Osteogenic Differentiation of Embryonic Stem Cells in 2D and 3D Culture

2010 ◽  
pp. 281-308 ◽  
Author(s):  
Lee Buttery ◽  
Robert Bielby ◽  
Daniel Howard ◽  
Kevin Shakesheff
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Osteogenic Differentiation ◽  
Embryonic Stem ◽  
3D Culture ◽  
2D And 3D

scholarly journals Differentiation of human embryonic stem cells into hepatocytes in 2D and 3D culture systems in vitro

2006 ◽  
Vol 50 (7) ◽  
pp. 645-652 ◽  
Author(s):  
Hossein Baharvand ◽  
Seyed M. Hashemi ◽  
Saeid Kazemi Ashtiani ◽  
Ali Farrokhi
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Embryonic Stem ◽  
3D Culture ◽  
Culture Systems ◽  
2D And 3D

Comparison of osteogenesis of human embryonic stem cells within 2D and 3D culture systems

2008 ◽  
Vol 68 (1) ◽  
pp. 58-67 ◽  
Author(s):  
X.‐F. Tian ◽  
B.‐C. Heng ◽  
Z. Ge ◽  
K. Lu ◽  
A. J. Rufaihah ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Embryonic Stem ◽  
3D Culture ◽  
Culture Systems ◽  
2D And 3D

scholarly journals Osteogenic Effect of Inducible Nitric Oxide Synthase (iNOS)-Loaded Mineralized Nanoparticles on Embryonic Stem Cells

2018 ◽  
Vol 51 (2) ◽  
pp. 746-762 ◽  
Author(s):  
Jin-Sun Lee ◽  
Hong Jae Lee ◽  
Jae Won Lee ◽  
Sang Cheon  Lee ◽  
Jung Sun Heo
Keyword(s):  
Nitric Oxide ◽  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Nitric Oxide Synthase ◽  
Cell Viability ◽  
Osteogenic Differentiation ◽  
Inducible Nitric Oxide Synthase ◽  
Embryonic Stem ◽  
Oxide Synthase ◽  
Inducible Nitric Oxide

Background/Aims: This study investigated the effect of inducible nitric oxide synthase-loaded mineralized nanoparticles (iNOS-MNPs) on the osteogenic differentiation of mouse embryonic stem cells (ESCs). Methods: We prepared iNOS-MNPs using an anionic block copolymer template-mediated calcium carbonate (CaCO3) mineralization process in the presence of iNOS. iNOS-MNPs were spherical and had a narrow size distribution. iNOS was stably loaded within MNPs without denaturation. In order to confirm the successful introduction of iNOS-MNPs into the cytosol of ESCs, intracellular levels of nitric oxide (NO) was determined with a fluorometric analysis. A NO effector molecule, cyclic guanosine 3’,5’ monophosphate (cGMP) was also quantified with a competitive enzyme immunoassay. Cell viability in response to iNOS-MNP treatment was determined using the cell counting kit-8 (CCK-8) assay. Alkaline phosphatase (ALP) activity assay, intracellular calcium quantification assay, and Alizarin red S staining for matrix mineralization were performed to investigate osteogenic differentiation of ESCs. The protein levels of Runt-related transcription factor 2 (RUNX2), osteocalcin (OCN), and osterix (OSX) as osteogenic-related factors were also assessed by immunofluorescence staining and Western blot analysis. The complex pathways associated with iNOS-MNP-derived osteogenic differentiation of ESCs were evaluated by network-based analysis. Results: Cells with iNOS-MNPs displayed a significant increase in NO and cGMP concentration compared with the control group. When cells were exposed to iNOS-MNPs, there were no adverse effects on cell viability. Importantly, iNOS-MNP uptake promoted the osteogenic differentiation of ESCs. Using transcriptome profiling, we obtained 1,836 differentially-induced genes and performed functional enrichment analysis with ClueGO and KEGG. These analyses identified significantly enriched and interconnected molecular pathways such as protein kinase activity, estrogen receptor activity, bone morphogenetic protein (BMP) receptor binding, ligand-gated ion channel activity, and phosphatidylinositol 3-phosphate binding. Conclusion: These findings suggest that iNOS-MNPs can induce osteogenic differentiation in ESCs by integrating complex signaling pathways.

Collagen-alginate microspheres as a 3D culture system for mouse embryonic stem cells differentiation to primordial germ cells

Biologicals ◽  
2017 ◽  
Vol 48 ◽  
pp. 114-120 ◽  
Author(s):  
Vahid Mansouri ◽  
Mohammad Salehi ◽  
Mir davood Omrani ◽  
Zahra Niknam ◽  
Abdolreza Ardeshirylajimi
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Germ Cells ◽  
Culture System ◽  
Primordial Germ Cells ◽  
Embryonic Stem ◽  
3D Culture ◽  
Mouse Embryonic Stem Cells ◽  
3D Culture System

A Novel, Three Dimensional (3D) Culture System for the Efficient, Single-Step Erythropoietic Differentiation of Mouse Embryonic Stem Cells (mESCs)

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2044-2044
Author(s):  
Iliana Fauzi ◽  
Nicki Panoskaltsis ◽  
Athanasios Mantalaris
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Conditioned Medium ◽  
Hematopoietic Cells ◽  
Embryonic Stem ◽  
3D Culture ◽  
Single Step ◽  
Rotating Wall ◽  
Rotating Wall Vessel ◽  
Mesoderm Formation

Abstract Abstract 2044 Current established protocols for the culture and differentiation of embryonic stem cells (ESCs) utilise two-dimensional (2D) tissue culture flasks/dishes. These culture methods are cumbersome and inefficient involving three stages: a) maintenance/expansion of undifferentiated ESCs, b) spontaneous differentiation through formation of embryoid bodies (EBs), and c) dissociation of EBs and replating leading to the terminal differentiation to the desired lineages. One of the major challenges in the use of ESCs for the production of red blood cells is controlling their differentiation pathway(s). Optimal culture conditions and requirements as well as precise differentiation mechanisms and cellular interactions within EBs are still not well characterised, resulting in sub-optimal control of homogenous differentiation especially due to the formation of all three germ layers. Furthermore, cavitation within EBs results in loss of available cell numbers, which reduces the yield and quality of the cellular product outcomes. To date, the most efficient protocols for the generation of oxygen-transporting, enucleated red blood cells from ESCs require co-culture with feeder cells and a multi-step process that lasts for approximately one month rendering such protocols difficult to scale-up. We have developed an integrated, single-step bioprocess that: a) uses conditioned medium (CM) derived from HepG2, a human hepatocarcinoma cell line, that stimulates mesoderm formation, b) facilitates 3D culture through encapsulation of undifferentiated mESCs in hydrogels, c) bypasses EB formation, and d) involves culture in a rotating wall vessel bioreactor that does not require passaging of the cells and is scalable and automatable. Previously, we have shown that in traditional 2D culture systems use of HepG2-CM facilitated early differentiation of mESCs into hematopoietic cells, as shown by expression of C-Myb, C-kit, Gata-2, SCL, and beta-globin genes, in comparison with that of control cultures. A significantly higher number (p≤0.001) of hematopoietic colonies was also achieved in conditioned medium-treated murine embryonic stem cells (CM)-mESC, at day 7 and 14, with a two-fold enhancement of all myeloid-erythroid progenitor colonies. Nucleated eythrocytes and macrophages were identified in the CM-mESC group as early as day 7 of culture. However, attempting to bypass the EB formation step in the 2D culture system did not produce any hematopoietic cells even by the conditioned medium-treated embryonic stem cells. Here, we now demonstrate, that single-step 3D cultures of encapsulated mESCs can produce hematopoietic cells bypassing EB formation. Specifically, undifferentiated mESCs were encapsulated (20,000 cells per hydrogel bead) and placed inside the rotating wall vessel bioreactor. The experimental group was exposed to conditioned medium supplemented with LIF for 3 days to stimulate mesoderm formation bypassing EB formation and terminal hematopoietic differentiation was accomplished by simply changing the culture medium and replacing it with 3U/ml hEPO and 40ng/ml mSCF. A significant increase in the number (p≤ 0.05) of hematopoietic colonies was observed from CM-mESCs at day 14, with a total five-fold expansion as well as enhancement of erythroid progenitors, BFU-E and CFU-E formation. A higher expression of hematopoietic genes, C-Myb, C-kit, Gata-2, SCL, as well as erythroid genes, EKLF and beta-major globin was also reported at 3 weeks of culture in low concentrations of cytokines in the bioreactor. Immunophenotypic analysis of the highly viable cells collected from the CM-mESCs group confirmed the positive expression of the proerythroblast markers (TER-119 and CD71). In conclusion, we have devised a scalable, automatable, single-step process for the derivation of mature erythrocytes from mESCs which may be used for further study of erythroid development and applications in the human system. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Utilization of transgenic models in the evaluation of osteogenic differentiation of embryonic stem cells

2013 ◽  
Vol 54 (4-5) ◽  
pp. 297-305 ◽  
Author(s):  
Dario Repic ◽  
Elena Torreggiani ◽  
Tiziana Franceschetti ◽  
Brya G. Matthews ◽  
Sanja Ivcevic ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Osteogenic Differentiation ◽  
Embryonic Stem ◽  
Transgenic Models

scholarly journals Generation of Otic Sensory Neurons from Mouse Embryonic Stem Cells in 3D Culture

2017 ◽  
Vol 11 ◽  
Author(s):  
Michael Perny ◽  
Ching-Chia Ting ◽  
Sonja Kleinlogel ◽  
Pascal Senn ◽  
Marta Roccio
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Sensory Neurons ◽  
Embryonic Stem ◽  
3D Culture ◽  
Mouse Embryonic Stem Cells
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