Human embryonic stem cells differentiated in dopaminergic neurons
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE43235 | GEO | 2013/01/03
SECONDARY ACCESSION(S): PRJNA185132
REPOSITORIES: GEO
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Project description:Human embryonic stem cells have been a great interest due to its potential to provide a renewable source of surrogate dopamine neurons for transplantation into ParkinsonM-bM-^@M-^Ys disease patients. We have differentiated hESCs with a monolayer of PA6 cells under 2D and 3D systems for 28 days. Genome-wide microarray analysis showed that our 3D samples increased expression of genes that are involved in neuronal developments such as Wnt, hedgehog and mitogen-activated protein kinase (MAPK) signalling pathways. The results suggest that the 3D differentiation system may have affected the regulatory or signalling mechanisms which enhanced the rate of differentiation towards ectoderm. 3 biological replicates for 2D samples and 2 biological replicates for 3D samples.
2013-01-03 | E-GEOD-43235 | biostudies-arrayexpress
Project description:The identified stromal factors SDF1alpha, sFRP1 and VEGFD induce dopaminergic neuron differentiation of human pluripotent stem cells. Human embryonic stem cell (hESC)-derived dopaminergic (DA) neurons are potentially useful for treating Parkinson’s disease (PD) through cell replacement therapy. Generation of DA neurons from hESCs has been achieved by co-culture with the stromal cell line PA6, a source of stromal cell-derived inducing activity (SDIA). However, the factor(s) produced by stromal cells that constitute SDIA is unknown. We previously reported that medium conditioned by PA6 cells can generate functional DA neurons in the human embryonal carcinoma stem cell line, NTera2. Here we further examined the effects of PA6-conditioned medium and found that it can induce DA neuronal differentiation in both the NTera2 cell line and the hESC line, I6. To identify the factor(s) responsible for SDIA, we used large-scale microarray analysis of gene expression combined with proteomic analysis of PA6-conditioned medium. Four candidate factors (hepatocyte growth factor (HGF), stromal cell-derived factor-1 alpha (SDF1alpha), secreted frizzled-related protein 1 (sFRP1) and vascular endothelial growth factor D (VEGFD)) were identified and immunoaffinity capillary electrophoresis (ICE) was used to establish the protein concentration of these factors in conditioned medium. Upon addition of SDF1alpha, sFRP1, and VEGFD, we observed an increase in the number of tyrosine hydroxylase- and TuJ1- positive cells in both the NTera2 and I6 cell lines. These results indicate that SDF1alpha, sFRP1 and VEGF-D are major components of SDIA, and suggest the potential use of these defined factors to elicit dopaminergic differentiation of pluripotent stem cells as a therapeutic intervention in PD. Mouse embryonic fibroblasts were grown in DMEM medium supplemented with 10% FBS; this conditioned media was used as the control. PA6, mouse stromal cells, were grown in MEM-alpha supplemented with 10% FBS; this conditioned media is known to induce differentiation in hES cells.
2012-06-24 | E-GEOD-20503 | biostudies-arrayexpress
Project description:The identified stromal factors SDF1alpha, sFRP1 and VEGFD induce dopaminergic neuron differentiation of human pluripotent stem cells. Human embryonic stem cell (hESC)-derived dopaminergic (DA) neurons are potentially useful for treating Parkinson’s disease (PD) through cell replacement therapy. Generation of DA neurons from hESCs has been achieved by co-culture with the stromal cell line PA6, a source of stromal cell-derived inducing activity (SDIA). However, the factor(s) produced by stromal cells that constitute SDIA is unknown. We previously reported that medium conditioned by PA6 cells can generate functional DA neurons in the human embryonal carcinoma stem cell line, NTera2. Here we further examined the effects of PA6-conditioned medium and found that it can induce DA neuronal differentiation in both the NTera2 cell line and the hESC line, I6. To identify the factor(s) responsible for SDIA, we used large-scale microarray analysis of gene expression combined with proteomic analysis of PA6-conditioned medium. Four candidate factors (hepatocyte growth factor (HGF), stromal cell-derived factor-1 alpha (SDF1alpha), secreted frizzled-related protein 1 (sFRP1) and vascular endothelial growth factor D (VEGFD)) were identified and immunoaffinity capillary electrophoresis (ICE) was used to establish the protein concentration of these factors in conditioned medium. Upon addition of SDF1alpha, sFRP1, and VEGFD, we observed an increase in the number of tyrosine hydroxylase- and TuJ1- positive cells in both the NTera2 and I6 cell lines. These results indicate that SDF1alpha, sFRP1 and VEGF-D are major components of SDIA, and suggest the potential use of these defined factors to elicit dopaminergic differentiation of pluripotent stem cells as a therapeutic intervention in PD.
2012-06-25 | GSE20503 | GEO
Project description:Tridimensional cardiac differentiation from hiPSCs has been largely described in the literature. However, the exact impact that 3D culture has throughout the entire process of cardiac differentiation remains poorly defined. We developed a robust and efficient 3D platform for cardiomyocyte differentiation from hiPSCs, based on the temporal modulation of WNT signalling using small molecules. 3D aggregates of hiPSCs were generated by forced aggregation in microwells and subsequently differentiated. In order to determine the differences in gene expression profile due to 3D culture throughout the different stages of cardiac differentiation, we compared transcriptional changes between cells in 3D aggregates and standard 2D monolayer cardiac differentiation. Analysis of these data suggests a faster commitment of hiPSCs toward the cardiac lineage and also higher degree of cardiomyocyte functional maturation after 20 days of culture in the 3D aggregates when compared with the 2D monolayer.
2019-06-24 | GSE116574 | GEO
Project description:Purpose: Assess changes in the three-dimensional (3D) chromatin architecture during surface ectoderm commitment and evaluate changes in 3D chromatin architecture in human embryonic stem cells and surface ectoderm cells with and without p63. Methods: Cohesin HiChIP profiles for human embryonic stem cells and surface ectoderm cells with and without p63 were generated in triplicate with deep sequencing on the Illumina HiSeq 4000 sequencer. Using HiC-Pro, paired end reads were aligned to hg19, duplicate reads were removed, and then were assigned to MboI restriction fragments, filtered for valid interactions, and used to generate binned interaction matrices of 10 kb resolution. Contacts were called using FitHiC and filtered for counts >= 10 and FDR < 0.001. Results: The greatest change in 3D chromatin architecture was observed between human embryonic stem cells and surface ectoderm cells, while moderate changes were observed in human embryonic stem cells with and without p63, and limited changes were observed in surface ectoderm cells with and without p63. Conclusion: Both p63 and the surface ectoderm morphogens direct changes in chromatin folding to establish the 3D chromatin landscape in the surface ectoderm.
2018-11-06 | GSE120294 | GEO
Project description:Background: Three-dimensional (3D) in vitro culture systems using human induced pluripotent stem cells (hiPSCs) represent impactful platforms to model neurodegenerative disease biology in physiologically relevant microenvironments. Though many successful biomaterials-based 3D model systems have been established for other neurogenerative diseases, such as Alzheimer’s Disease, relatively few exist for Parkinson’s Disease (PD) research. Methods: We employed tissue engineering approaches to construct a 3D silk scaffold-based platform for the culture of hiPSC-dopaminergic (DA) neurons derived from healthy individuals and PD patients harboring LRRK2 G2019S or GBA N370S mutations. We then compared results from protein, gene expression and metabolic analyses obtained from two-dimensional (2D) and 3D culture systems. Results: The 3D platform enabled the formation of dense dopamine neuronal network architectures and developed biological profiles both similar and distinct from 2D culture systems in healthy and PD disease lines. 3D PD cultures showed elevated levels of α-synuclein and alterations in purine metabolite profiles. Furthermore, computational network analysis of transcriptome networks nominated several novel molecular interactions occurring in neurons from patients with mutations in LRRK2 and GBA. Conclusion: The brain-like 3D system presented here is a realistic platform to interrogate molecular mechanisms underlying PD biology. The key advantages of silk-based bioengineering technology include long-term culture and the ability to incorporate multiple brain-relevant cell types to parse cell-cell interactions in development, disease, and aging.
2022-03-16 | GSE172409 | GEO
Project description:In this work we investigated the molecular mechanisms that sustain the endothelial differentiation of murine embryonic stem cells (ES). When ES cells are co-cultured with the stromal PA6 cells in serum-free medium they differentiate mainly into neurons, thanks to the neural inducing activity exerted by the stroma. The addition of exogenous FGF2 allows also the differentiation of endothelial cells whereas, in presence of exogenous BMP4, ES cells differentiate exclusively into endothelium. A pivotal role in ES cells differentiation is played by the stromal PA6 cells.To analyze the early signals released by PA6 cells in the medium of the co-cultures in response to FGF2 and BMP4, monolayers of PA6 cells were treated with recombinant FGF2 or BMP4 for 3 hours and the gene expression analysis of treated PA6 cultures versus the untreated ones was performed. In this work we investigated the molecular mechanisms that sustain the endothelial differentiation of murine embryonic stem cells (ES). When ES cells are co-cultured with the stromal PA6 cells in serum-free medium they differentiate mainly into neurons, thanks to the neural inducing activity exerted by the stroma. The addition of exogenous FGF2 allows also the differentiation of endothelial cells whereas, in presence of exogenous BMP4, ES cells differentiate exclusively into endothelium. A pivotal role in ES cells differentiation is played by the stromal PA6 cells.To analyze the early signals released by PA6 cells in the medium of the co-cultures in response to FGF2 and BMP4, monolayers of PA6 cells were treated with recombinant FGF2 or BMP4 for 3 hours and the gene expression analysis of treated PA6 cultures versus the untreated ones was performed.
2021-09-05 | GSE164630 | GEO
Project description:In order to investigate the impact of MMP-14 (MT1-MMP) and three-dimensional (3D) culture conditions on the transcriptomes of a human breast adenocarcinoma cell line, we performed a microarray analysis from RNAs isolated from MCF-7 cells expressing either an empty vector (CTRL) or human MMP-14 cDNA (MT1) in monolayer (2D) and 3D collagen (3D Col) growth conditions. MCF-7 cells were stably transfected with either an empty vector (pcDNA3.1/Zeo) or human MMP-14 cDNA (pcDNA3.1-MMP-14/Zeo). Cells were grown for 24, 48 and 72 hours in three-dimensional (3D) type I collagen gels or in monolayer culture conditions. Cells were then lysed in TRIzol and total RNA was isolated. For each experimental condition, total RNAs isolated from 4 independant biological replicates were pooled.
2014-12-15 | E-GEOD-49595 | biostudies-arrayexpress
Project description:A genomic expression comparison was done among neural progenitor cells cultured on 2D substrates, 3D porous polystyrene scaffolds, and as 3D neural spheres (in vivo surrogate), with the goal of assessing the feasibility of establishing the meaning of 3D and associated physiological relevance at the molecular level Neural progenitor cells were cultured on 2D surfaces, in 3D scaffolds and as 3D neural spheres. Chemical cues are controlled by coating. Only spacial properties of the culture systems were compared.
2009-12-22 | E-GEOD-13715 | biostudies-arrayexpress
Project description:The goal of this assay is to investigate the signaling pathways involved in dopamine's promotion on Th2 differentiation and proliferation and to clarify the role of dopaminergic nerve innervation in neonatal asthma pathogenesis, to help identifying potential targets for therapy. To do that, we isolated naïve CD4+ T cells from spleens of non-transgenic C57BL/6 mice according to established protocols, and we cultured them under Th0 condition with/without dopamine treatment. To obtain an unbiased view of gene regulation in mouse CD4+ T cells exposed to dopamine, we analyzed the transcriptome of untreated control CD4+ T cells ('Th0') and dopamine-treated CD4+ T cells ('Th0+Dopa') via explorative RNA-sequencing.
2019-09-26 | GSE137968 | GEO