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Project description:We systematically screened novel culture systems designed for naïve human embryonic stem cells via high-content screening and small molecule combinations. Our findings indicate that the cell lines established through this innovative system closely mimic the in vivo Inner Cell Mass (ICM) concerning both transcriptome profiles and methylation patterns. Additionally, we observed an elevated level of methylation specifically on transposable elements within these cell lines. Furthermore, our investigations suggest that the genomic stability of this novel system may exhibit superior qualities.
2025-12-01 | GSE252313 | GEO
Project description:We systematically screened novel culture systems designed for naïve human embryonic stem cells via high-content screening and small molecule combinations. Our findings indicate that the cell lines established through this innovative system closely mimic the in vivo Inner Cell Mass (ICM) concerning both transcriptome profiles and methylation patterns. Additionally, we observed an elevated level of methylation specifically on transposable elements within these cell lines. Furthermore, our investigations suggest that the genomic stability of this novel system may exhibit superior qualities.
2025-12-01 | GSE252312 | GEO
Project description:Novel culture systems for naive human embryonic stem cells based on chemical molecule screening
| PRJNA1059630 | ENA
Project description:Novel culture systems for naive human embryonic stem cells based on chemical molecule screening [WGBS]
| PRJNA1059629 | ENA
Project description:Novel culture systems for naive human embryonic stem cells based on chemical molecule screening [RNA-seq]
| PRJNA1059627 | ENA
Project description:Mouse embryonic stem cell (ESC) lines, and more recently human ESC lines, have become valuable tools for studying early mammalian development. Increasing interest in ESCs and their differentiated progeny in drug discovery and as potential therapeutic agents has highlighted the fact that current two-dimensional (2D) static culturing techniques are inadequate for large-scale production. The culture of mammalian cells in three-dimensional (3D) agitated systems has been shown to overcome many of the restrictions of 2D and is therefore likely to be effective for ESC proliferation. Using murine ESCs as our initial model, we investigated the effectiveness of different 3D culture environments for the expansion of pluripotent ESCs. Solohill Collagen, Solohill FACT, and Cultispher-S microcarriers were employed and used in conjunction with stirred bioreactors. Initial seeding parameters, including cell number and agitation conditions, were found to be critical in promoting attachment to microcarriers and minimizing the size of aggregates formed. While all microcarriers supported the growth of undifferentiated mESCs, Cultispher-S out-performed the Solohill microcarriers. When cultured for successive passages on Cultispher-S microcarriers, mESCs maintained their pluripotency, demonstrated by self-renewal, expression of pluripotency markers and the ability to undergo multi-lineage differentiation. When these optimized conditions were applied to unweaned human ESCs, Cultispher-S microcarriers supported the growth of hESCs that retained expression of pluripotency markers including SSEA4, Tra-1-60, NANOG, and OCT-4. Our study highlights the importance of optimization of initial seeding parameters and provides proof-of-concept data demonstrating the utility of microcarriers and bioreactors for the expansion of hESCs.
| S-EPMC3580883 | biostudies-other
Project description:The naïve pluripotent state has been shown in mice to lead to broad and more robust developmental potential relative to primed mouse epiblast cells. The human naïve ES cell state has eluded derivation without the use of transgenes, and forced expression of OCT4, KLF4, and KLF2 allows maintenance of human cells in a naïve state [Hanna J, et al. (2010) Proc Natl Acad Sci USA 107(20):9222-9227]. We describe two routes to generate nontransgenic naïve human ES cells (hESCs). The first is by reverse toggling of preexisting primed hESC lines by preculture in the histone deacetylase inhibitors butyrate and suberoylanilide hydroxamic acid, followed by culture in MEK/ERK and GSK3 inhibitors (2i) with FGF2. The second route is by direct derivation from a human embryo in 2i with FGF2. We show that human naïve cells meet mouse criteria for the naïve state by growth characteristics, antibody labeling profile, gene expression, X-inactivation profile, mitochondrial morphology, microRNA profile and development in the context of teratomas. hESCs can exist in a naïve state without the need for transgenes. Direct derivation is an elusive, but attainable, process, leading to cells at the earliest stage of in vitro pluripotency described for humans. Reverse toggling of primed cells to naïve is efficient and reproducible.
| S-EPMC3970494 | biostudies-literature
Project description:Efforts to develop more effective therapies for acute leukemia may benefit from high-throughput screening systems that reflect the complex physiology of the disease, including leukemia stem cells (LSCs) and supportive interactions with the bone-marrow microenvironment. The therapeutic targeting of LSCs is challenging because LSCs are highly similar to normal hematopoietic stem and progenitor cells (HSPCs) and are protected by stromal cells in vivo. We screened 14,718 compounds in a leukemia-stroma co-culture system for inhibition of cobblestone formation, a cellular behavior associated with stem-cell function. Among those that inhibited malignant cells but spared HSPCs was the cholesterol-lowering drug lovastatin. Lovastatin showed anti-LSC activity in vitro and in an in vivo bone marrow transplantation model. Mechanistic studies demonstrated that the effect was on-target, via inhibition of HMG-CoA reductase. These results suggest that statins should be re-evaluated as anti-leukemia agents, and illustrate the power of merging physiologically-relevant models with high-throughput screening. Freshly isolated LSCe cells were treated with either 5M-BM-5M BRD7116 or DMSO vehicle control for 6 hours in suspension in IMDM (12440053, Invitrogen), 10% FBS. Total RNA was extracted from the cells, labeled, and hybridized to MouseRef-8 v2.0 Expression BeadChips. Hartwell, K.A. et al., In Press (full citation forthcoming)
2013-09-20 | E-GEOD-51033 | biostudies-arrayexpress
Project description:The development of new high-tech systems for screening anticancer drugs is one of the main problems of preclinical screening. Poor correlation between preclinical in vitro and in vivo data with clinical trials remains a major concern. The choice of the correct tumor model at the stage of in vitro testing provides reduction in both financial and time costs during later stages due to the timely screening of ineffective agents. In view of the growing incidence of oncology, increasing the pace of the creation, development and testing of new antitumor agents, the improvement and expansion of new high-tech systems for preclinical in vitro screening is becoming very important. The pharmaceutical industry presently relies on several widely used in vitro models, including two-dimensional models, three-dimensional models, microfluidic systems, Boyden's chamber and models created using 3D bioprinting. This review outlines and describes these tumor models including their use in research, in addition to their characteristics. This review therefore gives an insight into in vitro based testing which is of interest to researchers and clinicians from differing fields including pharmacy, preclinical studies and cell biology.
| S-EPMC7160228 | biostudies-literature
Project description:ObjectiveHuman embryonic stem cells (hESCs) have the potential to give rise to all types of cells in the human body when appropriately induced to differentiate. Stem cells can differentiate spontaneously into the three-germ layer derivatives by embryoid bodies (EBs) formation. However, the two-dimensional (2D) adherent culture of hESCs under defined conditions is commonly used for directed differentiation toward a specific type of mature cells. In this study, we aimed to determine the propensity of the Royan hESC lines based on comparison of expression levels of 46 lineage specific markers.Materials and methodsIn this experimental study, we have compared the expression of lineage-specific markers in hESC lines during EB versus adherent-based spontaneous differentiation. We used quantitative real-time polymerase chain reaction (qRT-PCR) to assess expressions of 46 lineage-specific markers in 4 hESC lines, Royan H1 (RH1), RH2, RH5, and RH6, during spontaneous differentiation in both EB and adherent cultures at 0, 10, and 30 days after initiation of differentiation.ResultsBased on qRT-PCR data analysis, the liver and neuronal markers had higher expression levels in EBs, whereas skin-specific markers expressed at higher levels in the adherent culture. The results showed differential expression patterns of some lineage-specific markers in EBs compared with the adherent cultures.ConclusionAccording to these results, possibly the spontaneous differentiation technique could be a useful method for optimization of culture conditions to differentiate stem cells into specific cell types such ectoderm, neuron, endoderm and hepatocyte. This approach might prove beneficial for further work on maximizing the efficiency of directed differentiation and development of novel differentiation protocols.
| S-EPMC6582416 | biostudies-literature