chiba

Shinichiro Okamoto

Hiromitsu Nakauchi

Satoshi Yamazaki

Affiliations

Soon will be listed here.

Abstract

Haematopoietic stem cells (HSCs) are a rare cell type that reconstitute the entire blood and immune systems after transplantation and can be used as a curative cell therapy for a variety of haematological diseases. However, the low number of HSCs in the body makes both biological analyses and clinical application difficult, and the limited extent to which human HSCs can be expanded ex vivo remains a substantial barrier to the wider and safer therapeutic use of HSC transplantation. Although various reagents have been tested in attempts to stimulate the expansion of human HSCs, cytokines have long been thought to be essential for supporting HSCs ex vivo. Here we report the establishment of a culture system that allows the long-term ex vivo expansion of human HSCs, achieved through the complete replacement of exogenous cytokines and albumin with chemical agonists and a caprolactam-based polymer. A phosphoinositide 3-kinase activator, in combination with a thrombopoietin-receptor agonist and the pyrimidoindole derivative UM171, were sufficient to stimulate the expansion of umbilical cord blood HSCs that are capable of serial engraftment in xenotransplantation assays. Ex vivo HSC expansion was further supported by split-clone transplantation assays and single-cell RNA-sequencing analysis. Our chemically defined expansion culture system will help to advance clinical HSC therapies.

Citing Articles

Soluplus-Based Pharmaceutical Formulations: Recent Advances in Drug Delivery and Biomedical Applications.

Guembe-Michel N, Nguewa P, Gonzalez-Gaitano G Int J Mol Sci. 2025; 26(4).

PMID: 40003966 PMC: 11855892. DOI: 10.3390/ijms26041499.

UM171 glues asymmetric CRL3-HDAC1/2 assembly to degrade CoREST corepressors.

Yeo M, Zhang O, Xie X, Nam E, Payne N, Gosavi P Nature. 2025; 639(8053):232-240.

PMID: 39939761 PMC: 11882444. DOI: 10.1038/s41586-024-08532-4.

Methylmalonic acid at the serum level in the elderly contributes to cell growth via mitochondrial dysfunction in colorectal cancer cell spheroids.

Tanaka A, Murakami C, Yamamoto H Biochem Biophys Rep. 2025; 41:101909.

PMID: 39886070 PMC: 11780164. DOI: 10.1016/j.bbrep.2024.101909.

Regulatory cellular and molecular networks in the bone microenvironment during aging.

Zhang L, Wang Z, Zhang Y, Ji R, Li Z, Zou J Life Med. 2025; 3(3):lnae019.

PMID: 39871887 PMC: 11749081. DOI: 10.1093/lifemedi/lnae019.

Regulation and therapy: the role of ferroptosis in DLBCL.

Wang Y, He Z, Dong X, Yao Y, Chen Q, Shi Y Front Pharmacol. 2025; 15():1458412.

PMID: 39834804 PMC: 11743434. DOI: 10.3389/fphar.2024.1458412.

References

Copelan E . Hematopoietic stem-cell transplantation. N Engl J Med. 2006; 354(17):1813-26. DOI: 10.1056/NEJMra052638. View

Cohen S, Roy J, Lachance S, Delisle J, Marinier A, Busque L . Hematopoietic stem cell transplantation using single UM171-expanded cord blood: a single-arm, phase 1-2 safety and feasibility study. Lancet Haematol. 2019; 7(2):e134-e145. DOI: 10.1016/S2352-3026(19)30202-9. View

Pineault N, Abu-Khader A . Advances in umbilical cord blood stem cell expansion and clinical translation. Exp Hematol. 2015; 43(7):498-513. DOI: 10.1016/j.exphem.2015.04.011. View

Gluckman E, Broxmeyer H, Auerbach A, Friedman H, Douglas G, Devergie A . Hematopoietic reconstitution in a patient with Fanconi's anemia by means of umbilical-cord blood from an HLA-identical sibling. N Engl J Med. 1989; 321(17):1174-8. DOI: 10.1056/NEJM198910263211707. View

Orkin S, Zon L . Hematopoiesis: an evolving paradigm for stem cell biology. Cell. 2008; 132(4):631-44. PMC: 2628169. DOI: 10.1016/j.cell.2008.01.025. View

Weissman I . Stem cells: units of development, units of regeneration, and units in evolution. Cell. 2000; 100(1):157-68. DOI: 10.1016/s0092-8674(00)81692-x. View

Wilkinson A, Igarashi K, Nakauchi H . Haematopoietic stem cell self-renewal in vivo and ex vivo. Nat Rev Genet. 2020; 21(9):541-554. PMC: 7894993. DOI: 10.1038/s41576-020-0241-0. View

Boitano A, Wang J, Romeo R, Bouchez L, Parker A, Sutton S . Aryl hydrocarbon receptor antagonists promote the expansion of human hematopoietic stem cells. Science. 2010; 329(5997):1345-8. PMC: 3033342. DOI: 10.1126/science.1191536. View

Fares I, Chagraoui J, Gareau Y, Gingras S, Ruel R, Mayotte N . Cord blood expansion. Pyrimidoindole derivatives are agonists of human hematopoietic stem cell self-renewal. Science. 2014; 345(6203):1509-12. PMC: 4372335. DOI: 10.1126/science.1256337. View

10.

Wagner Jr J, Brunstein C, Boitano A, DeFor T, McKenna D, Sumstad D . Phase I/II Trial of StemRegenin-1 Expanded Umbilical Cord Blood Hematopoietic Stem Cells Supports Testing as a Stand-Alone Graft. Cell Stem Cell. 2015; 18(1):144-55. PMC: 4881386. DOI: 10.1016/j.stem.2015.10.004. View

11.

Bai T, Li J, Sinclair A, Imren S, Merriam F, Sun F . Expansion of primitive human hematopoietic stem cells by culture in a zwitterionic hydrogel. Nat Med. 2019; 25(10):1566-1575. DOI: 10.1038/s41591-019-0601-5. View

12.

Grey W, Chauhan R, Piganeau M, Huerga Encabo H, Garcia-Albornoz M, McDonald N . Activation of the receptor tyrosine kinase RET improves long-term hematopoietic stem cell outgrowth and potency. Blood. 2020; 136(22):2535-2547. PMC: 7714096. DOI: 10.1182/blood.2020006302. View

13.

Huang J, Nguyen-McCarty M, Hexner E, Danet-Desnoyers G, Klein P . Maintenance of hematopoietic stem cells through regulation of Wnt and mTOR pathways. Nat Med. 2012; 18(12):1778-85. PMC: 3518679. DOI: 10.1038/nm.2984. View

14.

Wilkinson A, Ishida R, Kikuchi M, Sudo K, Morita M, Crisostomo R . Long-term ex vivo haematopoietic-stem-cell expansion allows nonconditioned transplantation. Nature. 2019; 571(7763):117-121. PMC: 7006049. DOI: 10.1038/s41586-019-1244-x. View

15.

Wilkinson A, Ishida R, Nakauchi H, Yamazaki S . Long-term ex vivo expansion of mouse hematopoietic stem cells. Nat Protoc. 2020; 15(2):628-648. PMC: 7206416. DOI: 10.1038/s41596-019-0263-2. View

16.

Seita J, Ema H, Ooehara J, Yamazaki S, Tadokoro Y, Yamasaki A . Lnk negatively regulates self-renewal of hematopoietic stem cells by modifying thrombopoietin-mediated signal transduction. Proc Natl Acad Sci U S A. 2007; 104(7):2349-54. PMC: 1892983. DOI: 10.1073/pnas.0606238104. View

17.

Park H, Li J, Hannah R, Biddie S, Leal-Cervantes A, Kirschner K . Cytokine-induced megakaryocytic differentiation is regulated by genome-wide loss of a uSTAT transcriptional program. EMBO J. 2015; 35(6):580-94. PMC: 4801948. DOI: 10.15252/embj.201592383. View

18.

Yamazaki S, Iwama A, Takayanagi S, Morita Y, Eto K, Ema H . Cytokine signals modulated via lipid rafts mimic niche signals and induce hibernation in hematopoietic stem cells. EMBO J. 2006; 25(15):3515-23. PMC: 1538571. DOI: 10.1038/sj.emboj.7601236. View

19.

Miyamoto K, Araki K, Naka K, Arai F, Takubo K, Yamazaki S . Foxo3a is essential for maintenance of the hematopoietic stem cell pool. Cell Stem Cell. 2008; 1(1):101-112. DOI: 10.1016/j.stem.2007.02.001. View

20.

Tadokoro Y, Hoshii T, Yamazaki S, Eto K, Ema H, Kobayashi M . Spred1 Safeguards Hematopoietic Homeostasis against Diet-Induced Systemic Stress. Cell Stem Cell. 2018; 22(5):713-725.e8. DOI: 10.1016/j.stem.2018.04.002. View