Abstract
Efforts have been made to establish various human pluripotent stem cell lines. However, such methods have not yet been duplicated in non-human primate cells. Here, we introduce a multiplexed single-cell sequencing technique to profile the molecular features of monkey pluripotent stem cells in published culture conditions. The results demonstrate suboptimized maintenance of pluripotency and show that the selected signaling pathways for resetting human stem cells can also be interpreted for establishing monkey cell lines. Overall, this work legitimates the translation of novel human cell line culture conditions to monkey cells and provides guidance for exploring chemical cocktails for monkey stem cell line derivation.
摘要
尽管人们已经开发了多种构建人类多能干细胞的方法, 但这些方法仍未应用在非人灵长类细胞中. 在本文中, 我们利用多样品标记单细胞测序技术对不同培养条件下的猴多能干细胞进行了分子特征的表征. 结果表明, 虽然用于重置人类干细胞的信号通路可以用来构建猴干细胞系, 但其配方有待优化才能更好地维持猴干细胞多能性. 总而言之, 本研究证实了新型的人类干细胞培养体系可以转化到猴干细胞体系中, 这对于未来猴干细胞系培养配方的探索具有指导意义.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Ai ZY, Niu BH, Duan K, et al., 2020. Modulation of Wnt and Activin/Nodal supports efficient derivation, cloning and suspension expansion of human pluripotent stem cells. Biomaterials, 249:120015. https://doi.org/10.1016/j.biomaterials.2020.120015
Bayerl J, Ayyash M, Shani T, et al., 2021. Principles of signaling pathway modulation for enhancing human naive pluripotency induction. Cell Stem Cell, 28(9):1549–1565.E12. https://doi.org/10.1016/j.stem.2021.04.001
Bredenoord AL, Clevers H, Knoblich JA, 2017. Human tissues in a dish: the research and ethical implications of organoid technology. Science, 355(6322):eaaf9414. https://doi.org/10.1126/science.aaf9414
Chen CX, Ji WZ, Niu YY, 2021. Primate organoids and gene-editing technologies toward next-generation biomedical research. Trends Biotechnol, 39(12):1332–1342. https://doi.org/10.1016/j.tibtech.2021.03.010
Chen HW, Aksoy I, Gonnot F, et al., 2015. Reinforcement of STAT3 activity reprogrammes human embryonic stem cells to naive-like pluripotency. Nat Commun, 6:7095. https://doi.org/10.1038/ncomms8095
Dong C, Beltcheva M, Gontarz P, et al., 2020. Derivation of trophoblast stem cells from naïve human pluripotent stem cells. eLife, 9:e52504. https://doi.org/10.7554/eLife.52504
Girgin MU, Broguiere N, Hoehnel S, et al., 2021. Bioengineered embryoids mimic post-implantation development in vitro. Nat Commun, 12:5140. https://doi.org/10.1038/s41467-021-25237-8
Jiang YQ, Chen CX, Randolph LN, et al., 2021. Generation of pancreatic progenitors from human pluripotent stem cells by small molecules. Stem Cell Rep, 16(9):2395–2409. https://doi.org/10.1016/j.stemcr.2021.07.021
Kempf H, Olmer R, Haase A, et al., 2016. Bulk cell density and Wnt/TGFbeta signalling regulate mesendodermal patterning of human pluripotent stem cells. Nat Commun, 7: 13602. https://doi.org/10.1038/ncomms13602
Kinoshita M, Barber M, Mansfield W, et al., 2021. Capture of mouse and human stem cells with features of formative pluripotency. Cell Stem Cell, 28(3):453–471.e8. https://doi.org/10.1016/j.stem.2020.11.005
Liu DH, Wang XY, He DJ, et al., 2018. Single-cell RNA-sequencing reveals the existence of naive and primed pluripotency in pre-implantation rhesus monkey embryos. Genome Res, 28(10):1481–1493. https://doi.org/10.1101/gr.233437.117
Ma HX, Zhai JL, Wan HF, et al., 2019. In vitro culture of cynomolgus monkey embryos beyond early gastrulation. Science, 366(6467):eaax7890. https://doi.org/10.1126/science.aax7890
Moris N, Anlas K, van den Brink SC, et al., 2020. An in vitro model of early anteroposterior organization during human development. Nature, 582(7812):410–415. https://doi.org/10.1038/s41586-020-2383-9
Niu YY, Sun NQ, Li C, et al., 2019. Dissecting primate early post-implantation development using long-term in vitro embryo culture. Science, 366(6467):eaaw5754. https://doi.org/10.1126/science.aaw5754
Pastor WA, Chen D, Liu WL, et al., 2016. Naive human pluripotent cells feature a methylation landscape devoid of blastocyst or germline memory. Cell Stem Cell, 18(3):323–329. https://doi.org/10.1016/j.stem.2016.01.019
Posfai E, Schell JP, Janiszewski A, et al., 2021. Evaluating totipotency using criteria of increasing stringency. Nat Cell Biol, 23(1):49–60. https://doi.org/10.1038/s41556-020-00609-2
Stoeckius M, Hafemeister C, Stephenson W, et al., 2017. Simultaneous epitope and transcriptome measurement in single cells. Nat Methods, 14(9):865–868. https://doi.org/10.1038/nmeth.4380
Stoeckius M, Zheng SW, Houck-Loomis B, et al., 2018. Cell Hashing with barcoded antibodies enables multiplexing and doublet detection for single cell genomics. Genome Biol, 19:224. https://doi.org/10.1186/s13059-018-1603-1
Stuart HT, van Oosten AL, Radzisheuskaya A, et al., 2014. NANOG amplifies STAT3 activation and they synergistically induce the naive pluripotent program. Curr Biol, 24(3):340–346. https://doi.org/10.1016/j.cub.2013.12.040
Stuart HT, Stirparo GG, Lohoff T, et al., 2019. Distinct molecular trajectories converge to induce naive pluripotency. Cell Stem Cell, 25(3):388–406.e8. https://doi.org/10.1016/j.stem.2019.07.009
Su J, Morgani SM, David CJ, et al., 2020. TGF-β orchestrates fibrogenic and developmental EMTs via the RAS effector RREB1. Nature, 577(7791):566–571. https://doi.org/10.1038/s41586-019-1897-5
Takahashi K, Tanabe K, Ohnuki M, et al., 2007. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell, 131(5):861–872. https://doi.org/10.1016/J.CELL.2007.11.019
Takashima Y, Guo G, Loos R, et al., 2015. Resetting transcription factor control circuitry toward ground-state pluripotency in human. Cell, 162(2):452–453. https://doi.org/10.1016/j.cell.2015.06.052
Tesar PJ, Chenoweth JG, Brook FA, et al., 2007. New cell lines from mouse epiblast share defining features with human embryonic stem cells. Nature, 448(7150):196–199. https://doi.org/10.1038/nature05972
Theunissen TW, Powell BE, Wang HY, et al., 2014. Systematic identification of culture conditions for induction and maintenance of naive human pluripotency. Cell Stem Cell, 15(4):471–487. https://doi.org/10.1016/j.stem.2014.07.002
Theunissen TW, Friedli M, He YP, et al., 2016. Molecular criteria for defining the naive human pluripotent state. Cell Stem Cell, 19(4):502–515. https://doi.org/10.1016/j.stem.2016.06.011
Tyser RCV, Mahammadov E, Nakanoh S, et al., 2021. Single-cell transcriptomic characterization of a gastrulating human embryo. Nature, 600(7888):285–289. https://doi.org/10.1038/s41586-021-04158-y
Yang Y, Liu B, Xu J, et al., 2017. Derivation of pluripotent stem cells with in vivo embryonic and extraembryonic potency. Cell, 169(2):243–257.e25. https://doi.org/10.1016/j.cell.2017.02.005
Yu LQ, Wei YL, Duan JL, et al., 2021. Blastocyst-like structures generated from human pluripotent stem cells. Nature, 591(7851):620–626. https://doi.org/10.1038/s41586-021-03356-y
Acknowledgments
This work was supported by the National Key R&D Program of China (Nos. 2021YFA0805700 and 2021YFA1102000), the National Natural Science Foundation of China (No. U2102204), and the Natural Science Foundation of Yunnan Province, China (Nos. 202001BC070001 and 202102AA100053).
Author information
Authors and Affiliations
Contributions
Yuyu NIU and Chuanxin CHEN conceived the structure of manuscript. Yuyu NIU and Shuang LI designed the experiments. Shuang LI performed the experiments and analyzed data. Zhenzhen CHEN prepared the cell lines. Chuanxin CHEN and Shuang LI wrote the context and generated the figures. Yuyu NIU, Chuanxin CHEN, and Shuang LI contributed to revision of the manuscript. Bioinformatics analysis was supported by SequMed BioTechnology Inc. (Guangzhou, China). All authors have read and approved the final manuscript, and therefore, have full access to all the data in the study and take responsibility for the integrity and security of the data.
Corresponding authors
Ethics declarations
Shuang LI, Zhenzhen CHEN, Chuanxin CHEN, and Yuyu NIU declare that they have no conflict of interest.
This article does not contain any studies with human or animal subjects performed by any of the authors.
Additional information
Materials and methods
Detailed methods are provided in the electronic supplementary materials of this paper.
Supplementary information
Materials and methods; Figs. S1 and S2
Electronic supplementary material
Rights and permissions
About this article
Cite this article
Li, S., Chen, Z., Chen, C. et al. Multiplexed single-cell transcriptome analysis reveals molecular characteristics of monkey pluripotent stem cell lines. J. Zhejiang Univ. Sci. B 24, 418–429 (2023). https://doi.org/10.1631/jzus.B2200440
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1631/jzus.B2200440
Key words
- Monkey pluripotent stem cell
- Multiplexed single-cell sequencing
- Naive pluripotency
- Extended pluripotent stem cells