Reprogramming of human fibroblasts to pluripotent stem cells using mRNA of four transcription factors

Yakubov, Eduard; Rechavi, Gideon; Rozenblatt, Shmuel; Givol, David

doi:10.1016/j.bbrc.2010.02.150

Cited by 217 publications

(149 citation statements)

References 12 publications

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“…This method directly introduces synthetic mRNA into the host cell for the expression of reprogramming factors, thus eliminating the need of viral or DNA vectors. The successful generation of RNA-induced iPSCs from fibroblasts [12][13][14][15] has led to the belief that this strategy is the beginning of the new era of cell reprogramming [11]. This strategy could in principle be expanded to reprogram any type of cell as long as the genes that control the cell fate are identified.…”

Section: Introductionmentioning

confidence: 99%

Mouse Embryonic Stem Cells Have Underdeveloped Antiviral Mechanisms That Can Be Exploited for the Development of mRNA-Mediated Gene Expression Strategy

Wang

Teng

Spangler

et al. 2014

Stem Cells and Development

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We have recently reported that mouse embryonic stem cells (mESCs) are deficient in expressing type I interferons (IFN) when exposed to viral infection and double-stranded RNA. In this study, we extended our investigation and demonstrated that single-stranded RNA and protein-encoding mRNA can induce strong IFN expression and cytotoxicity in fibroblasts and epithelial cells, but none of the effects associated with these antiviral responses were observed in mESCs. Our results provided additional data to support the conclusion that mESCs are intrinsically deficient in antiviral responses. While our findings represent a novel feature of mESCs that in itself is important for understanding innate immunity development, we exploited this property to develop a novel mRNA-mediated gene expression cell model. Direct introduction of synthetic mRNA to express desired genes has been shown as an effective alternative to DNA/viral vector-based gene expression. However, a major biological challenge is that a synthetic mRNA is detected as a viral RNA analog by the host cell, resulting in a series of adverse effects associated with antiviral responses. We demonstrate that the lack of antiviral responses in mESCs effectively avoids this problem. mESCs can tolerate repeated transfection and effectively express proteins from their synthetic mRNA with expected biological functions, as demonstrated by the expression of green fluorescent protein and the transcription factor Etv2. Therefore, mRNA-based gene expression could be developed into a novel ESC differentiation strategy that avoids safety concerns associated with viral/DNA-based vectors in regenerative medicine.

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Section: Introductionmentioning

confidence: 99%

Mouse Embryonic Stem Cells Have Underdeveloped Antiviral Mechanisms That Can Be Exploited for the Development of mRNA-Mediated Gene Expression Strategy

Wang

Teng

Spangler

et al. 2014

Stem Cells and Development

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“…Yakubov et al were the first to propose a mRNA-based approach as a solution to minimize genome integration as well as to increase reprogramming efficiency. In 2010, they demonstrated that lipid-based mRNA encoding four reprogramming factors could be used to induce expression of pluripotency markers in human fibroblasts [140]. Unfortunately, this study was limited by the absence of pluripotency verification tests, leading to the question whether these iPSCs were able to functionally differentiate into each of the three germ layers.…”

Section: Current State Of Non-immunotherapy Related Mrna Applicationsmentioning

confidence: 99%

Evading innate immunity in nonviral mRNA delivery: don’t shoot the messenger

Devoldere

Dewitte

Smedt

2016

Drug Discovery Today

113

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Teaser:This review presents an overview of the immune-related hurdles that limit mRNA advance for non-immunotherapy related applications and suggest some promising methods to reduce this 'unwanted' innate immune response. Author photographs and biographiesJoke Devoldere (1990) Her project lies on the interface between material science and immunology as it aims to develop novel micro-and nanomaterials for the delivery of antigen-coding mRNA to induce antitumor immunity. With her research, she won several awards, among which the "Therapeutic use of microbubbles" oral presentation award (Rotterdam, The Netherlands) and the Jan Feijen poster prize at the 13 th European symposium on controlled drug delivery (Egmond aan Zee, The Netherlands). Evading innate immunity in non-viral mRNA delivery: don't shoot the messenger AbstractIn de field of non-viral gene therapy, in vitro transcribed (IVT) mRNA has emerged as a promising tool for the delivery of genetic information. Over the past few years it has become widely known the introduction of IVT mRNA into mammalian cells elicits an innate immune response which has favored mRNA use towards immunotherapeutic vaccination strategies. However, for non-immunotherapy related applications this intrinsic immune-stimulatory activity directly interferes with the aimed therapeutic outcome, as it can seriously compromise the expression of the desired protein. This review presents an overview of the immune-related obstacles that limit mRNA advance for non-immunotherapy related applications.

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“…mRNA synthesized in vitro from cDNA of the reprogramming factors was demonstrated to be successful in 2010. 30 This method utilized host cells translation machinery, although it requires five consecutive transfections to be successful. Protein delivery is an alternative to nucleic acid introduction.…”

Section: Technical Improvement and Remaining Challenges Ipsc Generatimentioning

confidence: 99%

A practical guide to induced pluripotent stem cell research using patient samples

Santostefano

Hamazaki

Biel

et al. 2015

Laboratory Investigation

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Approximately 3 years ago, we assessed how patient induced pluripotent stem cell (iPSC) research could potentially impact human pathobiology studies in the future. Since then, the field has grown considerably with numerous technical developments, and the idea of modeling diseases 'in a dish' is becoming increasingly popular in biomedical research. Likely, it is even acceptable to include patient iPSCs as one of the standard research tools for disease mechanism studies, just like knockout mice. However, as the field matures, we acknowledge there remain many practical limitations and obstacles for their genuine application to understand diseases, and accept that it has not been as straightforward to model disorders as initially proposed. A major practical challenge has been efficient direction of iPSC differentiation into desired lineages and preparation of the large numbers of specific cell types required for study. Another even larger obstacle is the limited value of in vitro outcomes, which often do not closely represent disease conditions. To overcome the latter issue, many new approaches are underway, including three-dimensional organoid cultures from iPSCs, xenotransplantation of human cells to animal models and in vitro interaction of multiple cell types derived from isogenic iPSCs. Here we summarize the areas where patient iPSC studies have provided truly valuable information beyond existing skepticism, discuss the desired technologies to overcome current limitations and include practical guidance for how to utilize the resources. Undoubtedly, these human patient cells are an asset for experimental pathology studies. The future rests on how wisely we use them.

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Reprogramming of human fibroblasts to pluripotent stem cells using mRNA of four transcription factors

Cited by 217 publications

References 12 publications

Mouse Embryonic Stem Cells Have Underdeveloped Antiviral Mechanisms That Can Be Exploited for the Development of mRNA-Mediated Gene Expression Strategy

Mouse Embryonic Stem Cells Have Underdeveloped Antiviral Mechanisms That Can Be Exploited for the Development of mRNA-Mediated Gene Expression Strategy

Evading innate immunity in nonviral mRNA delivery: don’t shoot the messenger

A practical guide to induced pluripotent stem cell research using patient samples

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