Generation of Human-Induced Pluripotent Stem Cells from Gut Mesentery-Derived Cells by Ectopic Expression of OCT4/SOX2/NANOG
ABSTRACT Induced pluripotent stem (iPS) cells have been generated from human somatic cells by ectopic expression of defined transcription factors. Application of this approach in human cells may have enormous potential to generate patient-specific pluripotent stem cells. However, traditional methods of reprogramming in human somatic cells involve the use of oncogenes c-MYC and KLF4, which are not applicable to clinical translation. In the present study, we investigated whether human fetal gut mesentery-derived cells (hGMDCs) could be successfully reprogrammed into induced pluripotent stem (iPS) cells by OCT4, SOX2, and NANOG alone. We used lentiviruses to express OCT4, SOX2, NANOG, in hGMDCs, then generated iPS cells that were identified by morphology, presence of pluripotency markers, global gene expression profile, DNA methylation status, capacity to form embryoid bodies (EBs), and terotoma formation. iPS cells resulting from hGMDCs were similar to human embryonic stem (ES) cells in morphology, proliferation, surface markers, gene expression, and epigenetic status of pluripotent cell-specific genes. Furthermore, these cells were able to differentiate into cell types of all three germ layers both in vitro and in vivo, as shown by EB and teratoma formation assays. DNA fingerprinting showed that the human iPS cells were derived from the donor cells, and are not a result of contamination. Our results provide proof that hGMDCs can be reprogrammed into pluripotent cells by ectopic expression of three factors (OCT4, SOX2, and NANOG) without the use of oncogenes c-MYC and KLF4.
- SourceAvailable from: Fabienne Archer
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- "While OCT4, a member of the POU (Pit-Oct-Unc) transcription factor family, is essential for the maintenance of self-renewal capacities, NANOG (a downstream target of OCT4) contributes to the cell fate determination of pluripotent cells during embryogenesis [36,37]. OCT4 and NANOG are among the few key factors that enable the reprogramming of adult somatic cells into pluripotent stem cells [38-42]. Interestingly, induced pluripotent cells (iPSCs) generated from sheep fibroblasts have recently been demonstrated to exhibit an embryonic stem cell-like morphology and to express OCT4 and NANOG among other intracellular and surface markers associated with undifferentiated cells, as previously demonstrated in humans and mice . "
ABSTRACT: Airways progenitors may be involved in embryogenesis and lung repair. The characterization of these important populations may enable development of new therapeutics to treat acute or chronic lung disease. In this study, we aimed to establish the presence of bronchioloalveolar progenitors in ovine lungs and to characterize their potential to differentiate into specialized cells. Lung cells were studied using immunohistochemistry on frozen sections of the lung. Immunocytochemistry and flow cytometry were conducted on ex-vivo derived pulmonary cells. The bronchioloalveolar progenitors were identified by their co-expression of CCSP, SP-C and CD34. A minor population of CD34pos/SP-Cpos/CCSPpos cells (0.33% +/- 0.31) was present ex vivo in cell suspensions from dissociated lungs. Using CD34 magnetic positive-cell sorting, undifferentiated SP-Cpos/CCSPpos cells were purified (>80%) and maintained in culture. Using synthetic media and various extracellular matrices, SP-Cpos/CCSPpos cells differentiated into either club cells (formerly named Clara cells) or alveolar epithelial type-II cells. Furthermore, these ex vivo and in vitro derived bronchioloalveolar progenitors expressed NANOG, OCT4 and BMI1, specifically described in progenitors or stem cells, and during lung development. We report for the first time in a large animal the existence of bronchioloalveolar progenitors with dual differentiation potential and the expression of specialized genes. These newly described cell population in sheep could be implicated in regeneration of the lung following lesions or in development of diseases such as cancers.BMC Veterinary Research 11/2013; 9(1):224. DOI:10.1186/1746-6148-9-224 · 1.78 Impact Factor
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- "The difficulties associated with the use of embryonic stem cells can be overcome by the use of induced pluripotent stem (iPS) cells. iPS cells were originally derived from adult fibroblasts by the introduction and expression of four genes, namely OCT3/4, SOX2, KLF4 and c-MYC . Since iPS cells derived by reprogramming adult cells using the oncogenes KLF4 and c-MYC, which when expressed can lead to the development of teratomas, a number of investigators have been successful at reprogramming adult cells into iPS cells using other gene sets [189, 190]. "
ABSTRACT: Retinal degenerations encompass a large number of diseases in which the retina and associated retinal pigment epithelial (RPE) cells progressively degenerate leading to severe visual disorders or blindness. Retinal degenerations can be divided into two groups, a group in which the defect has been linked to a specific gene and a second group that has a complex etiology that includes environmental and genetic influences. The first group encompasses a number of relatively rare diseases with the most prevalent being Retinitis pigmentosa that affects approximately 1 million individuals worldwide. Attempts have been made to correct the defective gene by transfecting the appropriate cells with the wild-type gene and while these attempts have been successful in animal models, human gene therapy for these inherited retinal degenerations has only begun recently and the results are promising. To the second group belong glaucoma, age-related macular degeneration (AMD) and diabetic retinopathy (DR). These retinal degenerations have a genetic component since they occur more often in families with affected probands but they are also linked to environmental factors, specifically elevated intraocular pressure, age and high blood sugar levels respectively. The economic and medical impact of these three diseases can be assessed by the number of individuals affected; AMD affects over 30 million, DR over 40 million and glaucoma over 65 million individuals worldwide. The basic defect in these diseases appears to be the relative lack of a neurogenic environment; the neovascularization that often accompanies these diseases has suggested that a decrease in pigment epithelium-derived factor (PEDF), at least in part, may be responsible for the neurodegeneration since PEDF is not only an effective neurogenic and neuroprotective agent but also a potent inhibitor of neovascularization. In the last few years inhibitors of vascularization, especially antibodies against vascular endothelial cell growth factors (VEGF), have been used to prevent the neovascularization that accompanies AMD and DR resulting in the amelioration of vision in a significant number of patients. In animal models it has been shown that transfection of RPE cells with the gene for PEDF and other growth factors can prevent or slow degeneration. A limited number of studies in humans have also shown that transfection of RPE cells in vivo with the gene for PEDF is effective in preventing degeneration and restore vision. Most of these studies have used virally mediated gene delivery with all its accompanying side effects and have not been widely used. New techniques using non-viral protocols that allow efficient delivery and permanent integration of the transgene into the host cell genome offer novel opportunities for effective treatment of retinal degenerations.Current Genomics 08/2012; 13(5):350-362. DOI:10.2174/138920212801619214 · 2.34 Impact Factor
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- "EXPRESSION DURING LIMB REGENERATION Limb regeneration in Xenopus involves numerous signaling pathways (Beck et al., 2009), including BMP (Beck et al., 2006), Fgf (Yokoyama et al., 2000), and Wnt (Kawakami et al., 2006). Regulation of Sall4 expression during reprogramming and maintenance of pluripotency in ESC and somatic stem cells and probably during patterning also involves numerous context-dependent signaling pathways (Li et al., 2010). For example, in Drosophila melanogaster spalt/Sal expression is regulated during embryogenesis by Shh, dpp (BMP), and EGF signaling pathways, depending on the context (de Celis and Barrio, 2009). "
ABSTRACT: A central feature of epimorphic regeneration during amphibian limb regeneration is cellular dedifferentiation. Two questions are discussed. First, what is the origin and nature of the soluble factors involved in triggering local cellular and tissue dedifferentiation? Secondly, what role does the key stem cell transcription factor Sall4 play in reprogramming gene expression during dedifferentiation? The pattern of Sall4 expression during Xenopus hindlimb regeneration is consistent with the hypothesis that Sall4 plays a role in dedifferentiation (reprogramming) and in maintaining limb blastema cells in an undifferentiated state. Sall4 is involved in maintenance of ESC pluripotency, is a major repressor of differentiation, plays a major role in reprogramming differentiated cells into iPSCs, and is a component of the stemness regulatory circuit of pluripotent ESCs and iPSCs. These functions suggest Sall4 as an excellent candidate to regulate reprogramming events that produce and maintain dedifferentiated blastema cells required for epimorphic regeneration.Developmental Dynamics 05/2011; 240(5):979-89. DOI:10.1002/dvdy.22554 · 2.38 Impact Factor