Purification of human iPSC-derived cells at large scale using microRNA switch and magnetic-activated cell sorting

Tsujisaka, Yuta; Hatani, Takeshi; Okubo, Chikako; Itô, Ryo; Kimura, Azuma; Narita, Megumi; Chonabayashi, Kazuhisa; Lucena-Cacace, Antonio; Toyoda, Tarō; Osafune, Kenji; Kimura, Takeshi; Saito, Hirohide; Yoshida, Yoshinori

doi:10.1016/j.stemcr.2022.05.003

Cited by 12 publications

(4 citation statements)

References 39 publications

(54 reference statements)

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“…Differentiation of iPSCs and ESCs. iPSCs/ESCs were directed to differentiate into pancreatic lineage cells as previously described 38,39 . The medium composition was as follows:…”

Section: Methodsmentioning

confidence: 99%

Elucidation of HHEX in pancreatic endoderm differentiation using a human iPSC differentiation model

Ito

Kimura

Hirose

et al. 2023

Sci Rep

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For pluripotent stem cell (PSC)-based regenerative therapy against diabetes, the differentiation efficiency to pancreatic lineage cells needs to be improved based on the mechanistic understanding of pancreatic differentiation. Here, we aimed to elucidate the molecular mechanisms underlying pancreatic endoderm differentiation by searching for factors that regulate a crucial pancreatic endoderm marker gene, NKX6.1. Unbiasedly screening an siRNA knockdown library, we identified a candidate transcription factor, HHEX. HHEX knockdown suppressed the expression of another pancreatic endoderm marker gene, PTF1A, as well as NKX6.1, independently of PDX1, a known regulator of NKX6.1 expression. In contrast, the overexpression of HHEX upregulated the expressions of NKX6.1 and PTF1A. RNA-seq analysis showed decreased expressions of several genes related to pancreatic development, such as NKX6.1, PTF1A, ONECUT1 and ONECUT3, in HHEX knockdown pancreatic endoderm. These results suggest that HHEX plays a key role in pancreatic endoderm differentiation.

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“…Differentiation of iPSCs and ESCs. iPSCs/ESCs were directed to differentiate into pancreatic lineage cells as previously described 38,39 . The medium composition was as follows:…”

Section: Methodsmentioning

confidence: 99%

Elucidation of HHEX in pancreatic endoderm differentiation using a human iPSC differentiation model

Ito

Kimura

Hirose

et al. 2023

Sci Rep

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“…Recently, microRNA-responsive, synthetic modified mRNA switch (miR switch), in which the synthetically modified mRNA was designed to contain the miRNA target sequence complimentary to specific miRNA and fluorescent reporter sequence, was used to purify hPSC-derived cardiomyocytes, endothelial cells, hepatocytes, and insulin-producing cells from undifferentiated hPSCs and other differentiated cells ( Miki et al, 2015 ). Interestingly, large-scale purification of hiPSC-derived cells such as cardiomyocytes and pancreatic insulin-producing cells has been accomplished by a combination of the miR switch and magnetic-activating cell sorting (MACS), the so-called miR-switch-MACS ( Tsujisaka et al, 2022 ). Furthermore, if one unique miRNA cannot specifically identify the cell type of interest, the more robust miRNA ON and OFF switch has been reported to enable the use of multiple miRNAs for precisely purifying such cells ( Fujita et al, 2022 ).…”

Section: Challenges and Future Perspectivesmentioning

confidence: 99%

iPSC-based approach for human hair follicle regeneration

Vatanashevanopakorn

Sartyoungkul

2023

Front. Cell Dev. Biol.

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Hair follicles (HFs) are a multifunctional structure involved in physical protection, thermoregulation, sensational detection, and wound healing. Formation and cycling of HFs require dynamic interaction between different cell types of the follicles. Although the processes have been well studied, the generation of human functional HFs with a normal cycling pattern for clinical utilization has yet to be achieved. Recently, human pluripotent stem cells (hPSCs) serve as an unlimited cell source for generating various types of cells including cells of the HFs. In this review, HF morphogenesis and cycling, different cell sources used for HF regeneration, and potential strategies for HF bioengineering using induced pluripotent stem cells (iPSCs) are depicted. Challenges and perspectives toward the therapeutic use of bioengineered HFs for hair loss disorder are also discussed.

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“…The methodology of magnetic activated cell sorting (MACS) was published first in 1995 by Pesce and De Felici [10], to separate primordial germ cells of mouse embryos from somatic cells. It is commonly used in immunology, oncological research, neuroscience and stem cell research [11][12][13][14][15][16]. Dunlap et al [17] used the method to identify immunoglobulin (Ig) G-bound bacteria from bronchoalveolar lavage samples to study lung microbiome.…”

Section: Introductionmentioning

confidence: 99%

An Alternative Application of Magnetic-Activated Cell Sorting: CD45 and CD235a Based Purification of Semen and Testicular Tissue Samples

Czétány,

Balló,

Márk

et al. 2024

IJMS

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Magnetic activated cell sorting (MACS) is a well-known sperm selection technique, which is able to remove apoptotic spermatozoa from semen samples using the classic annexinV based method. Leukocytes and erythrocytes in semen samples or in testicular tissue processed for in vitro fertilization (IVF) could exert detrimental effects on sperm. In the current study, we rethought the aforementioned technique and used magnetic microbeads conjugated with anti-CD45/CD235a antibodies to eliminate contaminating leukocytes and erythrocytes from leukocytospermic semen samples and testicular tissue samples gained via testicular sperm extraction (TESE). With this technique, a 15.7- and a 30.8-fold reduction could be achieved in the ratio of leukocytes in semen and in the number of erythrocytes in TESE samples, respectively. Our results show that MACS is a method worth to reconsider, with more potential alternative applications. Investigations to find molecules labeling high-quality sperm population and the development of positive selection procedures based on these might be a direction of future research.

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Purification of human iPSC-derived cells at large scale using microRNA switch and magnetic-activated cell sorting

Cited by 12 publications

References 39 publications

Elucidation of HHEX in pancreatic endoderm differentiation using a human iPSC differentiation model

Elucidation of HHEX in pancreatic endoderm differentiation using a human iPSC differentiation model

iPSC-based approach for human hair follicle regeneration

An Alternative Application of Magnetic-Activated Cell Sorting: CD45 and CD235a Based Purification of Semen and Testicular Tissue Samples

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