Page 325 - 2021_02-Haematologica-web

Letters to the Editor
3C, cells harvested from WT- and G1s-clones showed similar morphology in P-erymk41(+) and P-erymk41(-). However, when each subpopulation was separately cul- tured, both fractions from the G1s-clones presented sig- nificantly higher levels in cell number expansion under the myeloid-specific differentiation condition (Figure 3D, E). This change suggested we could use this assay to delineate the impact of the GATA1 mutation in abnormal cell proliferation. Moreover, the difference in cell num- ber expansion revealed that P-erymk41(+) was signifi- cantly affected more by the GATA1 mutation only under myeloid-specific differentiation condition (Figure 3F; Online Supplementary Figure S7C) verifying the network analysis described above. On the other hand, under ery- throid-megakaryocytic condition, P-erymk41(+) and P-erymk41(-) G1s-clones did not show any significant increase in total cell numbers compared with WT-clones (Online Supplementary Figure S7D). These data together demonstrated P-erymk41(+) is a progenitor intermediate that contributes strongly to abnormal myeloid cell pro- liferation (Figure 3G).
In summary, the current study identified the disease- responsible progenitors and the impact of GATA1 muta- tion on these cells by dissecting the differentiation steps of PSC-based hematopoiesis. The abnormal cells observed in patients with TAM have long been thought as being derived from definitive hematopoiesis,13 and recent studies have shown that PSC-derived definitive hematopoiesis predominantly arises from KDR+ meso- dermal progenitors via CD235a-CD34+ subpopulations.14 Given that our data are provided from those specified subpopulations, our Ts21-PSC-based model likely describes definitive hematopoiesis in vitro. Thus, our hematopoietic induction model adequately reflects the pathogenesis of TAM.
P-erymk41(+) is an inflection stage for both impaired erythro-megakaryocyte maturation and abnormal cell proliferation in TAM specifically in response to erythro- megakaryocytic and myeloid-directed stimulation, respectively. Together, we speculate that P-erymk41(+) can be a target of pre-emptive treatment for TAM. Our data also suggest that DNA damage responses may accu- mulate in P-erymk41(+) in the presence of GATA1 muta- tion. As TAM is a condition preceding Down syndrome acute megakaryoblastic leukemia (DS-AMKL) and as DS-AMKL is considered the consequence of acquired gene mutations,2 it is important to identify the most appropriate cell population from which the functional and molecular consequences of GATA1 mutation begins. From that viewpoint, our model should provide novel insights into the pathogenesis, prediction, and treatment of not only TAM, but also acute megakaryocytic leukemia in Down syndrome, though further analysis is needed.
Yoko Nishinaka-Arai,1,2* Akira Niwa,1* Shiori Matsuo,1 Yasuhiro Kazuki,3,4 Yuwna Yakura,3 Takehiko Hiroma,5 Tsutomu Toki,6 Tetsushi Sakuma,7 Takashi Yamamoto,7 Etsuro Ito,6 Mitsuo Oshimura,3 Tatsutoshi Nakahata8 and Megumu K. Saito1
1Department of Clinical Application, Center for iPS cell Research and Application, Kyoto University, Kyoto; 2Department of Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto; 3Chromosome Engineering Research Center, Tottori University, Tottori; 4Division of Genome and Cellular Functions, Department of Molecular and Cellular Biology, School of Life Science, Faculty of Medicine, Tottori University, Tottori; 5Perinatal Medical Center, Nagano Children’s Hospital, Nagano; 6Department of Pediatrics, Hirosaki University Graduate School of
Medicine, Hirosaki; 7Division of Integrated Sciences for Life, Graduate School of Integrated Sciences for Life, Hiroshima University, Hiroshima and 8Drug Discovery Technology Development Office, Center for iPS cell Research and Application, Kyoto University, Kyoto, Japan
*YNA and AN contributed equally as co-first authors
Disclosures: No conflicts of interest to disclose.
Contributions: YNA and AN designed and performed experi- ments and analyzed data; SM performed experiments; YK, YY and MO provided Ts21-KhES1 cells and contributed to scientific discussion; TS and TY designed and made plasmids for gene editing; TH, TT and EI obtained informed consent from a patient and harvested primary cells; TN and MKS supervised the project and experimental design; YNA, AN and MKS wrote the paper.
Funding: this work was supported by grants from the Ministry of Health, Labour and Welfare of Japan, Ministry of Education, Culture, Sports, Science and Technology of Japan, Japan Society for the Promotion of Science (JSPS) Research Fellowships for Young Scientists [YNA], KAKENHI Grant Number 19K17358 [YNA], KAKENHI Grant Number 25221308 [MO] and KAK- ENHI Grant Number 16K10026 [AN], Regional Innovation Support Program from the Ministry of Education, Culture, Sports, Science and Technology of Japan [MO and YK], the Leading Project of the Ministry of Education, Culture, Sports, Science and Technology [TN], the Funding Program for World-Leading Innovative Research and Development on Science and Technology of the Japan Society for the Promotion of Science [TN and MKS], CREST [TN], the Core Center for iPS Cell Research of Research Center Network for Realization of Regenerative Medicine from the Japan Agency for Medical Research and Development (AMED) [TN and MKS], the Program for Intractable Diseases Research utilizing Disease-specific iPS cells of AMED (17935423) [YNA, TN and MKS], and Practical Research Project for Rare/Intractable Diseases of AMED (17930095) [MKS].
Correspondence:
MEGUMU K. SAITO - msaito@cira.kyoto-u.ac.jp AKIRA NIWA - akiranw@cira.kyoto-u.ac.jp
doi:10.3324/haematol.2019.242693
References
1. Gamis AS, Alonzo TA, Gerbing RB. Natural history of transient myeloproliferative disorder clinically diagnosed in Down syn- drome neonates: A report from the Children’s Oncology Group Study A2971. Blood. 2011;118(26):6752-6759.
2.Yoshida K, Toki T, Okuno Y, et al. The landscape of somatic mutations in Down syndrome-related myeloid disorders. Nat Genet. 2013;45(11):1293-1299.
3. Chou ST, Byrska-Bishop M, Tober JM, et al. Trisomy 21-associat- ed defects in human primitive hematopoiesis revealed through induced pluripotent stem cells. Proc Nat Acad Sci U S A. 2012;109(43):17573-17578.
4.Byrska-Bishop M, VanDorn D, Campbell AE, et al. Pluripotent stem cells reveal erythroid-specific activities of the GATA1 N-ter- minus. J Clin Invest. 2015;125(3):993-1005.
5. Banno K, Omori S, Hirata K, et al. Systematic cellular disease models reveal synergistic interaction of trisomy 21 and GATA1 mutations in hematopoietic abnormalities. Cell Rep. 2016;15(6):1228-1241.
6. Niwa A, Heike T, Umeda K, et al. A novel serum-free monolayer culture for orderly hematopoietic differentiation of human pluripotent cells via mesodermal progenitors. PLoS One. 2011;6(7):e22261.
7. Yanagimachi MD, Niwa A, Tanaka T, et al. Robust and highly- efficient differentiation of functional monocytic cells from human pluripotent stem cells under serum- and feeder cell-free condi- tions. PLoS One. 2013;8(4):e59243.
8. Sakuma T, Ochiai H, Kaneko T, et al. Repeating pattern of non- RVD variations in DNA-binding modules enhances TALEN activ- ity. Sci Rep. 2013;3:3379.
9. Kazuki Y, Hoshiya H, Kai Y, et al. Correction of a genetic defect
haematologica | 2021; 106(2)
639