A Passage-free, Simplified, and Scalable Novel Method for IPSC Generation in Three-dimensional Culture

Overview

Journal Regen Ther

Date 2024 Mar 18

PMID 38496009

Authors

Masaya Tsukamoto

Tomoyuki Kawasaki

Mohan C Vemuri

Akihiro Umezawa

Hidenori Akutsu

Affiliations

Soon will be listed here.

Abstract

Induced pluripotent stem cells (iPSCs) have immense potential for use in disease modeling, etiological studies, and drug discovery. However, the current workflow for iPSC generation and maintenance poses challenges particularly during the establishment phase when specialized skills are required. Although three-dimensional culture systems offer scalability for maintaining established iPSCs, the enzymatic dissociation step is complex and time-consuming. In this study, a novel approach was developed to address these challenges by enabling iPSC generation, maintenance, and differentiation without the need for two-dimensional culture or enzymatic dissociation. This streamlined method offers a more convenient workflow, reduces variability and labor for technicians, and opens up avenues for advancements in iPSC research and broader applications.

Citing Articles

Synthetic hydrogel substrate for human induced pluripotent stem cell definitive endoderm differentiation.

Mulero-Russe A, Mora-Boza A, Marquez E, Ziegelski M, Helmrath M, Garcia A Biomaterials. 2024; 315:122920.

PMID: 39504708 PMC: 11625597. DOI: 10.1016/j.biomaterials.2024.122920.

Alternative Method for Obtaining Human-Induced Pluripotent Stem Cell Lines and Three-Dimensional Growth: A Simplified, Passage-Free Approach that Minimizes Labor.

Tsukamoto M, Kawasaki T, Umezawa A, Akutsu H Bio Protoc. 2024; 14(19):e5081.

PMID: 39399592 PMC: 11470374. DOI: 10.21769/BioProtoc.5081.

References

Nakagawa M, Taniguchi Y, Senda S, Takizawa N, Ichisaka T, Asano K . A novel efficient feeder-free culture system for the derivation of human induced pluripotent stem cells. Sci Rep. 2014; 4:3594. PMC: 3884228. DOI: 10.1038/srep03594. View

Fattahi P, Rahimian A, Slama M, Gwon K, Gonzalez-Suarez A, Wolf J . Core-shell hydrogel microcapsules enable formation of human pluripotent stem cell spheroids and their cultivation in a stirred bioreactor. Sci Rep. 2021; 11(1):7177. PMC: 8010084. DOI: 10.1038/s41598-021-85786-2. View

Nath S, Horie M, Nagamori E, Kino-Oka M . Size- and time-dependent growth properties of human induced pluripotent stem cells in the culture of single aggregate. J Biosci Bioeng. 2017; 124(4):469-475. DOI: 10.1016/j.jbiosc.2017.05.006. View

Bai Q, Ramirez J, Becker F, Pantesco V, Lavabre-Bertrand T, Hovatta O . Temporal analysis of genome alterations induced by single-cell passaging in human embryonic stem cells. Stem Cells Dev. 2014; 24(5):653-62. PMC: 4333508. DOI: 10.1089/scd.2014.0292. View

Morita K, Nakamura A, Machida M, Kawasaki T, Nakanishi R, Ichida J . Efficient reprogramming of human fibroblasts using RNA reprogramming with DAPT and iDOT1L under normoxia conditions. Regen Ther. 2022; 21:389-397. PMC: 9493288. DOI: 10.1016/j.reth.2022.09.002. View

Assou S, Girault N, Plinet M, Bouckenheimer J, Sansac C, Combe M . Recurrent Genetic Abnormalities in Human Pluripotent Stem Cells: Definition and Routine Detection in Culture Supernatant by Targeted Droplet Digital PCR. Stem Cell Reports. 2020; 14(1):1-8. PMC: 6962701. DOI: 10.1016/j.stemcr.2019.12.004. View

Galvanauskas V, Grincas V, Simutis R, Kagawa Y, Kino-Oka M . Current state and perspectives in modeling and control of human pluripotent stem cell expansion processes in stirred-tank bioreactors. Biotechnol Prog. 2016; 33(2):355-364. DOI: 10.1002/btpr.2431. View

Huang C, Liu C, Ting C, Chiu Y, Cheng Y, Nicholson M . Human iPSC banking: barriers and opportunities. J Biomed Sci. 2019; 26(1):87. PMC: 6819403. DOI: 10.1186/s12929-019-0578-x. View

Castro-Vinuelas R, Sanjurjo-Rodriguez C, Pineiro-Ramil M, Rodriguez-Fernandez S, Lopez-Baltar I, Fuentes-Boquete I . Tips and tricks for successfully culturing and adapting human induced pluripotent stem cells. Mol Ther Methods Clin Dev. 2021; 23:569-581. PMC: 8640166. DOI: 10.1016/j.omtm.2021.10.013. View

10.

Hookway T, Butts J, Lee E, Tang H, McDevitt T . Aggregate formation and suspension culture of human pluripotent stem cells and differentiated progeny. Methods. 2015; 101:11-20. DOI: 10.1016/j.ymeth.2015.11.027. View

11.

Wei H, Wang C, Guo R, Takahashi K, Naruse K . Development of a model of ischemic heart disease using cardiomyocytes differentiated from human induced pluripotent stem cells. Biochem Biophys Res Commun. 2019; 520(3):600-605. DOI: 10.1016/j.bbrc.2019.09.119. View

12.

Hansen S, Stummann T, Borland H, Hasholt L, Tumer Z, Nielsen J . Induced pluripotent stem cell - derived neurons for the study of spinocerebellar ataxia type 3. Stem Cell Res. 2016; 17(2):306-317. DOI: 10.1016/j.scr.2016.07.004. View

13.

Tsankov A, Akopian V, Pop R, Chetty S, Gifford C, Daheron L . A qPCR ScoreCard quantifies the differentiation potential of human pluripotent stem cells. Nat Biotechnol. 2015; 33(11):1182-92. PMC: 4636964. DOI: 10.1038/nbt.3387. View

14.

Lin Z, Garbern J, Liu R, Li Q, Mancheno Juncosa E, Elwell H . Tissue-embedded stretchable nanoelectronics reveal endothelial cell-mediated electrical maturation of human 3D cardiac microtissues. Sci Adv. 2023; 9(10):eade8513. PMC: 9995081. DOI: 10.1126/sciadv.ade8513. View

15.

Nagasaka R, Matsumoto M, Okada M, Sasaki H, Kanie K, Kii H . Visualization of morphological categories of colonies for monitoring of effect on induced pluripotent stem cell culture status. Regen Ther. 2018; 6:41-51. PMC: 6134894. DOI: 10.1016/j.reth.2016.12.003. View

16.

Wiegand C, Banerjee I . Recent advances in the applications of iPSC technology. Curr Opin Biotechnol. 2019; 60:250-258. DOI: 10.1016/j.copbio.2019.05.011. View

17.

Lei Y, Schaffer D . A fully defined and scalable 3D culture system for human pluripotent stem cell expansion and differentiation. Proc Natl Acad Sci U S A. 2013; 110(52):E5039-48. PMC: 3876251. DOI: 10.1073/pnas.1309408110. View

18.

Isono W, Kawasaki T, Ichida J, Ayabe T, Hiraike O, Umezawa A . The combination of dibenzazepine and a DOT1L inhibitor enables a stable maintenance of human naïve-state pluripotency in non-hypoxic conditions. Regen Ther. 2021; 15:161-168. PMC: 7770342. DOI: 10.1016/j.reth.2020.08.001. View