Generation of Human-Induced Pluripotent Stem Cells by a Nonintegrating RNA Sendai Virus Vector in Feeder-Free or Xeno-Free Conditions

MacArthur, Chad C.; Fontes, Andrew; Ravinder, Namritha; Kuninger, David; Kaur, Jasmeet; Bailey, Matthew; Taliana, Antje; Vemuri, Mohan C.; Lieu, Pauline T.

doi:10.1155/2012/564612

Cited by 75 publications

(53 citation statements)

References 29 publications

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“…SeV reprogramming of human PGCs was performed, as previously described [45,46], using a Cytotune reprogramming kit (Lifetech) according to the manufacturer's instructions. The day before transduction, *100,000 PGCs were plated in PGCIII medium in a 6-well plate, and on day 0, the cells were incubated with virus at a MOI of 3 for 24 h. Afterward, the media was changed, and the cells fed every other day.…”

Section: Sev Reprogrammingmentioning

confidence: 99%

Direct Reprogramming of Human Primordial Germ Cells into Induced Pluripotent Stem Cells: Efficient Generation of Genetically Engineered Germ Cells

Bazley

Liu

Yuan

et al. 2015

Stem Cells and Development

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Primordial germ cells (PGCs) share many properties with embryonic stem cells (ESCs) and innately express several key pluripotency-controlling factors, including OCT4, NANOG, and LIN28. Therefore, PGCs may provide a simple and efficient model for studying somatic cell reprogramming to induced pluripotent stem cells (iPSCs), especially in determining the regulatory mechanisms that fundamentally define pluripotency. Here, we report a novel model of PGC reprogramming to generate iPSCs via transfection with SOX2 and OCT4 using integrative lentiviral. We also show the feasibility of using nonintegrative approaches for generating iPSC from PGCs using only these two factors. We show that human PGCs express endogenous levels of KLF4 and C-MYC protein at levels similar to embryonic germ cells (EGCs) but lower levels of SOX2 and OCT4. Transfection with both SOX2 and OCT4 together was required to induce PGCs to a pluripotent state at an efficiency of 1.71%, and the further addition of C-MYC increased the efficiency to 2.33%. Immunohistochemical analyses of the SOderived PGC-iPSCs revealed that these cells were more similar to ESCs than EGCs regarding both colony morphology and molecular characterization. Although leukemia inhibitory factor (LIF) was not required for the generation of PGC-iPSCs like EGCs, the presence of LIF combined with ectopic exposure to C-MYC yielded higher efficiencies. Additionally, the SO-derived PGC-iPSCs exhibited differentiation into representative cell types from all three germ layers in vitro and successfully formed teratomas in vivo. Several lines were generated that were karyotypically stable for up to 24 subcultures. Their derivation efficiency and survival in culture significantly supersedes that of EGCs, demonstrating their utility as a powerful model for studying factors regulating pluripotency in future studies.

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Section: Sev Reprogrammingmentioning

confidence: 99%

Direct Reprogramming of Human Primordial Germ Cells into Induced Pluripotent Stem Cells: Efficient Generation of Genetically Engineered Germ Cells

Bazley

Liu

Yuan

et al. 2015

Stem Cells and Development

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“…Sun et al (21) generated human iPSCs from adult human adipose stem cells, and Beltrão-Braga et al (29) subsequently derived iPSCs from human immature dental pulp stem cells without feeder cells. Recently, Macarthur et al (30) suggested that human neonatal fibroblast cells were able to be reprogrammed by transcription factors using a xeno-free culture system.…”

Section: R E T R a C T E Dmentioning

confidence: 99%

Generation of induced pluripotent stem cells using skin fibroblasts from patients with myocardial infarction under feeder-free conditions

Cheng

Tian

et al. 2014

Molecular Medicine Reports

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Abstract. Myocardial infarction (MI) is an increasing medical problem; however, its pathogenesis has yet to be elucidated and more effective treatment strategies are required. Induced pluripotent stem cells (iPSCs) were recently successfully generated using human somatic cells transfected with four transcription factors. The present study aimed to generate iPSCs from cells from patients with myocardial infarction. Six patients who had been diagnosed with myocardial infarction were enrolled in this study. The fibroblast cells from the biopsied skin were reprogrammed using octamer-binding transcription factor 4 (Oct-4), SRY-related HMG-box gene 2 (Sox-2), Kruppel-like factor 4 (Klf-4) and cellular myelocytomatosis oncogene (c-Myc) transcription factors. The generated cells were identified by karyotyping, in vitro and in vivo differentiation ability and staining for specific markers. These human MI-iPSCs expressed pluripotent genes and cell surface markers, and exhibited normal proliferation. The iPSCs also showed in vivo and in vitro differentiation ability, as indicated by teratoma and embryoid body formation, respectively. Moreover, the iPSCs differentiated into cardiomyocytes and neuronal cells. In conclusion, human iPSCs were successfully generated from skin fibroblasts from patients with MI under feeder-independent conditions, which increases their potential suitability for clinical applications. These results may encourage further study of MI pathogenesis and facilitate the development of safe downstream clinical applications of iPSC-based cell therapies.

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“…Therefore the risks of tumorigenesis can be avoided at the step of reprogramming factor induction [20][21][22][23][24] . Additionally, the feeder-free conditions, which use a combination of defined culture medium and matrigel instead of a feeder layer, make it possible to minimize the potential risks of exposure to unknown exogenous factors.…”

Section: Discussionmentioning

confidence: 99%

Generation of Induced Pluripotent Stem Cells from Human Peripheral T Cells Using Sendai Virus in Feeder-free Conditions

Kishino¹,

Seki

Yuasa³

et al. 2015

JoVE

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Recently, iPSCs have attracted attention as a new source of cells for regenerative therapies. Although the initial method for generating iPSCs relied on dermal fibroblasts obtained by invasive biopsy and retroviral genomic insertion of transgenes, there have been many efforts to avoid these disadvantages. Human peripheral T cells are a unique cell source for generating iPSCs. iPSCs derived from T cells contain rearrangements of the T cell receptor (TCR) genes and are a source of antigen-specific T cells. Additionally, T cell receptor rearrangement in the genome has the potential to label individual cell lines and distinguish between transplanted and donor cells. For safe clinical application of iPSCs, it is important to minimize the risk of exposing newly generated iPSCs to harmful agents. Although fetal bovine serum and feeder cells have been essential for pluripotent stem cell culture, it is preferable to remove them from the culture system to reduce the risk of unpredictable pathogenicity. To address this, we have established a protocol for generating iPSCs from human peripheral T cells using Sendai virus to reduce the risk of exposing iPSCs to undefined pathogens. Although handling Sendai virus requires equipment with the appropriate biosafety level, Sendai virus infects activated T cells without genome insertion, yet with high efficiency. In this protocol, we demonstrate the generation of iPSCs from human peripheral T cells in feeder-free conditions using a combination of activated T cell culture and Sendai virus.

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Generation of Human-Induced Pluripotent Stem Cells by a Nonintegrating RNA Sendai Virus Vector in Feeder-Free or Xeno-Free Conditions

Cited by 75 publications

References 29 publications

Direct Reprogramming of Human Primordial Germ Cells into Induced Pluripotent Stem Cells: Efficient Generation of Genetically Engineered Germ Cells

Direct Reprogramming of Human Primordial Germ Cells into Induced Pluripotent Stem Cells: Efficient Generation of Genetically Engineered Germ Cells

Generation of induced pluripotent stem cells using skin fibroblasts from patients with myocardial infarction under feeder-free conditions

Generation of Induced Pluripotent Stem Cells from Human Peripheral T Cells Using Sendai Virus in Feeder-free Conditions

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