Project description:Induced pluripotent stem cells (iPSCs) offer opportunity for insight into the genetic requirements of the X chromosome for somatic and germline development. Turner syndrome is caused by complete or partial loss of the second sex chromosome; while more than 90% of Turner cases result in spontaneous fetal loss, survivors display an array of somatic and germline clinical characteristics. Here, we derived iPSCs from Turner syndrome and control individuals and examined germ cell development as a function of X chromosome composition. We analyzed gene expression profiles of derived iPSCs and in vitro differentiated cells by single cell qRT-PCR and RNA-seq. We whoed that two X chromosomes are not necessary for reprogramming or pluripotency maintenance. Genes that escape X chromosome inactivation (XCI) between control iPSCs and those with X chromosome aneuploidies revealed minimal expression differences relative to a female hESC line. Moreover, when we induced germ cell differentiation via murine xenotransplantation of iPSC lines into the seminiferous tubules of busulfan-treated mice, we observed that undifferentiated iPSCs, independent of X chromosome composition, when placed within the correct somatic environment, are capable of forming early germ cells in vivo. Results indicate that two intact X chromosomes are not required for germ cell formation; however, clinical data suggest that two sex chromosomes are required for maintenance of human germ cells. RNA-seq of H9 cells, iPSCs from Turner syndrome and control individuals and in vitro differentiated cells
Project description:Induced pluripotent stem cells (iPSCs) offer opportunity for insight into the genetic requirements of the X chromosome for somatic and germline development. Turner syndrome is caused by complete or partial loss of the second sex chromosome; while more than 90% of Turner cases result in spontaneous fetal loss, survivors display an array of somatic and germline clinical characteristics. Here, we derived iPSCs from Turner syndrome and control individuals and examined germ cell development as a function of X chromosome composition. We analyzed gene expression profiles of derived iPSCs and in vitro differentiated cells by single cell qRT-PCR and RNA-seq. We whoed that two X chromosomes are not necessary for reprogramming or pluripotency maintenance. Genes that escape X chromosome inactivation (XCI) between control iPSCs and those with X chromosome aneuploidies revealed minimal expression differences relative to a female hESC line. Moreover, when we induced germ cell differentiation via murine xenotransplantation of iPSC lines into the seminiferous tubules of busulfan-treated mice, we observed that undifferentiated iPSCs, independent of X chromosome composition, when placed within the correct somatic environment, are capable of forming early germ cells in vivo. Results indicate that two intact X chromosomes are not required for germ cell formation; however, clinical data suggest that two sex chromosomes are required for maintenance of human germ cells.
Project description:The molecular mechanisms of human primordial germ cell (PGC) specification are poorly understood due to the inaccessibility of cell materials and the lack of an alternative in vitro model that enables tracking of the earliest stages of germ cell development. Here, we introduce a defined and efficient differentiation system for the induction of pre-migratory PGC-like cells from human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). By step-wise differentiation, we generated an OCT4+/ T+/BLIMP1+ cell population that transitioned into STELLA expressing PGC-like cells that exhibited a similar key gene expression as mouse PGCs as well as global epigenetic reprogramming. Even though, these PGC-like cells expressed PRDM14 at very low levels, they underwent activation of pluripotency/PGC genes, suppression of neural induction and suppression of de novo DNA methylation, events that are regulated by Prdm14 during mouse PGC specification. This study demonstrates that human PGC commitment shares many key features with mouse PGC specification, but harbors unique and so far unknown mechanisms that, point to a novel human transcriptional regulation. 7 samples were analyzed. ESC: Human Embryonic Stem Cells, 1 biological rep iPSC: Human induced Pluripotent Stem Cells, 1 biological rep d2: Human induced Pluripotent Stem Cells, 2 days differentiation treatment , 2 biological rep d4PGCLC: Human induced Pluripotent Stem Cells, 4 days differentiation treatment towards Primordial Germ Cells Like Cells, 1 biological rep d6PGCLC: Human induced Pluripotent Stem Cells, 6 days differentiation treatment towards Primordial Germ Cells Like Cells, 2 biological rep
Project description:The purpose of this experiment was to compare differences in the transcript levels between RNA samples from induced pluripotent stem cells differentiated into neurons carrying a mutation in the SOD1 gene, an expanded GGGGCC repeat mutation in the chromosome 9 open reading frame 72 gene and those without any (control).
Project description:The Human Induced Pluripotent Stem Cells Initiative (HipSci) is generating a large, high-quality reference panel of human IPSC lines. This is a submission of mass-spectrometry analyses from 6 induced pluripotent stem cell lines generated by the HipSci project.
Project description:There are a total of four samples each for this analysis. Each sample consists of the cells grown on three 10 cm culture plates. Each plate should have 2x106 cells for a total of 6x106 cells per sample when all three plates are combined. The first sample is undifferentiated human embryonic stem cells, the second sample is human glutamatergic neurons derived from those human embryonic stem cells, the third sample is undifferentiated human induced pluripotent stem cells and the fourth sample is human glutamatergic neurons derived from those human induced pluripotent stem cells.
Project description:Background and aim: Human Induced pluripotent stem (iPS) cells have been derived from dermal fibroblasts, keratinocytes and blood cells by ectopic expression of defined transcription factors.1–5 Application of this approach in human cells would have enormous potential and generate patient-specific pluripotent stem cells to accelerate the implementation of stem cells for clinical treatment of degenerative diseases. In the present study, we investigated whether genetically marked human mesenchymal cells of gut mesentery may give rise to iPS cells. Methods: We used lentiviruses to express Oct4, Sox2, Nanog in mesenchymal cells of gut mesentery, then generated iPS cells were identified in many aspects including morphology, pluripotent markers, global gene expression profile, DNA methylation status at pluripotent cell-specific genes, embryoid bodies and terotomas formation. Results: The resulting iPS cells from mesenchymal cells of gut mesentery were similar to human embryonic stem (ES) cells in morphology, proliferation, surface antigens, gene expression, and epigenetic status of pluripotent cell-specific genes. Furthermore, these cells could differentiate into cell types of the three germ layers in vitro and in teratomas. DNA fingerprinting showed that the human iPS cells were derived from the donor cells and are not a result of contamination. one sample/variable