The events regulating human preimplantation development are still largely unknown, due to scarcity of material, ethical and legal limitations, and lack of reliable techniques to faithfully amplify the transcriptome of a single cell. Nonetheless, knowledge in human embryology is gathering renewed interest due to its close relationship with both stem cell biology and epigenetic reprogramming to pluripotency, and their centrality to regenerative medicine. Using carefully timed genome-wide transcript analyses on single oocytes and embryos, the analysis of the data allowed us to uncover a series of successive waves of embryonic transcriptional initiation which start as early as the 2 cell stage. In addition, we identified hierarchical activation of genes involved in the regulation of pluripotency. Finally, we developed HumER, a free database of human preimplantation human development gene expression to serve the scientific community. Importantly, our work links early transcription in the human embryo with the correct execution of the pluripotency program later in development, and paves the way for the identification of factors to improve epigenetic reprogramming.
Waves of early transcriptional activation and pluripotency program initiation during human preimplantation development.
Specimen part, Cell line
View SamplesParthenogenetic stem cells were derived from parthenotes, then differentiated to mesenchymal stem cells. These were further reprogrammed to induced pluripotent stem cells, which were finally differentiated to secondary mesenchymal stem cells.
Accumulation of instability in serial differentiation and reprogramming of parthenogenetic human cells.
Sex, Specimen part
View SamplesIschemic cardiopathy is the leading cause of death in the world, for which efficient regenerative therapy is not currently available. In mammals, after a myocardial infarction episode, the damaged myocardium is replaced by scar tissue featuring collagen deposition and tissue remodelling with negligible cardiomyocyte proliferation. Zebrafish, in contrast, display an extensive regenerative capacity as they are able to restore completely lost cardiac tissue after partial ventricular amputation. Due to the lack of genetic lineage tracing evidence, it is not yet clear if new cardiomyocytes arise from existing contractile cells or from an uncharacterised set of progenitors cells. Nonetheless, several genes and molecules have been shown to participate in this process, some of them being cardiomyocyte mitogens in vitro. Though questions as what are the early signals that drive the regenerative response and what is the relative role of each cardiac cell in this process still need to be answered, the zebrafish is emerging as a very valuable tool to understand heart regeneration and devise strategies that may be of potential value to treat human cardiac disease. Here, we performed a genome-wide transcriptome profile analysis focusing on the early time points of zebrafish heart regeneration and compared our results with those of previously published data. Our analyses confirmed the differential expression of several transcripts, and identified additional genes the expression of which is differentially regulated during zebrafish heart regeneration. We validated the microarray data by conventional and/or quantitative RT-PCR. For a subset of these genes, their expression pattern was analyzed by in situ hybridization and shown to be upregulated in the regenerating area of the heart. The specific role of these new transcripts during zebrafish heart regeneration was further investigated ex vivo using primary cultures of zebrafish cardiomyocytes and/or epicardial cells. Our results offer new insights into the biology of heart regeneration in the zebrafish and, together with future experiments in mammals, may be of potential interest for clinical applications.
Transcriptomics approach to investigate zebrafish heart regeneration.
Specimen part, Time
View SamplesGene expression from cord blood stem cells and respective derived neuronal cells at different times point of differentiation:CD133+ cells;
Cord blood-derived neuronal cells by ectopic expression of Sox2 and c-Myc.
Specimen part, Time
View SamplesHutchinson-Gilford progeria syndrome (HGPS) is a rare and fatal human premature aging disease1-5, characterized by premature atherosclerosis and degeneration of vascular smooth muscle cells (SMCs)6-8. HGPS is caused by a single-point mutation in the LMNA gene, resulting in the generation of progerin, a truncated mutant of lamin A. Accumulation of progerin leads to various aging-associated nuclear defects including disorganization of nuclear lamina and loss of heterochromatin9-12. Here, we report the generation of induced pluripotent stem cells (iPSCs) from fibroblasts obtained from patients with HGPS. HGPS-iPSCs show absence of progerin, and more importantly, lack the nuclear envelope and epigenetic alterations normally associated with premature aging. Upon differentiation of HGPS-iPSCs, progerin and its associated aging consequences are restored. In particular, directed differentiation of HGPS-iPSCs to SMCs leads to the appearance of premature senescent SMC phenotypes associated with vascular aging. Additionally, our studies identify DNA-dependent protein kinase catalytic subunit (DNAPKcs) as a component of the progerin-containing protein complex. The absence of nuclear DNAPKcs correlates with premature as well as physiological aging. Since progerin also accumulates during physiological aging6,12,13, our results provide an in vitro iPSC-based model with an acceleration progerin accumulation to study the pathogenesis of human premature and physiological vascular aging.
Recapitulation of premature ageing with iPSCs from Hutchinson-Gilford progeria syndrome.
Cell line
View SamplesThe generation of induced pluripotent stem (iPS) cells 1-4 has spawned unprecedented opportunities for investigating the molecular logic that underlies cellular pluripotency and reprogramming, as well as for obtaining patient-specific cells for future clinical applications. However, both prospects are hampered by the low efficiency of the reprogramming process. Here, we show that juvenile human primary keratinocytes can be efficiently reprogrammed to pluripotency by retroviral transduction with Oct4, Sox2, Klf4 and c-Myc. Keratinocyte-derived iPS (KiPS) cells appear indistinguishable from human embryonic stem (hES) cells in colony morphology, growth properties, expression of pluripotency-associated transcription factors and surface markers, as well as in vitro and in vivo differentiation potential. Notably, keratinocyte reprogramming to pluripotency is, at least, 100-fold more efficient and 2-fold faster than that of fibroblasts. This increase in reprogramming efficiency allowed us to expand the practicability of the technology and to generate KiPS cells from single plucked hairs from adult individuals.
Efficient and rapid generation of induced pluripotent stem cells from human keratinocytes.
No sample metadata fields
View SamplesInduced pluripotent stem (iPS) cells have generated interest for regenerative medicine as they allow for producing patient-specific progenitors in vitro with potential value for cell therapy. In many instances, however, an off-the-shelf approach would be desirable, such as for cell therapy of acute conditions or when the patient's somatic cells are altered as a consequence of chronic disease or aging. Cord blood (CB) stem cells appear ideally suited for this purpose as they are newborn, immunologically immature cells with minimal genetic and epigenetic alterations, and several hundred thousand immunotyped CB units are readily available through a worldwide network of CB banks. Here, we show that CB stem cells can be reprogrammed to pluripotency by retroviral transduction with OCT4, SOX2, KLF4, and c-MYC, in a process that is extremely efficient and fast. The resulting CB-derived iPS (CBiPS) cells are phenotypically and molecularly indistinguishable from human embryonic stem (hES) cells. Furthermore, we show that generation of CBiPS can be efficiently achieved without the use of the c-MYC and KLF4 oncogenes and just by overexpression of OCT4 and SOX2. Our studies set the basis for the creation of a comprehensive bank of HLA-matched CBiPS cells for off-the-shelf applications.
Generation of induced pluripotent stem cells from human cord blood using OCT4 and SOX2.
Specimen part
View SamplesGene expression data obtained from induced pluripotent stem cells derived from wild type fibroblasts (iPSc WT) and from Gaucher Disease type 2 fibroblasts (GD iPSc). Also, gene expression analysis from the initial fibroblasts was made (WT fibroblasts and GD- fibroblasts), as well as gene expression analysis from a human embryonic stem cell line (hES4).
Neuronopathic Gaucher's disease: induced pluripotent stem cells for disease modelling and testing chaperone activity of small compounds.
Specimen part, Cell line
View SamplesWe analyzed the role of the histone lysine methyltransferase Set7/9 in the differentiation of human embryonic stem (ES) cells. Human ES cell lines expressing a control short hairpin and a short hairpin against Set7/9 were established and the genome wide expression profile was compared between both cell lines at different days during differentiation. Analysis of both profiles indicates that in the absence of Set7/9 there is a delay in the silencing of self-renewal factors as well as in the induction of differentiation markers. These results indicate that Set7/9 plays an active role in the differentiation of human ES cells.
SETD7 Regulates the Differentiation of Human Embryonic Stem Cells.
Specimen part, Cell line
View Samples