Publications (7)99.42 Total impact
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Article: Generation of induced pluripotent stem cells from human renal proximal tubular cells with only two transcription factors, OCT4 and SOX2.
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ABSTRACT: The tubular epithelium of the kidney is susceptible to injury from a number of different causes, including inflammatory and immune disorders, oxidative stress, and nephrotoxins, among others. Primary renal epithelial cells remain one of the few tools for studying the biochemical and physiological characteristics of the renal tubular system. Nevertheless, differentiated primary cells are not suitable for recapitulation of disease properties that might arise during embryonic kidney formation and further maturation. Thus, cellular systems resembling kidney characteristics are in urgent need to model disease as well as to establish reliable drug-testing platforms. Induced pluripotent stem cells (iPSCs) bear the capacity to differentiate into every cell lineage comprising the adult organism. Thus, iPSCs bring the possibility for recapitulating embryonic development by directed differentiation into specific lineages. iPSC differentiation ultimately allows for both disease modeling in vitro and the production of cellular products with potential for regenerative medicine. Here, we describe the rapid, reproducible, and highly efficient generation of iPSCs derived from endogenous kidney tubular renal epithelial cells with only two transcriptional factors, OCT4 and SOX2. Kidney-derived iPSCs may provide a reliable cellular platform for the development of kidney differentiation protocols allowing drug discovery studies and the study of kidney pathology.Journal of Biological Chemistry 05/2012; 287(29):24131-8. · 4.77 Impact Factor -
Article: Generation of feeder-free pig induced pluripotent stem cells without Pou5f1.
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ABSTRACT: The pig represents an ideal large-animal model, intermediate between rodents and humans, for the preclinical assessment of emerging cell therapies. As no validated pig embryonic stem (pES) cell lines have been derived so far, pig induced pluripotent stem cells (piPSCs) should offer an alternative source of undifferentiated cells to advance regenerative medicine research from bench to clinical trial. We report here for the first time the derivation of piPSCs from adult fibroblast with only three transcription factors: Sox2 (sex determining region Y-box 2), Klf4 (Krüppel-like factor 4), and c-Myc (avian myelocytomatosis viral oncogene homolog). We have been able to demonstrate that exogenous Pou5f1 (POU domain class 5 transcription factor 1; abbreviated as Octamer-4: Oct4) is dispensable to achieve and maintain pluripotency in the generation of piPSCs. To the best of our knowledge, this is also the first report of somatic reprogramming in any species without the overexpression, either directly or indirectly, of Oct4. Moreover, we were able to generate piPSCs without the use of feeder cells, approaching thus xeno-free conditions. Our work paves the way for the derivation of clinical grade piPSCs for regenerative medicine.Cell Transplantation 09/2011; 21(5):815-25. · 5.13 Impact Factor -
Article: Simple generation of human induced pluripotent stem cells using poly-beta-amino esters as the non-viral gene delivery system.
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ABSTRACT: Reprogramming of somatic cells to induced pluripotent stem (iPS) cells can be achieved by the delivery of a combination of transcription factors, including Oct4, Sox2, Klf4, and c-Myc. Retroviral and lentiviral vectors are commonly used to express these four reprogramming factors separately and obtain reprogrammed iPS cells. Although efficient and reproducible, these approaches involve the time-consuming and labor-intensive production of retroviral or lentiviral particles together with a high risk of working with potentially harmful viruses overexpressing potent oncogenes, such as c-Myc. Here, we describe a simple method to produce bona fide iPS cells from human fibroblasts using poly-β-amino esters as the transfection reagent for the delivery of a single CAG-driven polycistronic plasmid expressing Oct4, Sox2, Klf4, c-Myc, and a GFP reporter gene (OSKMG). We demonstrate for the first time that poly-β-amino esters can be used to deliver a single polycistronic reprogramming vector into human fibroblasts, achieving significantly higher transfection efficiency than with conventional transfection reagents. After a protocol of serial transfections using poly-β-amino esters, we report a simple methodology to generate human iPS cells from human fibroblasts avoiding the use of viral vectors.Journal of Biological Chemistry 02/2011; 286(14):12417-28. · 4.77 Impact Factor -
Article: Methods for making induced pluripotent stem cells: reprogramming à la carte.
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ABSTRACT: Pluripotent stem-cell lines can be obtained through the reprogramming of somatic cells from different tissues and species by ectopic expression of defined factors. In theory, these cells--known as induced pluripotent stem cells (iPSCs)--are suitable for various purposes, including disease modelling, autologous cell therapy, drug or toxicity screening and basic research. Recent methodological improvements are increasing the ease and efficiency of reprogramming, and reducing the genomic modifications required to complete the process. However, depending on the downstream applications, certain technologies have advantages over others. Here, we provide a comprehensive overview of the existing reprogramming approaches with the aim of providing readers with a better understanding of the reprogramming process and a basis for selecting the most suitable method for basic or clinical applications.Nature Reviews Genetics 02/2011; 12(4):231-42. · 38.08 Impact Factor -
Article: Generation of pig iPS cells: a model for cell therapy.
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ABSTRACT: Reprogramming of pig somatic cells to induced pluripotent stem cells provides a tremendous advance in the field of regenerative medicine since the pig represents an ideal large animal model for the preclinical testing of emerging cell therapies. However, the current generation of pig-induced pluripotent stem cells (piPSCs) require the use of time-consuming and laborious retroviral or lentiviral transduction approaches, in order to ectopically express the pluripotency-associated transcription factors Oct4, Sox2, Klf4 and c-Myc, in the presence of feeder cells. Here, we describe a simple method to produce piPSC with a single transfection of a CAG-driven polycistronic plasmid expressing Oct4, Sox2, Klf4, c-Myc and a green fluorescent protein (GFP) reporter gene, in gelatine-coated plates, with or without feeder cells. In our system, the derivation of piPSCs from adult pig ear fibroblasts on a gelatine coating showed a higher efficiency and rate of reprogramming when compared with three consecutive retroviral transductions of a similar polycistronic construct. Our piPSCs expressed the classical embryonic stem cell markers, exhibit a stable karyotype and formed teratomas. Moreover, we also developed a simple method to generate in vitro spontaneous beating cardiomiocyte-like cells from piPSCs. Overall, our preliminary results set the bases for the massive production of xeno-free and integration-free piPSCs and provide a powerful tool for the preclinical application of iPSC technology in a large animal setting.Journal of Cardiovascular Translational Research 11/2010; 4(2):121-30. · 2.61 Impact Factor -
Article: Reprogramming of human fibroblasts to induced pluripotent stem cells under xeno-free conditions.
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ABSTRACT: The availability of induced pluripotent stem cells (iPSCs) has created extraordinary opportunities for modeling and perhaps treating human disease. However, all reprogramming protocols used to date involve the use of products of animal origin. Here, we set out to develop a protocol to generate and maintain human iPSC that would be entirely devoid of xenobiotics. We first developed a xeno-free cell culture media that supported the long-term propagation of human embryonic stem cells (hESCs) to a similar extent as conventional media containing animal origin products or commercially available xeno-free medium. We also derived primary cultures of human dermal fibroblasts under strict xeno-free conditions (XF-HFF), and we show that they can be used as both the cell source for iPSC generation as well as autologous feeder cells to support their growth. We also replaced other reagents of animal origin (trypsin, gelatin, matrigel) with their recombinant equivalents. Finally, we used vesicular stomatitis virus G-pseudotyped retroviral particles expressing a polycistronic construct encoding Oct4, Sox2, Klf4, and GFP to reprogram XF-HFF cells under xeno-free conditions. A total of 10 xeno-free human iPSC lines were generated, which could be continuously passaged in xeno-free conditions and maintained characteristics indistinguishable from hESCs, including colony morphology and growth behavior, expression of pluripotency-associated markers, and pluripotent differentiation ability in vitro and in teratoma assays. Overall, the results presented here demonstrate that human iPSCs can be generated and maintained under strict xeno-free conditions and provide a path to good manufacturing practice (GMP) applicability that should facilitate the clinical translation of iPSC-based therapies.Stem Cells 11/2009; 28(1):36-44. · 7.78 Impact Factor -
Article: Disease-corrected haematopoietic progenitors from Fanconi anaemia induced pluripotent stem cells.
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ABSTRACT: The generation of induced pluripotent stem (iPS) cells has enabled the derivation of patient-specific pluripotent cells and provided valuable experimental platforms to model human disease. Patient-specific iPS cells are also thought to hold great therapeutic potential, although direct evidence for this is still lacking. Here we show that, on correction of the genetic defect, somatic cells from Fanconi anaemia patients can be reprogrammed to pluripotency to generate patient-specific iPS cells. These cell lines appear indistinguishable from human embryonic stem cells and iPS cells from healthy individuals. Most importantly, we show that corrected Fanconi-anaemia-specific iPS cells can give rise to haematopoietic progenitors of the myeloid and erythroid lineages that are phenotypically normal, that is, disease-free. These data offer proof-of-concept that iPS cell technology can be used for the generation of disease-corrected, patient-specific cells with potential value for cell therapy applications.Nature 06/2009; 460(7251):53-9. · 36.28 Impact Factor
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Top Journals
- Journal of Biological Chemistry (2)
- Nature Reviews Genetics (1)
- Nature (1)
- Stem Cells (1)
- Cell Transplantation (1)
Institutions
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2009–2010
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Center of Regenerative Medicine in Barcelona
Barcelona, Catalonia, Spain
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