Suspension Culture of Undifferentiated Human Embryonic and Induced Pluripotent Stem Cells

Amit, Michal; Chebath, Judith; Margulets, Victoria; Laevsky, Ilana; Miropolsky, Yael; Shariki, Kohava; Peri, Meital; Blais, Idit; Slutsky, Guy; Revel, Michel; Itskovitz‐Eldor, Joseph

doi:10.1007/s12015-010-9149-y

Cited by 144 publications

(78 citation statements)

References 38 publications

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“…8 A classic cytogenetic protocol (G banding) is considered as the default approach for routine karyotypic assessment, 9,10 and is the method widely used to value chromosome integrity. 11 Karyotypic abnormalities of hESCs in long-term cultures have been reported before. [12][13][14][15] In hESCs, most of the alteration have been attributed to chromosomes 12 and 17, but chromosomes X and 20 were also repeatedly described.…”

Section: Introductionmentioning

confidence: 83%

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

et al. 2012

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Pluripotency and proliferative capacity of human embryonic stem cells (hESCs) make them a promising source for basic and applied research as well as in therapeutic medicine. The introduction of human induced pluripotent cells (hiPSCs) holds great promise for patient-tailored regenerative medicine therapies. However, for hESCs and hiPSCs to be applied for therapeutic purposes, long-term genomic stability in culture must be maintained. Until recently, G-banding analysis was considered as the default approach for detecting chromosomal abnormalities in stem cells. Our goal in this study was to apply fluorescence in-situ hybridization (FISH) and comparative genomic hybridization (CGH) for the screening of pluripotent stem cells, which will enable us identifying chromosomal abnormalities in stem cells genome with a better resolution. We studied three hESC lines and two hiPSC lines over long-term culture. Aneuploidy rates were evaluated at different passages, using FISH probes (12,13,16,17,18,21,X,Y). Genomic integrity was shown to be maintained at early passages of hESCs and hiPSCs but, at late passages, we observed low rates mosaiciam in hESCs, which implies a direct correlation between number of passages and increased aneuploidy rate. In addition, CGH analysis revealed a recurrent genomic instability, involving the gain of chromosome 1q. This finding was detected in two unrelated cell lines of different origin and implies that gains of chromosome 1q may endow a clonal advantage in culture. These findings, which could only partially be detected by conventional cytogenetic methods, emphasize the importance of using molecular cytogenetic methods for tracking genomic instability in stem cells.

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Section: Introductionmentioning

confidence: 83%

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

et al. 2012

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“…Another approach to producing hPSCs aggregates or clusters is to use soluble factors instead of biomaterials in a suspension culture [18][19][20][21]. However, karyotypic abnormalities were observed in some cell lines in the suspension culture [20].…”

mentioning

confidence: 99%

The Use of Nanofibrillar Cellulose Hydrogel As a Flexible Three-Dimensional Model to Culture Human Pluripotent Stem Cells

Lou

Kanninen

Kuisma

et al. 2014

Stem Cells and Development

195

161

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Human embryonic stem cells and induced pluripotent stem cells have great potential in research and therapies. The current in vitro culture systems for human pluripotent stem cells (hPSCs) do not mimic the threedimensional (3D) in vivo stem cell niche that transiently supports stem cell proliferation and is subject to changes which facilitate subsequent differentiation during development. Here, we demonstrate, for the first time, that a novel plant-derived nanofibrillar cellulose (NFC) hydrogel creates a flexible 3D environment for hPSC culture. The pluripotency of hPSCs cultured in the NFC hydrogel was maintained for 26 days as evidenced by the expression of OCT4, NANOG, and SSEA-4, in vitro embryoid body formation and in vivo teratoma formation. The use of a cellulose enzyme, cellulase, enables easy cell propagation in 3D culture as well as a shift between 3D and two-dimensional cultures. More importantly, the removal of the NFC hydrogel facilitates differentiation while retaining 3D cell organization. Thus, the NFC hydrogel represents a flexible, xeno-free 3D culture system that supports pluripotency and will be useful in hPSC-based drug research and regenerative medicine.

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“…Suspension SCs like hematopoietic progenitor cells without requirement of surface attachment for growth were successfully cultured in stirring bioreactors. Adherent SCs, such as ESCs, iPSCs and MSCs, are attached to cell microcarriers for their suspension and are cultured in stirring bioreactors [19,20]. The growth efficiency and homogeneous of cultured SCs in stirring bioreactor were improved in somehow, due to homogeneous nature of culture system.…”

Section: Stirred Tank Bioreactormentioning

confidence: 99%

Application of Bioreactor in Stem Cell Culture

Zhang¹,

Wang²,

Pong³

et al. 2017

JBiSE

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Stem cells (SCs), the undifferentiated biological cells, have the infinite capacity to self-renew and the pluripotent ability to differentiate. SCs and their derived products offer great promise for biomedical applications such as cell therapy, tissue engineering, regenerative medicine and drug screening. However, the clinical applications of SCs require a large amount of SCs with high quality and the number of SCs from their tissue resources is very limited. Large-scale expansion is needed to generate homogeneous SCs with good biological characteristics for clinical application. This necessitates a bioreactor system to provide controllable and stable conditions for stem cell (SC) culture. Traditional methods of bioreactor for maintenance and expansion of cells rely on two-dimensional (2-D) culture techniques, leading to loss self-renewal ability and differentiation capacity upon long-term culture. New approaches for SC expansion with bioreactor employ three-dimensional (3-D) cell growth to mimic their environment in vivo. In this review, we summarize the application of bioreactors in SC culture.

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Suspension Culture of Undifferentiated Human Embryonic and Induced Pluripotent Stem Cells

Cited by 144 publications

References 38 publications

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

The Use of Nanofibrillar Cellulose Hydrogel As a Flexible Three-Dimensional Model to Culture Human Pluripotent Stem Cells

Application of Bioreactor in Stem Cell Culture

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