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Differentiation of Induced Pluripotent Stem Cells towards Mesenchymal Stromal Cells is Hampered by Culture in 3D Hydrogels [RNA-Seq]

ABSTRACT: Directed differentiation of induced pluripotent stem cells (iPSCs) towards specific lineages remains a major challenge in regenerative medicine, while there is a growing perception that this process can be influenced by the three-dimensional environment. In this study, we investigated whether iPSCs can differentiate towards mesenchymal stromal cells (MSCs) when embedded into fibrin hydrogels to enable a one-step differentiation procedure within a scaffold. Differentiation of iPSCs on tissue culture plastic or on top of fibrin hydrogels resulted in a typical MSC-like phenotype. In contrast, iPSCs embedded into fibrin gel gave rise to much smaller cells with heterogeneous growth patterns, absence of fibronectin, faint expression of CD73 and CD105, and reduced differentiation potential towards osteogenic and adipogenic lineage. Transcriptomic analysis demonstrated that characteristic genes for MSCs and extracellular matrix were upregulated on flat substrates, whereas genes of neural development were upregulated in 3D culture. Furthermore, the 3D culture had major effects on DNA methylation profiles, particularly within genes for neuronal and cardiovascular development, while there was no evidence for epigenetic maturation towards MSCs. Taken together, iPSCs could be differentiated towards MSCs on tissue culture plastic or on a flat fibrin hydrogel. In contrast, the differentiation process was heterogeneous and not directed towards MSCs when iPSCs were embedded into the hydrogel.

ORGANISM(S): Homo sapiens

PROVIDER: GSE122545 | GEO | 2019/10/04

REPOSITORIES: GEO

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Differentiation of Induced Pluripotent Stem Cells towards Mesenchymal Stromal Cells is Hampered by Culture in 3D Hydrogels [DNA methylation array]

Project description:Directed differentiation of induced pluripotent stem cells (iPSCs) towards specific lineages remains a major challenge in regenerative medicine, while there is a growing perception that this process can be influenced by the three-dimensional environment. In this study, we investigated whether iPSCs can differentiate towards mesenchymal stromal cells (MSCs) when embedded into fibrin hydrogels to enable a one-step differentiation procedure within a scaffold. Differentiation of iPSCs on tissue culture plastic or on top of fibrin hydrogels resulted in a typical MSC-like phenotype. In contrast, iPSCs embedded into fibrin gel gave rise to much smaller cells with heterogeneous growth patterns, absence of fibronectin, faint expression of CD73 and CD105, and reduced differentiation potential towards osteogenic and adipogenic lineage. Transcriptomic analysis demonstrated that characteristic genes for MSCs and extracellular matrix were upregulated on flat substrates, whereas genes of neural development were upregulated in 3D culture. Furthermore, the 3D culture had major effects on DNA methylation profiles, particularly within genes for neuronal and cardiovascular development, while there was no evidence for epigenetic maturation towards MSCs. Taken together, iPSCs could be differentiated towards MSCs on tissue culture plastic or on a flat fibrin hydrogel. In contrast, the differentiation process was heterogeneous and not directed towards MSCs when iPSCs were embedded into the hydrogel.

2019-10-04 | GSE122469 | GEO

Soft Hydrogels Support Differentiation of Induced Pluripotent Stem Cells toward Mesenchymal Stromal Cells [expression]

Project description:Induced pluripotent stem cells (iPSCs) can be differentiated toward mesenchymal stromal cells (MSCs), but at least on epigenetic level this transition remains incomplete with the current culture conditions. Hydrogels provide a more physiologic three-dimensional environment for in vitro cell culture than conventional tissue culture plastic (TCP). In this study, we followed the hypothesis that growth and differentiation of primary MSCs and of iPSC-derived MSCs (iMSCs) can be enhanced on hydrogels. To this end, we used a hydrogel made of human platelet lysate (hPL). MSCs were effectively cultured on and inside hPL-gel and demonstrated more structured deposition of extracellular matrix (ECM) components than TCP. Furthermore, hPL-gel supported differentiation of iPSCs toward MSCs. Unexpectedly, the differentiation process seemed to be hardly affected by the substrate: iMSCs generated either on TCP or hPL-gel did not reveal differences in morphology, immunophenotype, or differentiation potential. Moreover, global gene expression and DNA-methylation profiles were almost identical in iMSCs generated on TCP or hPL-gel. Our results indicate that matrix elasticity is less crucial for directed lineage-specific differentiation toward MSCs than expected.

2018-03-13 | GSE95060 | GEO

Soft Hydrogels Support Differentiation of Induced Pluripotent Stem Cells toward Mesenchymal Stromal Cells [methylation]

2018-03-13 | GSE95058 | GEO

RNA-Seq of human spinal cord organoids generated in Matrigel and alginate

Project description:This experiment compares the transcriptome profile of human spinal cord organoids generated from iPSCs in Matrigel, 1% alginate hydrogel, or 2% alginate hydrogel. Spinal cord organoids were generated from human iPSCs and then encapsulated in the respective hydrogels before further maturation. Organoids were harvested on days 30, 60, and 90 for each group. In addition to the differences between hydrogel groups, neuronal and glial markers at each time point were also assessed for the development of the organoids.

2022-12-26 | E-MTAB-12270 | biostudies-arrayexpress

Tissue- and Aging-specific DNA-Methylation Patterns are erased in Mesenchymal Stromal Cells derived from Induced Pluripotent Stem Cells. [Expression profiling]

Project description:Standardization of mesenchymal stromal cells (MSCs) remains a major obstacle in regenerative medicine. Starting material and culture expansion affect cell preparations and render comparison between studies difficult. In contrast, induced pluripotent stem cells (iPSCs) assimilate towards a ground-state and may therefore give rise to more standardized cell preparations. We reprogrammed bone marrow MSCs into iPSCs which were subsequently re-differentiated towards MSCs. These iPS-MSCs revealed similar morphology, immunophenotype, in vitro differentiation potential, and gene expression profiles as primary MSCs. DNA methylation (DNAm) profiles of iPSCs maintained some donor-specific characteristics, whereas tissue-specific, senescence-associated, and age-related DNAm patterns were erased during reprogramming. iPS-MSCs reacquired senescence-associated DNAm during culture expansion but they remained rejuvenated with regard to age-related DNAm. Overall, iPS-MSCs and MSCs are similar in function but differ in their epigenetic makeup. 8 samples were hybridized to the GeneChip Human Gene 1.0 ST Array (Affymetrix)

2014-09-24 | E-GEOD-54766 | biostudies-arrayexpress

Tissue- and Aging-specific DNA-Methylation Patterns are erased in Mesenchymal Stromal Cells derived from Induced Pluripotent Stem Cells. [Methylation profiling]

Project description:Standardization of mesenchymal stromal cells (MSCs) remains a major obstacle in regenerative medicine. Starting material and culture expansion affect cell preparations and render comparison between studies difficult. In contrast, induced pluripotent stem cells (iPSCs) assimilate towards a ground-state and may therefore give rise to more standardized cell preparations. We reprogrammed bone marrow MSCs into iPSCs which were subsequently re-differentiated towards MSCs. These iPS-MSCs revealed similar morphology, immunophenotype, in vitro differentiation potential, and gene expression profiles as primary MSCs. DNA methylation (DNAm) profiles of iPSCs maintained some donor-specific characteristics, whereas tissue-specific, senescence-associated, and age-related DNAm patterns were erased during reprogramming. iPS-MSCs reacquired senescence-associated DNAm during culture expansion but they remained rejuvenated with regard to age-related DNAm. Overall, iPS-MSCs and MSCs are similar in function but differ in their epigenetic makeup. 12 samples were hybridized to the Illumina Infinium 450k Human Methylation Beadchip

2014-09-24 | E-GEOD-54767 | biostudies-arrayexpress

Directed Differentiation of Human iPSCs into Mesenchymal Lineages by Optogenetic Control of TGF-β Signaling

Project description:For chondrogenic studies of optogenetically activated TGF-β signaling, optogenetic human iPSC-derived MSCs were encapsulated in hydrogels (20 million cells/mL of 2% agarose hydrogel). Groups received either no soluble TGF-β or optogenetic stimulation, or soluble TGF-β3 alone, or optogenetic stimulation alone. After 21 days of differentiation, we performed global quantitative proteomics on samples from two independent experiments, with n=3 replicates per group.

2023-07-20 | PXD040232 | Pride

Cholangiocyte organoid culture in hydrogels from decellularized liver tissue

Project description:Culture of human cholangiocyte organoids in hydrogels derived from healthy liver extracellular matrix (LECM) extracts prepared from decellularized human livers are evaluated in an effort to establish a platform for production of cholangiocyte organoids for clinical regenerative applications. Human intrahepatic cholangiocyte organoids (ICO) grown in hydrogels made from LECM are compared those grown in mouse tumor derived basement membrane extracts (BME). Culture was performed with amino acids labeled with stable heavy isotopes to enable separation of ECM from the hydrogels from that of the produced by the cells with mass spectrometry (MS). MS data were used to evaluate the protein production of ICO comparing the different hydrogel substrates. The study also contains evaluation of the properties of the hydrogel substrates and focuses on expansion and differentiation of the ICO.

2022-04-13 | PXD028417 | Pride

Fibrin Hydrogels Induce Mixed Dorsal/Ventral Spinal Neuron Identities During Differentiation of Human Induced Pluripotent Stem Cells

Project description:We hypothesized that generating spinal motor neurons (sMNs) from human induced pluripotent stem cell (hiPSC)-derived neural aggregates (NAs) using a chemically-defined differentiation protocol would be more effective inside of 3D fibrin hydrogels compared to 2D poly-L-ornithine(PLO)/laminin-coated tissue culture plastic surfaces. We performed targeted RNA-Seq using next generation sequencing to determine the substrate-specific differences in gene expression that regulate cell phenotype. Cells cultured on both substrates expressed sMN genes CHAT and MNX1, though persistent WNT signaling contributed to a higher expression of genes associated with interneurons in NAs cultured in 3D fibrin scaffolds. Cells in fibrin also expressed lower levels of astrocyte progenitor genes and higher levels of the neuronal-specific gene TUBB3, suggesting a purer population of neurons compared to 2D cultures. STATEMENT OF SIGNIFICANCE: Fibrin scaffolds can support the neuronal differentiation of pluripotent stem cells. This study provides insight into how fibrin hydrogels affect neuronal induction by analyzing of the signaling pathways activated during the differentiation process. These insights can then be used to tailor the properties of these hydrogels to optimize the generation of sMNs for regenerative medicine applications.

2017-01-17 | GSE79442 | GEO

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