Differentiation of Human Pluripotent Stem Cells into Functional Endothelial Cells in Scalable Suspension Culture

Olmer, Ruth; Engels, Lena; Usman, Abdulai; Menke, Sandra; Malik, Muhammad Nasir Hayat; Peßler, Frank; Göhring, Gudrun; Bornhorst, Dorothee; Bolten, Svenja Nicolin; Abdelilah‐Seyfried, Salim; Scheper, Thomas; Kempf, Henning; Zweigerdt, Robert; Martin, Ulrich

doi:10.1016/j.stemcr.2018.03.017

Cited by 80 publications

(68 citation statements)

References 73 publications

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“…However, while the process of iPS generation from fully differentiated somatic cells is now well‐established, efficient differentiation of iPS to endothelial cells is still hampered by a gap in knowledge regarding the molecular mechanism that determines endothelial cell phenotype . We aimed to address this question by taking advantage of the procedure of reprogramming and differentiation to explore the processes that participate in repression and activation of the highly endothelial‐specific gene coding for VWF.…”

Section: Discussionmentioning

confidence: 99%

Regulation of von Willebrand Factor Gene in Endothelial Cells That Are Programmed to Pluripotency and Differentiated Back to Endothelial Cells

et al. 2019

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Endothelial cells play a central role in physiological function and pathophysiology of blood vessels in health and disease. However, the molecular mechanism that establishes the endothelial phenotype, and contributes to its signature cell type-specific gene expression, is not yet understood. We studied the regulation of a highly endothelial-specific gene, von Willebrand factor (VWF), in induced pluripotent stem cells generated from primary endothelial cells (human umbilical vein endothelial cells [HUVEC] into a pluripotent state [HiPS]) and subsequently differentiated back into endothelial cells. This allowed us to explore how VWF expression is regulated when the endothelial phenotype is revoked (endothelial cells to HiPS), and re-established (HiPS back to endothelial cells [EC-Diff]). HiPS were generated from HUVECs, their pluripotency established, and then differentiated back to endothelial cells. We established phenotypic characteristics and robust angiogenic function of EC-Diff. Gene array analyses, VWF chromatin modifications, and transacting factors binding assays were performed on the three cell types (HUVEC, HiPS, and EC-Diff). The results demonstrated that generally cohorts of transacting factors that function as transcriptional activators, and those that contribute to histone acetylation and DNA demethylation, were significantly decreased in HiPS compared with HUVECs and EC-Diff. In contrast, there were significant increases in the gene expression levels of epigenetic modifiers that function as methyl transferases in HiPS compared with endothelial cells. The results demonstrated that alterations in chromatin modifications of the VWF gene, in addition to expression and binding of transacting factors that specifically function as activators, are responsible for establishing endothelial specific regulation of the VWF gene. STEM CELLS 2019;37:542-554 SIGNIFICANCE STATEMENTIn these studies, a novel approach involving generation and differentiation of induced pluripotent stem cells is used to revoke and subsequently re-establish a specific cell phenotype, namely endothelial cells, to explore transcriptional regulatory networks that are associated with establishing cell type specific gene regulation. It provides a novel approach, compared with classic analyses of differential gene regulation among cells of distinct phenotypes, toward determining how cell type specific gene expression is regulated. The process allows for determining simultaneously what the critical components that contribute to silencing are, as well as activating target cell-type specific gene regulation.

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Section: Discussionmentioning

confidence: 99%

Regulation of von Willebrand Factor Gene in Endothelial Cells That Are Programmed to Pluripotency and Differentiated Back to Endothelial Cells

et al. 2019

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“…S6bc). For transplant vascularization, endothelial cells (hiPSC-ECs) were generated based on published protocols 39 with minor modification (Fig. S8a).…”

Section: Platelet-derived Factors Promote Mesodermal Specification Ofmentioning

confidence: 99%

“…The maturation phase was extended from passage 4 to passage >8 in α-MEM/10% hPL to obtain mature CD90 + hiPSC-FBs. Bone marrow and umbilical cord blood derived hiPSCs were used for endothelial differentiation according to an established protocol with modifications 39 . In brief, 5 x 10 4 hiPSCs per cm² were seeded as single cells followed by mesoderm induction using the mesoderm induction medium (05221, STEMCELL Technologies) for 48 hours.…”

Section: Isolation and 2d Propagation Of Primary Cellsmentioning

confidence: 99%

Self-assembly of progenitor cells under the aegis of platelet factors facilitates human skin organoid formation and vascularized wound healing

Peking

Krisch

Wolf

et al. 2020

Preprint

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Stem/progenitor cells can self-organize into organoids modelling tissue function and regeneration. Here we demonstrate that human platelet-derived factors can orchestrate 3D self-assembly of clonally expanded adult skin fibroblasts, keratinocytes and endothelial progenitors forming skin organoids within three days. Organoids showed distinct signaling patterns in response to inflammatory stimuli that clearly differed from separated cell types. Human induced pluripotent stem cell (hiPSC)-derived skin cell progenitors also self-assembled into stratified human skin within two weeks, healing deep wounds of immune-deficient mice. Co-transplantation of endothelial progenitors significantly accelerated vascularization. Mechanistically, platelet-derived extracellular vesicles mediated the platelet-derived trophic effects. Long-term fitness of epidermal cells was accelerated further by keratinocyte growth factor mRNA transfection. No tumorigenesis was observed upon xenografting. This permits novel rapid 3D skin-related pharmaceutical testing opportunities and facilitates development of iPSC-based skin regeneration strategies.

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“…In vitro tube formation assay. Tube formation assay was performed as described previously (34). Human umbilical vein endothelial cells (HUVECs; ScienCell Research Laboratories, Inc.) were serum starved overnight at 37˚C in 1640 medium (cat.…”

Section: Cell Differentiation Analysis and Reverse Transcription-quanmentioning

confidence: 99%

Human cardiac extracellular matrix‑chitosan‑gelatin composite scaffold and its endothelialization

Liu

Shi

et al. 2019

Exp Ther Med

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The present study developed a cardiac extracellular matrix-chitosan-gelatin (cECM-CG) composite scaffold that can be used as a tissue-engineered heart patch and investigated its endothelialization potential by incorporating CD34 + endothelial progenitor cells (EPCs). The cECM-CG composite scaffold was prepared by blending cardiac extracellular matrix (cECM) with biodegradable chitosan-gelatin (CG). The mixture was lyophilized using vacuum freeze-drying. CD34 + EPCs were isolated and seeded on the scaffolds, and then the endothelialization effect was subsequently investigated. Effects of the scaffolds on CD34 + EPCs survival and proliferation were evaluated by immunofluorescence staining and MTT assay. Cell differentiation into endothelial cells and the influence of the scaffolds on cell differentiation were investigated by reverse transcription-quantitative PCR (RT-qPCR), immunofluorescence staining and tube formation assay. The present results indicated that most cells were removed after decellularization, but the main extracellular matrix components were retained. Scanning electron microscopy imaging illustrated three-dimensional and porous scaffolds. The present results suggested the cECM-CG composite scaffold had a higher water absorption ability compared with the CG scaffold. Additionally, compared with the CG scaffold, the cECM-CG composite scaffold significantly increased cell survival and proliferation, which suggested its non-toxicity and biocompatibility. Furthermore, RT-qPCR, immunofluorescence and tube formation assay results indicated that CD34 + EPCs differentiated into endothelial cells, and the cECM-CG composite scaffold promoted this differentiation process. In conclusion, the present results indicated that the human cECM-CG composite scaffold generated in the present study was a highly porous, biodegradable three-dimensional scaffold which supported endothelialization of seeded CD34 + EPCs. The present results suggested that this cECM-CG composite scaffold may be a promising heart patch for use in heart tissue engineering for congenital heart disease.

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Differentiation of Human Pluripotent Stem Cells into Functional Endothelial Cells in Scalable Suspension Culture

Cited by 80 publications

References 73 publications

Regulation of von Willebrand Factor Gene in Endothelial Cells That Are Programmed to Pluripotency and Differentiated Back to Endothelial Cells

Regulation of von Willebrand Factor Gene in Endothelial Cells That Are Programmed to Pluripotency and Differentiated Back to Endothelial Cells

Self-assembly of progenitor cells under the aegis of platelet factors facilitates human skin organoid formation and vascularized wound healing

Human cardiac extracellular matrix‑chitosan‑gelatin composite scaffold and its endothelialization

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