Cytoskeletal dynamics during in vitro neurogenesis of induced pluripotent stem cells (iPSCs)

Compagnucci, Claudia; Piermarini, Emanuela; Sferra, Antonella; Borghi, Rossella; Niceforo, Alessia; Petrini, Stefania; Piemonte, Fiorella; Bertini, Enrico

doi:10.1016/j.mcn.2016.10.002

Cited by 10 publications

(16 citation statements)

References 68 publications

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“…Static cultures, in particular, gave rise to very few organoids in the absence of ROCK inhibitor. Not only does ROCK inhibition promote the survival of human pluripotent stem cells upon dissociation , the effects of ROCK inhibition on the actin cytoskeleton and the role of the cytoskeleton in the process of tissue self‐organization has also been documented , and could be a reason for the enhanced retinal formation in our experiments. Lamas et al found that ROCK i enhanced the proliferation of motor neuron progenitors derived from hESCs and hiPSCs, meaning that the effect of ROCK i in neural cultures acts not only to prevent cell death following cellular dissociation, but may also act to promote the expansion of the neural population .…”

Section: Discussionmentioning

confidence: 68%

Systematic Comparison of Retinal Organoid Differentiation from Human Pluripotent Stem Cells Reveals Stage Specific, Cell Line, and Methodological Differences

Mellough

Collin

Queen

et al. 2019

Stem Cells Translational Medicine

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A major goal in the stem cell field is to generate tissues that can be utilized as a universal tool for in vitro models of development and disease, drug development, or as a resource for patients suffering from disease or injury. Great efforts are being made to differentiate human pluripotent stem cells in vitro toward retinal tissue, which is akin to native human retina in its cytoarchitecture and function, yet the numerous existing retinal induction protocols remain variable in their efficiency and do not routinely produce morphologically or functionally mature photoreceptors. Herein, we determine the impact that the method of embryoid body (EB) formation and maintenance as well as cell line background has on retinal organoid differentiation from human embryonic stem cells and human induced pluripotent stem cells. Our data indicate that cell line‐specific differences dominate the variables that underline the differentiation efficiency in the early stages of differentiation. In contrast, the EB generation method and maintenance conditions determine the later differentiation and maturation of retinal organoids. Of the latter, the mechanical method of EB generation under static conditions, accompanied by media supplementation with Y27632 for the first 48 hours of differentiation, results in the most consistent formation of laminated retinal neuroepithelium containing mature and electrophysiologically responsive photoreceptors. Collectively, our data provide substantive evidence for stage‐specific differences in the ability to give rise to laminated retinae, which is determined by cell line‐specific differences in the early stages of differentiation and EB generation/organoid maintenance methods at later stages.

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Section: Discussionmentioning

confidence: 68%

Systematic Comparison of Retinal Organoid Differentiation from Human Pluripotent Stem Cells Reveals Stage Specific, Cell Line, and Methodological Differences

Mellough

Collin

Queen

et al. 2019

Stem Cells Translational Medicine

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“…Studies have shown that the neural differentiation process involves significant reorganization of the cytoskeleton including actin, intermediate filaments, and microtubules 25,26 . A key biomarker for neural stem cells is nestin, an intermediate filament protein whose expression is tightly regulated during differentiation 27 .…”

Section: Resultsmentioning

confidence: 99%

High-throughput physical phenotyping of cell differentiation

et al. 2017

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In this report, we present multiparameter deformability cytometry (m-DC), in which we explore a large set of parameters describing the physical phenotypes of pluripotent cells and their derivatives. m-DC utilizes microfluidic inertial focusing and hydrodynamic stretching of single cells in conjunction with high-speed video recording to realize high-throughput characterization of over 20 different cell motion and morphology-derived parameters. Parameters extracted from videos include size, deformability, deformation kinetics, and morphology. We train support vector machines that provide evidence that these additional physical measurements improve classification of induced pluripotent stem cells, mesenchymal stem cells, neural stem cells, and their derivatives compared to size and deformability alone. In addition, we utilize visual interactive stochastic neighbor embedding to visually map the high-dimensional physical phenotypic spaces occupied by these stem cells and their progeny and the pathways traversed during differentiation. This report demonstrates the potential of m-DC for improving understanding of physical differences that arise as cells differentiate and identifying cell subpopulations in a label-free manner. Ultimately, such approaches could broaden our understanding of subtle changes in cell phenotypes and their roles in human biology.

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“…GFAP is a protein component of IFs in astroglial cells and thus participates in regulating their morphology and motility performances. In this view, several studies have investigated cytoskeleton changes involved in the proliferation and migration of NPCs during adult neurogenesis [21]. In particular, the splicing variant of GFAP, GFAP-δ has been shown specifically expressed in a ribbon below the adult human SVZ and in the SGZ of hippocampus which are commonly recognized as NSC niches and in the subpial cortex where recently active adult neurogenesis has also been demonstrated [2].…”

Section: Discussionmentioning

confidence: 99%

GFAP Delta as Divergent Marker of Human Glial Progenitors

Zalfa¹,

Grasselli²,

Santilli³

et al. 2018

J Stem Cell Res Ther

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Background: Human Neural Stem Cells (hNSCs) are responsible for brain development at prenatal and postnatal stages and for tissue homeostasis and repair after injury on adulthood. The identification of hNSC specific markers is still a debated argument. GFAP-δ, the delta isoform of Glial Fibrillary Acidic Protein (GFAP), is particularly expressed in the Subventricular Zone (SVZ) of the brain and its expression has been related to long-term quiescent NSC. On the other hand, another cytoskeletal protein, gelsolin, is expressed in Neural Progenitor Cells (NPCs) migrating from the SVZ. Methods: In this study, we intended to investigate by immunocytochemistry the co-expression of GFAP-δ and gelsolin in different sources of hNSCs, and hNPCs, with particular emphasis on Good Manifacture Procedures (GMPs) grade hNSC currently used in two clinical trials on Amyotrophic Lateral Sclerosis (ALS) and Multiple Sclerosis (MS) patients. Results: We found that the two proteins are co-expressed by undifferentiated GFAP+ cells but tend to localize in different sub-cellular compartments and to segregate in divergent GFAP+ progenies along with differentiation. Interestingly, we proved that, after transplantation into the brain of rodent models of focal demyelination or transient global ischemia, hNSC integrating in the SVZ still retain the co-expression of GFAP-δ and gelsolin, indicating that hNSC intrinsically co-express the two proteins at the stem stage. Conclusion: These findings suggest that GFAP-δ and gelsolin may represent two candidate markers to distinguish NSC from NSC-deriving divergent astroglial phenotypes.

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Cytoskeletal dynamics during in vitro neurogenesis of induced pluripotent stem cells (iPSCs)

Cited by 10 publications

References 68 publications

Systematic Comparison of Retinal Organoid Differentiation from Human Pluripotent Stem Cells Reveals Stage Specific, Cell Line, and Methodological Differences

Systematic Comparison of Retinal Organoid Differentiation from Human Pluripotent Stem Cells Reveals Stage Specific, Cell Line, and Methodological Differences

High-throughput physical phenotyping of cell differentiation

GFAP Delta as Divergent Marker of Human Glial Progenitors

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