RESUMEN
Embryonic stem cells (ESCs) have previously been reported to reprogram somatic cells following fusion. The resulting ES-somatic cell hybrids have been shown to adopt the transcriptional profile of ESCs, suggesting that the pluripotent program is dominant. ES-somatic cell hybrids have most characteristics of pluripotent cells in vitro; however, it remains unclear whether the somatic genome is an active partner in the hybrid cells or simply retained predominately as silent cargo. Furthermore, the functional properties of ES-somatic cell hybrids in vivo have been limited to studies on their contribution to teratomas and developing embryos/chimeras. The extent of their pluripotency remains largely unclear. Here we determined that the somatic genome is actively transcribed by generating ES-somatic cell hybrids using Rag2-deficient ESCs fused to autologous wild-type somatic cells. Rag2 expression was detected during in vitro differentiation, suggesting that the somatic genome follows the correct temporal cues during differentiation. Furthermore, ES-somatic cell hybrids maintain their tetraploid state following 4 weeks of differentiation in vivo and are immune tolerated when transferred into matched individuals. The ES-somatic cell hybrids can efficiently differentiate into hematopoietic precursors in both myeloid and lymphoid lineages in vitro, suggesting that the somatic genome is actively transcribed following cell fusion based reprogramming. However, the ES-somatic cell hybrids showed an altered hematopoietic potential following in vitro differentiation and were unable to show hematopoietic engraftment in a mouse model.
Asunto(s)
Células Madre Embrionarias/citología , Células Híbridas/citología , Animales , Secuencia de Bases , Diferenciación Celular , Linaje de la Célula , Cartilla de ADN , Proteínas de Unión al ADN/genética , Técnicas In Vitro , Ratones , Ratones Endogámicos C57BL , TetraploidíaRESUMEN
We compared bona fide human induced pluripotent stem cells (iPSCs) derived from umbilical cord blood (CB) cells and neonatal keratinocytes (K). As a consequence of both incomplete erasure of tissue-specific methylation and aberrant de novo methylation, CB-iPSCs and K-iPSCs were distinct in genome-wide DNA methylation profiles and differentiation potential. Extended passage of some iPSC clones in culture did not improve their epigenetic resemblance to embryonic stem cells, implying that some human iPSCs retain a residual 'epigenetic memory' of their tissue of origin.
Asunto(s)
Diferenciación Celular , Linaje de la Célula , Metilación de ADN , Epigénesis Genética , Sangre Fetal/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Queratinocitos/metabolismo , Sangre Fetal/citología , Regulación de la Expresión Génica , Genoma Humano , Humanos , Células Madre Pluripotentes Inducidas/citología , Queratinocitos/citología , Análisis por MicromatricesRESUMEN
Human dermal fibroblasts obtained by skin biopsy can be reprogrammed directly to pluripotency by the ectopic expression of defined transcription factors. Here, we describe the derivation of induced pluripotent stem cells from CD34+ mobilized human peripheral blood cells using retroviral transduction of OCT4/SOX2/KLF4/MYC. Blood-derived human induced pluripotent stem cells are indistinguishable from human embryonic stem cells with respect to morphology, expression of surface antigens, and pluripotency-associated transcription factors, DNA methylation status at pluripotent cell-specific genes, and the capacity to differentiate in vitro and in teratomas. The ability to reprogram cells from human blood will allow the generation of patient-specific stem cells for diseases in which the disease-causing somatic mutations are restricted to cells of the hematopoietic lineage.
Asunto(s)
Células Sanguíneas/citología , Desdiferenciación Celular , Proliferación Celular , Células Madre Pluripotentes/citología , Adulto , Antígenos CD34/metabolismo , Células Sanguíneas/metabolismo , Técnicas de Cultivo de Célula , Desdiferenciación Celular/fisiología , Células Cultivadas , Humanos , Factor 4 Similar a Kruppel , Masculino , Modelos Biológicos , Células Madre Pluripotentes/metabolismoRESUMEN
Genetically matched pluripotent embryonic stem (ES) cells generated via nuclear transfer or parthenogenesis (pES cells) are a potential source of histocompatible cells and tissues for transplantation. After parthenogenetic activation of murine oocytes and interruption of meiosis I or II, we isolated and genotyped pES cells and characterized those that carried the full complement of major histocompatibility complex (MHC) antigens of the oocyte donor. Differentiated tissues from these pES cells engrafted in immunocompetent MHC-matched mouse recipients, demonstrating that selected pES cells can serve as a source of histocompatible tissues for transplantation.
Asunto(s)
Células Madre Embrionarias/inmunología , Antígenos H-2/genética , Antígenos H-2/inmunología , Histocompatibilidad , Complejo Mayor de Histocompatibilidad , Partenogénesis , Células Madre Pluripotentes/inmunología , Animales , Diferenciación Celular , Línea Celular , Segregación Cromosómica , Células Madre Embrionarias/citología , Células Madre Embrionarias/fisiología , Femenino , Genotipo , Heterocigoto , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/inmunología , Meiosis , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos CBA , Oocitos/citología , Oocitos/inmunología , Células Madre Pluripotentes/citología , Células Madre Pluripotentes/fisiología , Reacción en Cadena de la Polimerasa , Recombinación Genética , Trasplante de Células MadreRESUMEN
Parthenogenesis and somatic cell nuclear transfer (SCNT) are two methods for deriving embryonic stem (ES) cells that are genetically matched to the oocyte donor or somatic cell donor, respectively. Using genome-wide single nucleotide polymorphism (SNP) analysis, we demonstrate distinct signatures of genetic recombination that distinguish parthenogenetic ES cells from those generated by SCNT. We applied SNP analysis to the human ES cell line SCNT-hES-1, previously claimed to have been derived by SCNT, and present evidence that it represents a human parthenogenetic ES cell line. Genome-wide SNP analysis represents a means to validate the genetic provenance of an ES cell line.