3D ECM-rich environment sustains the identity of naive human iPSCs

Cesare, Elisa; Urciuolo, Anna; Stuart, Hannah; Torchio, Erika; Gesualdo, Alessia; Laterza, Cecilia; Gagliano, Onelia; Martewicz, Sebastian; Cui, Meihua; Manfredi, Anna; Filippo, Lucio Di; Sabatelli, Patrizia; Squarzoni, Stefano; Zorzan, Irene; Betto, Riccardo Massimiliano; Martello, Graziano; Cacchiarelli, Davide; Luni, Camilla; Elvassore, Nicola

doi:10.1016/j.stem.2022.11.011

Cited by 13 publications

(2 citation statements)

References 65 publications

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“…This agrees with the compacted morphology of the colonies (Fig. 2a ) and with a recent study revealing that extracellular matrix sustains naïve identity of hniPSCs 37 . Conversely, developmental and stem cell differentiation related genes ( PECAM1 , KIT , HAND1 , VASH1 , CDX2 and ADM ) were downregulated in CI-hnESCs (Fig.…”

Section: Resultssupporting

confidence: 92%

Self-renewing human naïve pluripotent stem cells dedifferentiate in 3D culture and form blastoids spontaneously

Guo,

Wu,

Chen

et al. 2024

Nat Commun

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Human naïve pluripotent stem cells (hnPSCs) can generate integrated models of blastocysts termed blastoids upon switch to inductive medium. However, the underlying mechanisms remain obscure. Here we report that self-renewing hnPSCs spontaneously and efficiently give rise to blastoids upon three dimensional (3D) suspension culture. The spontaneous blastoids mimic early stage human blastocysts in terms of structure, size, and transcriptome characteristics and are capable of progressing to post-implantation stages. This property is conferred by the glycogen synthase kinase-3 (GSK3) signalling inhibitor IM-12 present in 5iLAF self-renewing medium. IM-12 upregulates oxidative phosphorylation-associated genes that underly the capacity of hnPSCs to generate blastoids spontaneously. Starting from day one of self-organization, hnPSCs at the boundary of all 3D aggregates dedifferentiate into E5 embryo-like intermediates. Intermediates co-express SOX2/OCT4 and GATA6 and by day 3 specify trophoblast fate, which coincides with cavity and blastoid formation. In summary, spontaneous blastoid formation results from 3D culture triggering dedifferentiation of hnPSCs into earlier embryo-like intermediates which are then competent to segregate blastocyst fates.

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Section: Resultssupporting

confidence: 92%

Self-renewing human naïve pluripotent stem cells dedifferentiate in 3D culture and form blastoids spontaneously

Guo,

Wu,

Chen

et al. 2024

Nat Commun

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“…Interestingly, the development of 3D culture systems that support robust long-term and feeder-free self-renewal of pluripotent cells has been recently described and demonstrated to boost and stabilize high plasticity [15,25,41,42]. In line with these findings, encapsulation of miR-200-reprogrammed cells in PTFE micro-bioreactors coax cells to assemble in multicellular 3D spheroids reminiscent of ICM-like morphology and steadily transcribing for the main pluripotency-related genes.…”

Section: Discussionmentioning

confidence: 79%

Generation of Artificial Blastoids Combining miR-200-Mediated Reprogramming and Mechanical Cues

Pennarossa,

Arcuri,

Gandolfi

et al. 2024

Cells

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In vitro-generated blastocyst-like structures are of great importance since they recapitulate specific features or processes of early embryogenesis, thus avoiding ethical concerns as well as increasing scalability and accessibility compared to the use of natural embryos. Here, we combine cell reprogramming and mechanical stimuli to create 3D spherical aggregates that are phenotypically similar to those of natural embryos. Specifically, dermal fibroblasts are reprogrammed, exploiting the miR-200 family property to induce a high plasticity state in somatic cells. Subsequently, miR-200-reprogrammed cells are either driven towards the trophectoderm (TR) lineage using an ad hoc induction protocol or encapsulated into polytetrafluoroethylene micro-bioreactors to maintain and promote pluripotency, generating inner cell mass (ICM)-like spheroids. The obtained TR-like cells and ICM-like spheroids are then co-cultured in the same micro-bioreactor and, subsequently, transferred to microwells to encourage blastoid formation. Notably, the above protocol was applied to fibroblasts obtained from young as well as aged donors, with results that highlighted miR-200’s ability to successfully reprogram young and aged cells with comparable blastoid rates, regardless of the donor’s cell age. Overall, the approach here described represents a novel strategy for the creation of artificial blastoids to be used in the field of assisted reproduction technologies for the study of peri- and early post-implantation mechanisms.

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3D Multispheroid Assembly Strategies towards Tissue Engineering and Disease Modeling

Zhu,

Hu,

Cui

et al. 2024

Adv Healthcare Materials

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Cell spheroids (esp. organoids) as 3D culture platforms are popular models for representing cell–cell and cell–extracellular matrix (ECM) interactions, bridging the gap between 2D cell cultures and natural tissues. 3D cell models with spatially organized multiple cell types are preferred for gaining comprehensive insights into tissue pathophysiology and constructing in vitro tissues and disease models because of the complexities of natural tissues. In recent years, an assembly strategy using cell spheroids (or organoids) as living building blocks has been developed to construct complex 3D tissue models with spatial organization. Here, a comprehensive overview of recent advances in multispheroid assembly studies is provided. The different mechanisms of the multispheroid assembly techniques, i.e., automated directed assembly, noncontact remote assembly, and programmed self‐assembly, are introduced. The processing steps, advantages, and technical limitations of the existing methodologies are summarized. Applications of the multispheroid assembly strategies in disease modeling, drug screening, tissue engineering, and organogenesis are reviewed. Finally, this review concludes by emphasizing persistent issues and future perspectives, encouraging researchers to adopt multispheroid assembly techniques for generating advanced 3D cell models that better resemble real tissues.

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3D ECM-rich environment sustains the identity of naive human iPSCs

Cited by 13 publications

References 65 publications

Self-renewing human naïve pluripotent stem cells dedifferentiate in 3D culture and form blastoids spontaneously

Self-renewing human naïve pluripotent stem cells dedifferentiate in 3D culture and form blastoids spontaneously

Generation of Artificial Blastoids Combining miR-200-Mediated Reprogramming and Mechanical Cues

3D Multispheroid Assembly Strategies towards Tissue Engineering and Disease Modeling

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