This SuperSeries is composed of the SubSeries listed below.
Epigenetic rejuvenation of mesenchymal stromal cells derived from induced pluripotent stem cells.
Specimen part, Subject
View SamplesStandardization 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.
Epigenetic rejuvenation of mesenchymal stromal cells derived from induced pluripotent stem cells.
Specimen part, Subject
View SamplesMesenchymal stromal cells (MSC) were isolated from human bone marrow. Here, we have compared gene expression profiles of MSC at early and late passages and upon stimulation with transforming growth factor beta 1 (TGF-b1). Stimulation was performed with 1ng/mL TGF-b1 for 1, 4, or 12 hours as indicated. The goal of this study was to determine if senescence-associated gene expression changes and TGF-b1 induced gene expression changes are related.
TGF-beta1 does not induce senescence of multipotent mesenchymal stromal cells and has similar effects in early and late passages.
Specimen part, Treatment, Subject
View SamplesInduced pluripotent stem cells (iPSCs) are usually clonally derived. The selection of fully reprogrammed cells generally involves picking of individual colonies with morphology similar to embryonic stem cells (ESCs). However, successfully reprogrammed cells are highly proliferative and escape from cellular senescence - it is therefore conceivable that they outgrow non-pluripotent and partially reprogrammed cells during culture expansion without the need of clonal selection. In this study, we have reprogrammed human dermal fibroblasts (HDFs) with episomal plasmid vectors. Colony frequency and size was higher when using murine embryonic fibroblasts (MEFs) as stromal support instead of HDFs or human mesenchymal stromal cells (MSCs). We have then compared iPSCs which were either clonally derived by manual selection of a single colony, or derived from bulk-cultures of all initial colonies. After few passages their morphology, expression of pluripotency markers, and gene expression profiles did not reveal any significant differences. Furthermore, clonally-derived and bulk-cultured iPSCs had indistinguishable in vitro differentiation potential towards the three germ layers. Therefore, manual selection of individual colonies does not appear to be necessary for the generation of iPSCs this is of relevance for standardization and automation of cell culture procedures
To clone or not to clone? Induced pluripotent stem cells can be generated in bulk culture.
Sex, Specimen part
View SamplesPrimary cells enter replicative senescence after a limited number of cell divisions. This process is associated with reproducible changes in DNA methylation (DNAm) at specific sites in the genome. The mechanism that drives senescence-associated DNAm changes remains unknown and may arise through drift in DNAm or through regulated, senescence dependent modifications at specific sites in the genome. In this study, we analyzed the reorganization of nuclear architecture and DNA methylation during long-term culture of human fibroblasts and mesenchymal stromal cells (MSCs). [RNA-seq] Overall design: RNA was isolated from 1,000,000 cells of three MSC donors (59, 64, and 73 years old) at passage 4 and passage 13 using the miRNeasy Mini Kit (Qiagen). Gene expression profiles were analzyed by deep sequencing with IlluminaHiSeq 2000 technology with a read length of 50 bases at EMBL gene core facility (Heidelberg, Germany).
Replicative senescence is associated with nuclear reorganization and with DNA methylation at specific transcription factor binding sites.
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View SamplesSurface topography impacts on cell growth and differentiation, but it is not trivial to generate homogeneous surface structures and to define the specific morphological parameters of relevance. In this study, we have compared gene expression profiles of mesenchymal stem cells (MSCs) on nanostructured groove/ridge surfaces. Patterns were generated in polyimide using multi beam laser interference. These structures affected cell size and orientation of human MSCs. Furthermore, the nano-patterns with a periodicity of 650 nm increased differentiation towards osteogenic and adipogenic lineages. However, in absence of differentiation media the surface structures did neither induce differentiation, nor lineage-specific gene expression changes as assessed by genome wide gene expression profiles with Affymetrix microarray technology. Our results demonstrate that grooves and ridges at a periodicity of 650 nm enhance the propensity of MSCs to differentiate towards adipogenic and/or osteogenic lineages but they do not directly govern lineage-specific gene expression changes.
Surface topography enhances differentiation of mesenchymal stem cells towards osteogenic and adipogenic lineages.
Specimen part, Subject
View SamplesThis SuperSeries is composed of the SubSeries listed below.
Matrix elasticity, replicative senescence and DNA methylation patterns of mesenchymal stem cells.
Specimen part, Subject
View SamplesMatrix elasticity influences differentiation of mesenchymal stem cells (MSCs) but it is unclear if these effects are only transient - while the cells reside on the substrate - or if they reflect persistent lineage commitment. In this study, MSCs were continuously culture-expanded in parallel either on polydimethylsiloxane (PDMS) gels of different elasticity or on tissue culture plastic (TCP) to compare impact on replicative senescence, in vitro differentiation, gene expression, and DNA methylation (DNAm) profiles. The maximal number of cumulative population doublings was not affected by matrix elasticity. Differentiation towards adipogenic and osteogenic lineage was increased on soft and rigid biomaterials, respectively - but this propensity was no more evident if cells were transferred to TCP. Global gene expression profiles and DNAm profiles revealed relatively few differences in MSCs cultured on soft or rigid matrices. Furthermore, only moderate DNAm changes were observed upon culture on very soft hydrogels of human platelet lysate. Our results support the notion that matrix elasticity influences cellular differentiation while the cells are organized on the substrate, but it does not have major impact on cell-intrinsic lineage determination, replicative senescence or DNAm patterns.
Matrix elasticity, replicative senescence and DNA methylation patterns of mesenchymal stem cells.
Specimen part, Subject
View SamplesThis SuperSeries is composed of the SubSeries listed below.
The lncRNA HOTAIR impacts on mesenchymal stem cells via triple helix formation.
Specimen part, Treatment
View SamplesLong non coding RNAs are implemented in epigenetic changes and regulation of gene expression. HOTAIR is a promising lncRNA concerning epigenetic regulation. We performed HOTAIR overexpression and knockdown experiments in mesenchymal stromal cells derived from bone marrow. After two weeks cells were harvested and RNA and DNA were isolated. Analysis of gene expression was performed with Human Gene 2.0 ST Array (Affymetrix, Santa Clara, USA). Analysis of DNA methylation was performed with Infinium HumanMethylation450 BeadChips (Illumina, San Diego, USA)
The lncRNA HOTAIR impacts on mesenchymal stem cells via triple helix formation.
Specimen part, Treatment
View Samples