Surface Topography Guides Morphology and Spatial Patterning of Induced Pluripotent Stem Cell Colonies

Abagnale, Giulio; Sechi, Antonio; Steger, Michael F.; Zhou, Qihui; Kuo, Chao-Chung; Aydin, Gülcan; Schalla, Carmen; Müller‐Newen, Gerhard; Zenke, Martin; Costa, Ivan G.; Rijn, Patrick van; Gillner, Arnold; Wagner, Wolfgang

doi:10.1016/j.stemcr.2017.06.016

Cited by 125 publications

(111 citation statements)

References 57 publications

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“…Despite their uniform appearance, there is also marked heterogeneity within colonies of iPSCs [32,33]. Therefore, we analyzed if specific subclones become dominant during serial passaging of iPSCs.…”

Section: The Clonal Composition Of Ipscs Remains Relatively Stable Dumentioning

confidence: 99%

Genetic barcoding reveals clonal dominance in iPSC-derived mesenchymal stromal cells

et al. 2020

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Background: The use of mesenchymal stromal cells (MSCs) for research and clinical application is hampered by cellular heterogeneity and replicative senescence. Generation of MSC-like cells from induced pluripotent stem cells (iPSCs) may circumvent these limitations, and such iPSC-derived MSCs (iMSCs) are already tested in clinical trials. So far, a comparison of MSCs and iMSCs was particularly addressed in bulk culture. Despite the high hopes in cellular therapy, only little is known how the composition of different subclones changes in these cell preparations during culture expansion. Methods: In this study, we used multicolor lentiviral genetic barcoding for the marking of individual cells within cell preparations. Based on this, we could track the clonal composition of syngenic MSCs, iPSCs, and iMSCs during culture expansion. Furthermore, we analyzed DNA methylation patterns at senescence-associated genomic regions by barcoded bisulfite amplicon sequencing. The proliferation and differentiation capacities of individual subclones within MSCs and iMSCs were investigated with limiting dilution assays. Results: Overall, the clonal composition of primary MSCs and iPSCs gradually declined during expansion. In contrast, iMSCs became oligoclonal early during differentiation, indicating that they were derived from few individual iPSCs. This dominant clonal outgrowth of iMSCs was not associated with changes in chromosomal copy number variation. Furthermore, clonal dynamics were not clearly reflected by stochastically acquired DNA methylation patterns. Limiting dilution assays revealed that iMSCs are heterogeneous in colony formation and in vitro differentiation potential, while this was even more pronounced in primary MSCs. Conclusions: Our results indicate that the subclonal diversity of MSCs and iPSCs declines gradually during in vitro culture, whereas derivation of iMSCs may stem from few individual iPSCs. Differentiation regimen needs to be further optimized to achieve homogeneous differentiation of iPSCs towards iMSCs.

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Section: The Clonal Composition Of Ipscs Remains Relatively Stable Dumentioning

confidence: 99%

Genetic barcoding reveals clonal dominance in iPSC-derived mesenchymal stromal cells

et al. 2020

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“…Micropatterning tools have further uncovered the role of the cytoskeleton during differentiation through analysis of how RhoA/Rho-associated kinase (ROCK) impacts actomyosin contractility in MSCs undergoing fate decision toward adipocyte and osteocyte lineages (16)(17)(18)(19). Nonetheless, these studies rely on the presence of soluble cues, while studies exploring the undeveloped cytoskeletal structure of hiPSCs, as it relates to their differentiation capacity, have been relatively unexplored (20,21). Here we utilized micropatterning to show how the interplay between cytoskeletal tension (i.e., active forces developed by the cell) and local cell density, reported as the fraction of each local region occupied by nuclei, modulates mesodermal fate independently of soluble cues, which subsequently leads to vascular commitment from hiPSCs.…”

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confidence: 99%

Cytoskeletal tension regulates mesodermal spatial organization and subsequent vascular fate

Smith

Rochman

Carmo

et al. 2018

Proc. Natl. Acad. Sci. U.S.A.

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Morphogenesis during human development relies on the interplay between physiochemical cues that are mediated in part by cellular density and cytoskeletal tension. Here, we interrogated these factors on vascular lineage specification during human-induced pluripotent stem-cell (hiPSC) fate decision. We found that independent of chemical cues, spatially presented physical cues induce the self-organization of Brachyury-positive mesodermal cells, in a RhoA/Rho-associated kinase (ROCK)-dependent manner. Using unbiased support vector machine (SVM) learning, we found that density alone is sufficient to predict mesodermal fate. Furthermore, the long-withstanding presentation of spatial confinement during hiPSC differentiation led to an organized vascular tissue, reminiscent of native blood vessels, a process dependent on cell density as found by SVM analysis. Collectively, these results show how tension and density relate to vascular identity mirroring early morphogenesis. We propose that such a system can be applied to study other aspects of the stem-cell niche and its role in embryonic patterning.

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“…Hereinafter, our results revealed that tested PLA macro‐topographies similarly influenced different DPSC orientation and morphology: wavy surface determined more straightened morphology of cells, meanwhile, DPSC seeded on porous scaffold were flatter and bent over on the PLA filament. Moreover, scaffold's macro‐topographies also influenced DPSC nuclei shape, although previously it was thought that only micro‐ and nano‐structures can influence cell morphology. Therefore, our results imply that surface pattern in the scale of >100 μm still alters cell shape and induces different ECM tensions which undoubtedly affect cell fate.…”

Section: Discussionmentioning

confidence: 62%

The effect of larger than cell diameter polylactic acid surface patterns on osteogenic differentiation of rat dental pulp stem cells

Alksnė

Šimoliūnas

Kalvaitytė

et al. 2018

J Biomedical Materials Res

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Topography of the scaffold is one of the most important factors defining the quality of artificial bone. However, the production of precise micro-and nano-structured scaffolds, which is known to enhance osteogenic differentiation, is expensive and time-consuming. Meanwhile, little is known about macro-patterns (larger than cell diameter) effect on cell fate, while this kind of structures would significantly facilitate the manufacturing of artificial skeleton. Therefore, this research is focused on polylactic acid scaffold's macropattern impact on rat's dental pulp stem cells (DPSCs) morphology, proliferation, and osteogenic differentiation. For this study, two types of scaffolds were 3D printed: wavy and porous. Wavy scaffolds consisted of 188 μm wide joined threads, meaning that cells might have been curved on the filament as well as compressed in the groove. Porous scaffolds were designed to avoid groove formation and consisted of 500 μm threads, arranged in the woodpile manner, forming 300 μm diameter pores. We found that both macro-surfaces influenced DPSC morphology compared to control. As a consequence, enhanced DPSC proliferation and increased osteogenic differentiation potential was registered in cells grown on these scaffolds. Finally, our results showed that the construction of an artificial bone did not necessarily require the precise structuring of the scaffold, because both types of macrotopographic PLA scaffolds were sufficient enough to induce spontaneous DPSC osteogenic differentiation. How to cite this article: Alksne M, Simoliunas E, Kalvaityte M, Skliutas E, Rinkunaite I, Gendviliene I, Baltriukiene D, Rutkunas V, Bukelskiene V. 2019. The effect of larger than cell diameter polylactic acid surface patterns on osteogenic differentiation of rat dental pulp stem cells. J Biomed Mater Res Part A 2019:107A:174-186.

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Surface Topography Guides Morphology and Spatial Patterning of Induced Pluripotent Stem Cell Colonies

Cited by 125 publications

References 57 publications

Genetic barcoding reveals clonal dominance in iPSC-derived mesenchymal stromal cells

Genetic barcoding reveals clonal dominance in iPSC-derived mesenchymal stromal cells

Cytoskeletal tension regulates mesodermal spatial organization and subsequent vascular fate

The effect of larger than cell diameter polylactic acid surface patterns on osteogenic differentiation of rat dental pulp stem cells

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