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Quantitative Live Imaging of Human Embryonic Stem Cell Derived Neural Rosettes Reveals Structure-Function Dynamics Coupled to Cortical Development.

Ziv O, Zaritsky A, Yaffe Y, Mutukula N, Edri R, Elkabetz Y - PLoS Comput. Biol. (2015)

Bottom Line: In contrast, later derived rosettes, which are characterized by reduced NSC capacity and elevated numbers of differentiated neurons, and thus correspond to neurogenesis mode in the developing cortex, exhibit slower motions and decreased radial organization.Finally, molecular perturbations of INM by inhibition of actin or non-muscle myosin-II (NMII) reduced INM measures.Our framework enables quantification of cytoarchitecture NSC dynamics and may have implications in functional molecular studies, drug screening, and iPS cell-based platforms for disease modeling.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, Sackler School of Medicine, Tel Aviv University, Ramat Aviv, Tel Aviv, Israel.

ABSTRACT
Neural stem cells (NSCs) are progenitor cells for brain development, where cellular spatial composition (cytoarchitecture) and dynamics are hypothesized to be linked to critical NSC capabilities. However, understanding cytoarchitectural dynamics of this process has been limited by the difficulty to quantitatively image brain development in vivo. Here, we study NSC dynamics within Neural Rosettes--highly organized multicellular structures derived from human pluripotent stem cells. Neural rosettes contain NSCs with strong epithelial polarity and are expected to perform apical-basal interkinetic nuclear migration (INM)--a hallmark of cortical radial glial cell development. We developed a quantitative live imaging framework to characterize INM dynamics within rosettes. We first show that the tendency of cells to follow the INM orientation--a phenomenon we referred to as radial organization, is associated with rosette size, presumably via mechanical constraints of the confining structure. Second, early forming rosettes, which are abundant with founder NSCs and correspond to the early proliferative developing cortex, show fast motions and enhanced radial organization. In contrast, later derived rosettes, which are characterized by reduced NSC capacity and elevated numbers of differentiated neurons, and thus correspond to neurogenesis mode in the developing cortex, exhibit slower motions and decreased radial organization. Third, later derived rosettes are characterized by temporal instability in INM measures, in agreement with progressive loss in rosette integrity at later developmental stages. Finally, molecular perturbations of INM by inhibition of actin or non-muscle myosin-II (NMII) reduced INM measures. Our framework enables quantification of cytoarchitecture NSC dynamics and may have implications in functional molecular studies, drug screening, and iPS cell-based platforms for disease modeling.

No MeSH data available.


Related in: MedlinePlus

Radial score is associated with rosette size, and enhanced for E-RG rosettes.A. Radial score is associated with rosette size. RS of E-RG (Pearson Rho = -0.8, p = 1.55E-06) and M-RG (Pearson Rho = -0.68, p = 0.0112) are associated with rosette size. RS of most M-RG rosettes are above the linear fit of E-RG RS and rosette size (black line), implying reduced radial organization. B. Radial score is elevated for E-RG rosettes. Boxplots showing signed distances between RS of E-RG and M-RG rosettes to the linear fit of E-RG RS and rosette size. Value of 0 implies perfect fit, positive values indicate reduced radial organization. E-RG rosettes are characterized by enhanced radial organization (reduced RS) than M-RG rosettes (Wilcoxon rank-sum test, p = 0.048). 25 E-RG rosettes and 14 M-RG rosettes were analyzed.
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pcbi.1004453.g003: Radial score is associated with rosette size, and enhanced for E-RG rosettes.A. Radial score is associated with rosette size. RS of E-RG (Pearson Rho = -0.8, p = 1.55E-06) and M-RG (Pearson Rho = -0.68, p = 0.0112) are associated with rosette size. RS of most M-RG rosettes are above the linear fit of E-RG RS and rosette size (black line), implying reduced radial organization. B. Radial score is elevated for E-RG rosettes. Boxplots showing signed distances between RS of E-RG and M-RG rosettes to the linear fit of E-RG RS and rosette size. Value of 0 implies perfect fit, positive values indicate reduced radial organization. E-RG rosettes are characterized by enhanced radial organization (reduced RS) than M-RG rosettes (Wilcoxon rank-sum test, p = 0.048). 25 E-RG rosettes and 14 M-RG rosettes were analyzed.

Mentions: We hypothesized that larger rosettes are characterized by enhanced structured motion, as a response to increased mechanical constraints by the rosette cytoarchitecture. To test this, we examined the association between RS and rosette size. This property was first examined for E-RG rosettes, speculated to have a more structured dynamics compared to the more advanced M-RG rosettes. Indeed, RS of E-RG rosettes was found to be associated with rosette size (Fig 3A), indicating that larger rosettes exhibit enhanced radial migration. Similarly to E-RG rosettes, RS of M-RG rosettes was also found to be associated to rosette size (Fig 3A), suggesting that the association between rosette size and RS is an intrinsic property. This result also implies that the robustness of INM is augmented when larger numbers of nuclei move together towards apical or basal sites, as previously suggested for INM in pseudostratified neuroepithelial cells in vivo [12]. A similar model works also in other studies of collective cell migration showing enhanced group coordinated motility correlated with group size [28–32]. We therefore suggest that the confined structures of larger rosettes lead to more mechanical constraints that explain the increased radial organization of larger rosettes.


Quantitative Live Imaging of Human Embryonic Stem Cell Derived Neural Rosettes Reveals Structure-Function Dynamics Coupled to Cortical Development.

Ziv O, Zaritsky A, Yaffe Y, Mutukula N, Edri R, Elkabetz Y - PLoS Comput. Biol. (2015)

Radial score is associated with rosette size, and enhanced for E-RG rosettes.A. Radial score is associated with rosette size. RS of E-RG (Pearson Rho = -0.8, p = 1.55E-06) and M-RG (Pearson Rho = -0.68, p = 0.0112) are associated with rosette size. RS of most M-RG rosettes are above the linear fit of E-RG RS and rosette size (black line), implying reduced radial organization. B. Radial score is elevated for E-RG rosettes. Boxplots showing signed distances between RS of E-RG and M-RG rosettes to the linear fit of E-RG RS and rosette size. Value of 0 implies perfect fit, positive values indicate reduced radial organization. E-RG rosettes are characterized by enhanced radial organization (reduced RS) than M-RG rosettes (Wilcoxon rank-sum test, p = 0.048). 25 E-RG rosettes and 14 M-RG rosettes were analyzed.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4608579&req=5

pcbi.1004453.g003: Radial score is associated with rosette size, and enhanced for E-RG rosettes.A. Radial score is associated with rosette size. RS of E-RG (Pearson Rho = -0.8, p = 1.55E-06) and M-RG (Pearson Rho = -0.68, p = 0.0112) are associated with rosette size. RS of most M-RG rosettes are above the linear fit of E-RG RS and rosette size (black line), implying reduced radial organization. B. Radial score is elevated for E-RG rosettes. Boxplots showing signed distances between RS of E-RG and M-RG rosettes to the linear fit of E-RG RS and rosette size. Value of 0 implies perfect fit, positive values indicate reduced radial organization. E-RG rosettes are characterized by enhanced radial organization (reduced RS) than M-RG rosettes (Wilcoxon rank-sum test, p = 0.048). 25 E-RG rosettes and 14 M-RG rosettes were analyzed.
Mentions: We hypothesized that larger rosettes are characterized by enhanced structured motion, as a response to increased mechanical constraints by the rosette cytoarchitecture. To test this, we examined the association between RS and rosette size. This property was first examined for E-RG rosettes, speculated to have a more structured dynamics compared to the more advanced M-RG rosettes. Indeed, RS of E-RG rosettes was found to be associated with rosette size (Fig 3A), indicating that larger rosettes exhibit enhanced radial migration. Similarly to E-RG rosettes, RS of M-RG rosettes was also found to be associated to rosette size (Fig 3A), suggesting that the association between rosette size and RS is an intrinsic property. This result also implies that the robustness of INM is augmented when larger numbers of nuclei move together towards apical or basal sites, as previously suggested for INM in pseudostratified neuroepithelial cells in vivo [12]. A similar model works also in other studies of collective cell migration showing enhanced group coordinated motility correlated with group size [28–32]. We therefore suggest that the confined structures of larger rosettes lead to more mechanical constraints that explain the increased radial organization of larger rosettes.

Bottom Line: In contrast, later derived rosettes, which are characterized by reduced NSC capacity and elevated numbers of differentiated neurons, and thus correspond to neurogenesis mode in the developing cortex, exhibit slower motions and decreased radial organization.Finally, molecular perturbations of INM by inhibition of actin or non-muscle myosin-II (NMII) reduced INM measures.Our framework enables quantification of cytoarchitecture NSC dynamics and may have implications in functional molecular studies, drug screening, and iPS cell-based platforms for disease modeling.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, Sackler School of Medicine, Tel Aviv University, Ramat Aviv, Tel Aviv, Israel.

ABSTRACT
Neural stem cells (NSCs) are progenitor cells for brain development, where cellular spatial composition (cytoarchitecture) and dynamics are hypothesized to be linked to critical NSC capabilities. However, understanding cytoarchitectural dynamics of this process has been limited by the difficulty to quantitatively image brain development in vivo. Here, we study NSC dynamics within Neural Rosettes--highly organized multicellular structures derived from human pluripotent stem cells. Neural rosettes contain NSCs with strong epithelial polarity and are expected to perform apical-basal interkinetic nuclear migration (INM)--a hallmark of cortical radial glial cell development. We developed a quantitative live imaging framework to characterize INM dynamics within rosettes. We first show that the tendency of cells to follow the INM orientation--a phenomenon we referred to as radial organization, is associated with rosette size, presumably via mechanical constraints of the confining structure. Second, early forming rosettes, which are abundant with founder NSCs and correspond to the early proliferative developing cortex, show fast motions and enhanced radial organization. In contrast, later derived rosettes, which are characterized by reduced NSC capacity and elevated numbers of differentiated neurons, and thus correspond to neurogenesis mode in the developing cortex, exhibit slower motions and decreased radial organization. Third, later derived rosettes are characterized by temporal instability in INM measures, in agreement with progressive loss in rosette integrity at later developmental stages. Finally, molecular perturbations of INM by inhibition of actin or non-muscle myosin-II (NMII) reduced INM measures. Our framework enables quantification of cytoarchitecture NSC dynamics and may have implications in functional molecular studies, drug screening, and iPS cell-based platforms for disease modeling.

No MeSH data available.


Related in: MedlinePlus