Generation of embryonic stem cells derived from the inner cell mass of blastocysts of outbred ICR mice

Han, Ning; Baek, Song Ee; Kim, Hwa-Young; Lee, Kwon Young; Yun, Jong-Il; Choi, Jung Hoon; Lee, Eunsong; Park, Choon‐Keun; Lee, Seung Tae

doi:10.1080/19768354.2020.1752306

Cited by 6 publications

(4 citation statements)

References 36 publications

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“…5B ). The outbred ICR line has been considered a non-permissive strain to derive ESCs ( Lee et al, 2012 ; Han et al, 2020 ). Using 2iLA medium, we efficiently derived ESC lines from E3.5–4.5 ICR blastocysts.…”

Section: Resultsmentioning

confidence: 99%

Dual role of lipids for genome stability and pluripotency facilitates full potency of mouse embryonic stem cells

et al. 2023

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While Mek1/2 and Gsk3β inhibition (“2i”) supports the maintenance of murine embryonic stem cells (ESCs) in a homogenous naïve state, prolonged culture in 2i results in aneuploidy and DNA hypomethylation that impairs developmental potential. Additionally, 2i fails to support derivation and culture of fully potent female ESCs. Here we find that mouse ESCs cultured in 2i/LIF supplemented with lipid-rich albumin (AlbuMAX) undergo pluripotency transition yet maintain genomic stability and full potency over long-term culture. Mechanistically, lipids in AlbuMAX impact intracellular metabolism including nucleotide biosynthesis, lipid biogenesis, and TCA cycle intermediates, with enhanced expression of DNMT3s that prevent DNA hypomethylation. Lipids induce a formative-like pluripotent state through direct stimulation of Erk2 phosphorylation, which also alleviates X chromosome loss in female ESCs. Importantly, both male and female “all-ESC” mice can be generated from de novo derived ESCs using AlbuMAX-based media. Our findings underscore the importance of lipids to pluripotency and link nutrient cues to genome integrity in early development.

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Section: Resultsmentioning

confidence: 99%

Dual role of lipids for genome stability and pluripotency facilitates full potency of mouse embryonic stem cells

et al. 2023

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“…Formative pluripotency was recently proposed to describe an intermediate executive phase between the naïve and primed states [9–12, 21]. This phase of pluripotency is thought to be responsive to inductive cues for lineage differentiation and to express a group of unique formative genes [11, 43]. We performed RNA-seq of 2i-ESCs and AX-ESCs, finding 1637 genes that were differentially expressed and which are significantly enriched in pluripotency genes and the MAPK signaling pathway ( Extended data Fig.…”

Section: Resultsmentioning

confidence: 99%

“…7b ). The outbred ICR line has been considered a non-permissive strain to derive ESCs [43, 59]. Using 2iLA medium, we efficiently derived ICR ESC lines from E3.5-E4.5 blastocysts.…”

Section: Resultsmentioning

confidence: 99%

Lipids Maintain Genomic Stability and Developmental Potency of Murine Pluripotent Stem Cells

Wen

Zhong

Gordillo

et al. 2022

Preprint

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Lipids play vital roles in cellular homeostasis and regulate pluripotency of human stem cells. However, the impact of lipids on murine pluripotent stem cells is unclear. While Mek and Gsk3 inhibition supports the maintenance of murine embryonic stem cells in a homogenous naive state, prolonged culture in 2i results in aneuploidy and DNA hypomethylation that impairs developmental potential. Additionally, 2i fails to support derivation and culture of fully potent female ESCs. Here we find that mouse ESCs cultured in 2i/LIF supplemented with lipid-rich albumin (AlbuMAX) undergo pluripotency transition yet maintain genomic stability and full potency over long-term culture. Mechanistically, lipids in AlbuMAX impact intracellular metabolism including nucleotide biosynthesis, lipid biogenesis, and TCA cycle intermediates, with enhanced expression of ZCAN4 and DNMT3s that prevent telomere shortening and DNA hypomethylation. In concert with 2i, lipids induce a formative-like pluripotent state through direct stimulation of Mek-mediated Erk2 phosphorylation, which also alleviates X chromosome loss in female ESCs. Importantly, both male and female all-ESC mice can be generated from de novo derived ESCs using AlbuMAX-based media. Our findings underscore the importance of lipids to pluripotency and link nutrient cues to genome integrity in early development.

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“…Because cell immortalization is a critical process for carcinogenesis, numerous immortalized cell lines have been isolated from various human cancers. Different from mouse embryonic stem (ES) cells (Han et al 2020 ), normal diploid human cells eventually enter replicative senescence after a finite number of cell divisions (Hayflick 1965 ) and thus, to become immortalized, cells must escape replicative senescence, acquiring the capability of infinite cell division. Replicative senescence is attributable to shortening of telomeres, the ends of chromosomes, with each cell division (Blackburn 2000 ).…”

Section: Introductionmentioning

confidence: 99%

Immortalization of primary marmoset skin fibroblasts by CRISPR-Cas9-mediated gene targeting

Jeong

Cho

Kwak

et al. 2022

Animal Cells and Systems

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Immortalized cell lines can be used for diverse in vitro experiments, providing invaluable data before conducting in vivo studies Callithrix jacchus , the common marmoset, is a non-human primate model utilized for studying various human diseases. However, only a few immortalized marmoset cell lines are currently available. In the present study, we reveal that CRISPR-Cas9-mediated targeting of the p53 gene or CDKN2A locus is an effective means for immortalizing primary marmoset skin fibroblasts. In addition to frameshift mutations that result in premature stop codons, in-frame mutations potentially destroying the DNA-binding motif of p53 are frequently detected in immortalized cells. Like Cdkn2a -deficient mouse cells, CDKN2A -deficient marmoset cells express wild-type p53 proteins normally respond to genotoxic stresses, including adriamycin and etoposide. Taken together, these findings indicate that Cas9- mediated gene targeting of the p53 gene or CDKN2A locus is an effective tool for establishing immortalized marmoset cell lines with defined genetic alterations.

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Generation of embryonic stem cells derived from the inner cell mass of blastocysts of outbred ICR mice

Cited by 6 publications

References 36 publications

Dual role of lipids for genome stability and pluripotency facilitates full potency of mouse embryonic stem cells

Dual role of lipids for genome stability and pluripotency facilitates full potency of mouse embryonic stem cells

Lipids Maintain Genomic Stability and Developmental Potency of Murine Pluripotent Stem Cells

Immortalization of primary marmoset skin fibroblasts by CRISPR-Cas9-mediated gene targeting

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