||Sahoko Matsuoka, Raffaella Facchini, Tiago C Luis, Joana Carrelha, Petter S Woll, Takuo Mizukami, Bishan Wu, Hanane Boukarabila, Mario Buono, Ruggiero Norfo, Fumio Arai, Toshio Suda, Adam Mead, Claus Nerlov, Sten Eirik W Jacobsen, Loss of endothelial membrane KIT Ligand affects systemic KIT ligand levels but not bone marrow hematopoietic stem cells., Blood, 10.1182/blood.2022019018, 2023.08, A critical regulatory role of hematopoietic stem cell vascular niches in the bone marrow has been implicated to occur through endothelial niche cell expression of KIT Ligand. However, endothelial-derived KIT Ligand is expressed in both a soluble and membrane-bound form, and not unique to bone marrow niches and is also systemically distributed through the circulatory system. Here we confirm that upon deletion of both the soluble and membrane-bound form of endothelial-derived KIT Ligand hematopoietic stem cells are reduced in mouse bone marrow. However, deletion of endothelial-derived KIT Ligand was also accompanied by reduced soluble KIT Ligand levels in blood, precluding any conclusion as to whether the reduction in HSC numbers reflect reduced endothelial expression of KIT Ligand within HSC niches, elsewhere in the bone marrow and/or systemic sKIT Ligand produced by endothelial cells outside of the bone marrow. Notably, endothelial deletion specifically of the membrane bound form of KIT Ligand also reduced systemic levels of soluble KIT Ligand although with no effect on stem cell numbers, implicating a hematopoietic stem cell regulatory role primarily of soluble rather than membrane KIT Ligand expression in endothelial cells. In support of a role of systemic rather than local niche expression of soluble KIT Ligand, hematopoietic stem cells were unaffected in bones with deletion of KIT ligand when implanted in mice with normal systemic levels of soluble KIT Ligand. Our findings highlight the need for more specific tools to unravel niche-specific roles of regulatory cues expressed in hematopoietic niche cells in the bone marrow..
||Patrick S Stumpf, Fumio Arai, Ben D MacArthur, Modeling Stem Cell Fates using Non-Markov Processes., Cell stem cell, 10.1016/j.stem.2021.01.009, 28, 2, 187-190, 2021.02, Epigenetic memories play an important part in regulating stem cell identities. Tools from the theory of non-Markov processes may help us understand these memories better and develop a more integrated view of stem cell fate and function..
||Patrick S Stumpf, Xin Du, Haruka Imanishi, Yuya Kunisaki, Yuichiro Semba, Timothy Noble, Rosanna C G Smith, Matthew Rose-Zerili, Jonathan J West, Richard O C Oreffo, Katayoun Farrahi, Mahesan Niranjan, Koichi Akashi, Fumio Arai, Ben D MacArthur, Transfer learning efficiently maps bone marrow cell types from mouse to human using single-cell RNA sequencing., Communications biology, 10.1038/s42003-020-01463-6, 3, 1, 736-736, 2020.12, Biomedical research often involves conducting experiments on model organisms in the anticipation that the biology learnt will transfer to humans. Previous comparative studies of mouse and human tissues were limited by the use of bulk-cell material. Here we show that transfer learning-the branch of machine learning that concerns passing information from one domain to another-can be used to efficiently map bone marrow biology between species, using data obtained from single-cell RNA sequencing. We first trained a multiclass logistic regression model to recognize different cell types in mouse bone marrow achieving equivalent performance to more complex artificial neural networks. Furthermore, it was able to identify individual human bone marrow cells with 83% overall accuracy. However, some human cell types were not easily identified, indicating important differences in biology. When re-training the mouse classifier using data from human, less than 10 human cells of a given type were needed to accurately learn its representation. In some cases, human cell identities could be inferred directly from the mouse classifier via zero-shot learning. These results show how simple machine learning models can be used to reconstruct complex biology from limited data, with broad implications for biomedical research..
||Fumio Arai, Patrick S Stumpf, Yoshiko M Ikushima, Kentaro Hosokawa, Aline Roch, Matthias P Lutolf, Toshio Suda, Ben D MacArthur, Machine Learning of Hematopoietic Stem Cell Divisions from Paired Daughter Cell Expression Profiles Reveals Effects of Aging on Self-Renewal., Cell systems, 10.1016/j.cels.2020.11.004, 11, 6, 640-652, 2020.12, Changes in stem cell activity may underpin aging. However, these changes are not completely understood. Here, we combined single-cell profiling with machine learning and in vivo functional studies to explore how hematopoietic stem cell (HSC) divisions patterns evolve with age. We first trained an artificial neural network (ANN) to accurately identify cell types in the hematopoietic hierarchy and predict their age from single-cell gene-expression patterns. We then used this ANN to compare identities of daughter cells immediately after HSC divisions and found that the self-renewal ability of individual HSCs declines with age. Furthermore, while HSC cell divisions are deterministic and intrinsically regulated in young and old age, they are variable and niche sensitive in mid-life. These results indicate that the balance between intrinsic and extrinsic regulation of stem cell activity alters substantially with age and help explain why stem cell numbers increase through life, yet regenerative potency declines..
||Kentaro Hosokawa, Ben D. MacArthur, Yoshiko Matsumoto Ikushima, Hirofumi Toyama, Yoshikazu Masuhiro, Shigemasa Hanazawa, Toshio Suda, Fumio Arai, The telomere binding protein Pot1 maintains haematopoietic stem cell activity with age., Nature Communications, 10.1038/s41467-017-00935-4, 8, 1, 2017.10, Repeated cell divisions and aging impair stem cell function. However, the mechanisms by which this occurs are not fully understood. Here we show that protection of telomeres 1A (Pot1a), a component of the Shelterin complex that protects telomeres, improves haematopoietic stem cell (HSC) activity during aging. Pot1a is highly expressed in young HSCs, but
declines with age. In mouse HSCs, Pot1a knockdown increases DNA damage response (DDR) and inhibits self-renewal. Conversely, Pot1a overexpression or treatment with POT1a protein prevents DDR, maintained self-renewal activity and rejuvenated aged HSCs upon ex vivo culture. Moreover, treatment of HSCs with exogenous Pot1a inhibits the production of reactive oxygen species, suggesting a non-telomeric role for Pot1a in HSC maintenance. Consistent with these results, treatment with exogenous human POT1 protein maintains human HSC activity in culture. Collectively, these results show that Pot1a/POT1 sustains HSC activity and can be used to expand HSC numbers ex vivo..
||Kyoko Ito, Raphaël Turcotte, Jinhua Cui, Samuel E. Zimmerman, Sandra Pinho, Toshihide Mizoguchi, Fumio Arai, Judith M. Runnels, Clemens Alt, Julie Teruya-Feldstein, Jessica C. Mar, Rajat Singh, Toshio Suda, Charles P. Lin, Paul S. Frenette, Keisuke Ito, Self-renewal of a purified Tie2+ hematopoietic stem cell population relies on mitochondrial clearance, Science, 10.1126/science.aaf5530, 354, 6316, 1156-1160, 2016.12, A single hematopoietic stem cell (HSC) is capable of reconstituting hematopoiesis and maintaining homeostasis by balancing self-renewal and cell differentiation. The mechanisms of HSC division balance, however, are not yet defined. Here we demonstrate, by characterizing at the single-cell level a purified and minimally heterogeneous murine Tie2+ HSC population, that these top hierarchical HSCs preferentially undergo symmetric divisions. The induction of mitophagy, a quality control process in mitochondria, plays an essential role in self-renewing expansion of Tie2+ HSCs. Activation of the PPAR (peroxisome proliferator-activated receptor)-fatty acid oxidation pathway promotes expansion of Tie2+ HSCs through enhanced Parkin recruitment in mitochondria. These metabolic pathways are conserved in human TIE2+ HSCs. Our data thus identify mitophagy as a key mechanism of HSC expansion and suggest potential methods of cell-fate manipulation through metabolic pathways..
||Yoshiko Matsumoto Ikushima, Fumio Arai, Kentaro Hosokawa, Hirofumi Toyama, Keiyo Takubo, Tomoyuki Furuyashiki, Shuh Narumiya, Toshio Suda, Prostaglandin E2 regulates murine hematopoietic stem/progenitor cells directly via EP4 receptor and indirectly through mesenchymal progenitor cells, Blood, 10.1182/blood-2012-06-437889, 121, 11, 1995-2007, 2013.03, Key Points
PGE2 signaling positively regulates hematopoietic stem cells both directly and via activation of a nonhematopoietic cell population. EP4 is a major receptor for the PGE2-mediated regulation of hematopoietic stem and progenitor cells..
||Yuka Nakamura, Fumio Arai, Hiroko Iwasaki, Kentaro Hosokawa, Isao Kobayashi, Yumiko Gomei, Yoshiko Matsumoto, Hiroki Yoshihara, Toshio Suda, Isolation and characterization of endosteal niche cell populations that regulate hematopoietic stem cells, Blood, 10.1182/blood-2009-08-239194, 116, 9, 1422-1432, 2010.09, The endosteal niche is critical for the maintenance of hematopoietic stem cells (HSCs). However, it consists of a heterogeneous population in terms of differentiation stage and function. In this study, we characterized endosteal cell populations and examined their ability to maintain HSCs. Bone marrow endosteal cells were subdivided into immature mesenchymal cell-enriched ALCAM-Sca-1+ cells, osteoblast-enriched ALCAM +Sca-1-, and ALCAM-Sca-1- cells. We found that all 3 fractions maintained long-term reconstitution (LTR) activity of HSCs in an in vitro culture. In particular, ALCAM+Sca-1- cells significantly enhanced the LTR activity of HSCs by the up-regulation of homing-and cell adhesion-related genes in HSCs. Microarray analysis showed that ALCAM-Sca-1+ fraction highly expressed cytokine-related genes, whereas the ALCAM+Sca-1- fraction expressed multiple cell adhesion molecules, such as cadherins, at a greater level than the other fractions, indicating that the interaction between HSCs and osteoblasts via cell adhesion molecules enhanced the LTR activity of HSCs. Furthermore, we found an osteoblastic markerlow/- subpopulation in ALCAM +Sca-1- fraction that expressed cytokines, such as Angpt1 and Thpo, and stem cell marker genes. Altogether, these data suggest that multiple subsets of osteoblasts and mesenchymal progenitor cells constitute the endosteal niche and regulate HSCs in adult bone marrow..
||Kentaro Hosokawa, Fumio Arai, Hiroki Yoshihara, Hiroko Iwasaki, Yuka Nakamura, Yumiko Gomei, Toshio Suda, Knockdown of N-cadherin suppresses the long-term engraftment of hematopoietic stem cells, Blood, 10.1182/blood-2009-05-224857, 116, 4, 554-563, 2010.07, During postnatal life, the bone marrow (BM) supports both self-renewal and differentiation of hematopoietic stem cells (HSCs) in specialized microenvironments termed stem cell niches. Cell-cell and cell-extracellular matrix interactions between HSCs and their niches are critical for the maintenance of HSC properties. Here, we analyzed the function of Ncadherin in the regulation of the proliferation and long-term repopulation activity of hematopoietic stem/progenitor cells (HSPCs) by the transduction of N-cadherin shRNA. Inhibition of N-cadherin expression accelerated cell division in vitro and reduced the lodgment of donor HSPCs to the endosteal surface, resulting in a significant reduction in long-term engraftment. Cotransduction of N-cadherin shRNA and a mutant N-cadherin that introduced the silent mutations to shRNA target sequences rescued the accelerated cell division and reconstitution phenotypes. In addition, the requirement of N-cadherin for HSPC engraftment appears to be niche specific, as shN-cad-transduced lineage-Sca- 1+c-Kit+ cells successfully engrafted in spleen, which lacks an osteoblastic niche. These findings suggest that N-cad-mediated cell adhesion is functionally required for the establishment of hematopoiesis in the BM niche after BM transplantation..
||Kentaro Hosokawa, Fumio Arai, Hiroki Yoshihara, Hiroko Iwasaki, Mark Hembree, Tong Yin, Yuka Nakamura, Yumiko Gomei, Keiyo Takubo, Haruko Shiama, Sahoko Matsuoka, Linheng Li, Toshio Suda, Cadherin-Based Adhesion Is a Potential Target for Niche Manipulation to Protect Hematopoietic Stem Cells in Adult Bone Marrow, Cell stem cell, 10.1016/j.stem.2009.04.013, 6, 3, 194-198, 2010.03.
||Hiroki Yoshihara, Fumio Arai, Kentaro Hosokawa, Tetsuya Hagiwara, Keiyo Takubo, Yuka Nakamura, Yumiko Gomei, Hiroko Iwasaki, Sahoko Matsuoka, Kana Miyamoto, Hiroshi Miyazaki, Takao Takahashi, Toshio Suda, Thrombopoietin/MPL Signaling Regulates Hematopoietic Stem Cell Quiescence and Interaction with the Osteoblastic Niche, Cell stem cell, 10.1016/j.stem.2007.10.020, 1, 6, 685-697, 2007.12, Maintenance of hematopoietic stem cells (HSCs) depends on interaction with their niche. Here we show that the long-term (LT)-HSCs expressing the thrombopoietin (THPO) receptor, MPL, are a quiescent population in adult bone marrow (BM) and are closely associated with THPO-producing osteoblastic cells. THPO/MPL signaling upregulated β1-integrin and cyclin-dependent kinase inhibitors in HSCs. Furthermore, inhibition and stimulation of THPO/MPL pathway by treatments with anti-MPL neutralizing antibody, AMM2, and with THPO showed reciprocal regulation of quiescence of LT-HSC. AMM2 treatment reduced the number of quiescent LT-HSCs and allowed exogenous HSC engraftment without irradiation. By contrast, exogenous THPO transiently increased quiescent HSC population and subsequently induced HSC proliferation in vivo. Altogether, these observations suggest that THPO/MPL signaling plays a critical role of LT-HSC regulation in the osteoblastic niche..
||Fumio Arai, Atsushi Hirao, Masako Ohmura, Hidetaka Sato, Sahoko Matsuoka, Keiyo Takubo, Keisuke Ito, Gou Young Koh, Toshio Suda, Tie2/angiopoietin-1 signaling regulates hematopoietic stem cell quiescence in the bone marrow niche, Cell, 10.1016/j.cell.2004.07.004, 118, 2, 149-161, 2004.07, The quiescent state is thought to be an indispensable property for the maintenance of hematopoietic stem cells (HSCs). Interaction of HSCs with their particular microenvironments, known as the stem cell niches, is critical for adult hematopoiesis in the bone marrow (BM). Here, we demonstrate that HSCs expressing the receptor tyrosine kinase Tie2 are quiescent and antiapoptotic, and comprise a side-population (SP) of HSCs, which adhere to osteoblasts (OBs) in the BM niche. The interaction of Tie2 with its ligand Angiopoietin-1 (Ang-1) induced cobblestone formation of HSCs in vitro and maintained in vivo long-term repopulating activity of HSCs. Furthermore, Ang-1 enhanced the ability of HSCs to become quiescent and induced adhesion to bone, resulting in protection of the HSC compartment from myelosuppressive stress. These data suggest that the Tie2/Ang-1 signaling pathway plays a critical role in the maintenance of HSCs in a quiescent state in the BM niche..