Project description:The variation among induced pluripotent stem cells (iPSCs) in their differentiation capacity to specific lineages is frequently attributed to somatic memory. In this study, we compared hematopoietic differentiation capacity of 35 human iPSC lines derived from four different tissues and four embryonic stem cell lines. The analysis revealed that hematopoietic commitment capacity (PSCs to hematopoietic precursors) is correlated with the expression level of the IGF2 gene independent of the iPSC origins. In contrast, maturation capacity (hematopoietic precursors to mature blood) is affected by iPSC origin; blood-derived iPSCs showed the highest capacity. However, some fibroblast-derived iPSCs showed higher capacity than blood-derived clones. Tracking of DNA methylation changes during reprogramming reveals that maturation capacity is highly associated with aberrant DNA methylation acquired during reprogramming, rather than the types of iPSC origins. These data demonstrated that variations in the hematopoietic differentiation capacity of iPSCs are not attributable to somatic memories of their origins. Methyl-seq analysis for undifferentiated induced pluripotent stem cell (iPSC) lines (n = 21), human dermal fibroblast (HDF, n = 1), human peripheral blood (n = 1), and human keratinocyte (n = 1), and ATAC-seq analysis for 2 iPSC lines and an embryonic stem cell (ESC) line with two different culture conditions. CTCF-ChIP-seq analysis for an ESC line.
Project description:Trophoblast differentiation from human ESC has been achieved by exposing the cells to BMP4 with or without supplementation of ALK4/5/7 inhibitor (A83-01) and FGF2 signaling inhibitor (PD173074) (BAP). Here the two differentiation conditions, BMP4 and BAP were applied to two sets of human PSC lines, H1 ESC and iPSC that latter was generated by DOX-inducible lentiviral (V) transductions of umbilical cord mesenchymal cells. The V-iPSC showed residual transgene expressions from the viral vectors in DOX-free culture condition. When the both ESC and iPSC lines were differentiated simultaneously, similar time dependent morphological changes were observed but BMP4 treated V-iPSC showed a minor yet consistent lag in the differentiation progression compared to BMP4 treated hESC. Although both differentiated ESC and V-iPSC showed dominant trophoblast phenotypes, the BMP4 treated V-iPSC also expressed gene markers consistent with the presence of mesoendoderm. The BAP condition provided more efficient differentiation than BMP4 alone, and the BAP-differentiated iPSC and ESC never expressed mesoendoderm markers.
Project description:With the advent of the induced pluripotent stem cell (iPSC) technology, how to distinguish the developmental potentials of the iPSC clones with molecular approaches becomes an imperative issue. Herein, we demonstrated that histone variant H2A.X plays an unexpected role in distinguishing the developmental potentials of iPSC. We showed that H2A.X is specifically targeted to and negatively regulates extra-embryonic lineage gene expression in embryonic stem cell (ESCs) and therefore, it prevents trophectoderm (TE) lineage differentiation under inductive conditions. ESC-specific H2A.X deposition and functions are faithfully recapitulated in the iPSC lines that support the development of “all-iPS” animals. In iPSC lines that fail to support embryonic development, aberrant H2A.X depositions result in upregulation of extra-embryonic lineage genes and predisposition to extra-embryonic tissue differentiation. In summary, our work has revealed novel epigenetic mechanisms for maintaining cell lineage commitment, which can be used to distinguish the quality of the iPSC lines. Detect and compare different H2A.X deposition patterns in ES cells [GSE42306] and TS cells, with Illumina HiSeq 2000
Project description:Trophoblast differentiation from human ESC has been achieved by exposing the cells to BMP4 with or without supplementation of ALK4/5/7 inhibitor (A83-01) and FGF2 signaling inhibitor (PD173074) (BAP). Here the two differentiation conditions, BMP4 and BAP were applied to two sets of human PSC lines, H1 ESC and iPSC that latter was generated by DOX-inducible lentiviral (V) transductions of umbilical cord mesenchymal cells. The V-iPSC showed residual transgene expressions from the viral vectors in DOX-free culture condition. When the both ESC and iPSC lines were differentiated simultaneously, similar time dependent morphological changes were observed but BMP4 treated V-iPSC showed a minor yet consistent lag in the differentiation progression compared to BMP4 treated hESC. Although both differentiated ESC and V-iPSC showed dominant trophoblast phenotypes, the BMP4 treated V-iPSC also expressed gene markers consistent with the presence of mesoendoderm. The BAP condition provided more efficient differentiation than BMP4 alone, and the BAP-differentiated iPSC and ESC never expressed mesoendoderm markers. Five samples, one control of undifferentiated V-iPSC (FGF2) and four differentiated samples included two H1 ESC (treated with BMP4 or BMP4+A83-01) and similarly treated two V-iPSC were analyzed. Trophoblast differentiation was conducted with BMP4 (10 ng/ml) with or without supplementation of ALK4/5/7 inhibitor (A83-01; 1 μM) to H1 ESC and V-iPSC, respective reagents were added to FGF2-free MEF-CM from the second day culture of following passages for up to six additional days.
Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs.
Project description:The variation among induced pluripotent stem cells (iPSCs) in their differentiation capacity to specific lineages is frequently attributed to somatic memory. In this study, we compared hematopoietic differentiation capacity of 35 human iPSC lines derived from four different tissues and four embryonic stem cell lines. The analysis revealed that hematopoietic commitment capacity (PSCs to hematopoietic precursors) is correlated with the expression level of the IGF2 gene independent of the iPSC origins. In contrast, maturation capacity (hematopoietic precursors to mature blood) is affected by iPSC origin; blood-derived iPSCs showed the highest capacity. However, some fibroblast-derived iPSCs showed higher capacity than blood-derived clones. Tracking of DNA methylation changes during reprogramming reveals that maturation capacity is highly associated with aberrant DNA methylation acquired during reprogramming, rather than the types of iPSC origins. These data demonstrated that variations in the hematopoietic differentiation capacity of iPSCs are not attributable to somatic memories of their origins. iPSC/ESC-derived erythroblasts (n = 6)
Project description:Transcriptional profiling of human mesenchymal stem cells comparing normoxic MSCs cells with hypoxic MSCs cells. Hypoxia may inhibit senescence of MSCs during expansion. Goal was to determine the effects of hypoxia on global MSCs gene expression.
Project description:With the advent of the induced pluripotent stem cell (iPSC) technology, how to distinguish the developmental potentials of the iPSC clones with molecular approaches becomes an imperative issue. Herein, we demonstrated that histone variant H2A.X plays an unexpected role in distinguishing the developmental potentials of iPSC. We showed that H2A.X is specifically targeted to and negatively regulates extra-embryonic lineage gene expression in embryonic stem cell (ESCs) and therefore, it prevents trophectoderm (TE) lineage differentiation under inductive conditions. ESC-specific H2A.X deposition and functions are faithfully recapitulated in the iPSC lines that support the development of “all-iPS” animals. In iPSC lines that fail to support embryonic development, aberrant H2A.X depositions result in upregulation of extra-embryonic lineage genes and predisposition to extra-embryonic tissue differentiation. In summary, our work has revealed novel epigenetic mechanisms for maintaining cell lineage commitment, which can be used to distinguish the quality of the iPSC lines.