Optimization of Synthetic mRNA for Highly Efficient Translation and its Application in the Generation of Endothelial and Hematopoietic Cells from Human and Primate Pluripotent Stem Cells

Suknuntha, Kran; Tao, Lihong; Brok-Volchanskaya, Vera; D'Souza, Saritha S.; Kumar, Akhilesh; Slukvin, Igor I.

doi:10.1007/s12015-018-9805-1

Cited by 28 publications

(28 citation statements)

References 32 publications

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“…3B, fig. S5), which is consistent with previous reports (16) To further elucidate potential differences between h-iECs generated from our S1-modETV2, the S1-S2, and the early modETV2 protocols, we performed RNAseq analysis across multiple h-iECs samples generated from three independent h-iPSC lines using all three differentiation protocols.…”

Section: Time Of Etv2 Activation Affects the Transcriptional Profile supporting

confidence: 81%

“…Nevertheless, the efficiency of direct reprogramming somatic cells into ECs remains exceedingly low and achieving proper EC maturation requires long periods of time in culture. Alternatively, a few studies have recently proposed inducing ETV2 expression directly on h-iPSCs to induce EC differentiation (14)(15)(16). However, to date, methods have relied on early activation of ETV2 in the h-iPSCs, thus bypassing transition through an intermediate mesodermal stage.…”

Section: Introductionmentioning

confidence: 99%

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Robust differentiation of human pluripotent stem cells into endothelial cells via temporal modulation of ETV2 with modified mRNA

Wang

Lin

Hong

et al. 2020

Preprint

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Human induced pluripotent stem cell (h-iPSC)-derived endothelial cells (h-iECs) have become a valuable tool in regenerative medicine. However, current differentiation protocols remain inefficient and lack reliability. Here, we describe a method for rapid, consistent, and highly efficient generation of h-iECs. The protocol entails the delivery of modified mRNA encoding the transcription factor ETV2 at the intermediate mesodermal stage of differentiation. This approach reproducibly differentiated thirteen diverse h-iPSC lines into h-iECs with exceedingly high efficiency. In contrast, standard differentiation methods that relied on endogenous ETV2 were inefficient and notably inconsistent. Our h-iECs were functionally competent in many respects, including the ability to form perfused vascular networks in vivo. Importantly, timely activation of ETV2 was critical, and bypassing the mesodermal stage produced putative h-iECs with reduced expansion potential and inability to form functional vessels. Our protocol has broad applications and could reliably provide an unlimited number of h-iECs for vascular therapies.

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Section: Time Of Etv2 Activation Affects the Transcriptional Profile supporting

confidence: 81%

Section: Introductionmentioning

confidence: 99%

Robust differentiation of human pluripotent stem cells into endothelial cells via temporal modulation of ETV2 with modified mRNA

Wang

Lin

Hong

et al. 2020

Preprint

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“…The importance of incorporation of 5'-and 3'-UTRs has been noted during in vitro post-transcriptional regulation of gene expression [55]. The numerous roles that UTRs play include (i) regulation of mRNA export from the nucleus, (ii) regulation of translation efficiency [56], (iii) orchestration of subcellular localization [57], and (iv) mRNA stability [58]. Introduction of α-globin 3' end UTRs results in stabilization of mRNA, while the incorporation of beta-globin 5' end and 3' end UTRs leads to enhanced translational efficiency [59].…”

Section: ' and 3' Untranslated Regions (Utrs)mentioning

confidence: 99%

Opportunities and Challenges in the Delivery of mRNA-Based Vaccines

et al. 2020

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In the past few years, there has been increasing focus on the use of messenger RNA (mRNA) as a new therapeutic modality. Current clinical efforts encompassing mRNA-based drugs are directed toward infectious disease vaccines, cancer immunotherapies, therapeutic protein replacement therapies, and treatment of genetic diseases. However, challenges that impede the successful translation of these molecules into drugs are that (i) mRNA is a very large molecule, (ii) it is intrinsically unstable and prone to degradation by nucleases, and (iii) it activates the immune system. Although some of these challenges have been partially solved by means of chemical modification of the mRNA, intracellular delivery of mRNA still represents a major hurdle. The clinical translation of mRNA-based therapeutics requires delivery technologies that can ensure stabilization of mRNA under physiological conditions. Here, we (i) review opportunities and challenges in the delivery of mRNA-based therapeutics with a focus on non-viral delivery systems, (ii) present the clinical status of mRNA vaccines, and (iii) highlight perspectives on the future of this promising new type of medicine.

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“…127 hPSCs were also differentiated into typical endothelial phenotype by delivery of modRNA encoding ETV2. 128 However, the modRNA-based reprogramming protocol is complicated and laborious. 123 It requires multiple transfections in cell fate conversion.…”

Section: Genetically Modified Cellsmentioning

confidence: 99%

Vascular Tissue Engineering: Advanced Techniques and Gene Editing in Stem Cells for Graft Generation

Chen

Ugwu

Li³

et al. 2021

Tissue Engineering Part B: Reviews

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The common occurrence of cardiovascular diseases and the lack of proper autologous tissues prompt and promote the pressing development of tissue-engineered vascular grafts. Current progress on scaffold production, genetically modified cells and use of nanotechnology-based monitoring has considerably improved the long-term patency of engineered tissue grafts. However, challenges abound in the autologous materials and manipulation of genes and cells for tissue engineering. This review overviews current development in tissue-engineered vascular grafts and discusses recent improvements in scaffolding techniques as well as the efficiency of gene-editing tools and their ability to fill the existing gaps in stem-cell and regenerative therapies. Current advances in 3D-printing approaches for fabrication of engineered tissues are also reviewed together with specific biomaterials for vascular tissues. In addition, the natural and synthetic polymers that hold increasing significance for vascular tissue engineering are highlighted. Both animal models and nanotechnology-based monitoring are proposed for pre-clinical evaluation of engineered grafts in view of their historical significance in tissue engineering. The ultimate success of tissue regeneration, which is yet to be fully realized, depends on the optimal performance of culture systems, biomaterial constructs and stem cells in a suitable artificial physiological environment.

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Optimization of Synthetic mRNA for Highly Efficient Translation and its Application in the Generation of Endothelial and Hematopoietic Cells from Human and Primate Pluripotent Stem Cells

Cited by 28 publications

References 32 publications

Robust differentiation of human pluripotent stem cells into endothelial cells via temporal modulation of ETV2 with modified mRNA

Robust differentiation of human pluripotent stem cells into endothelial cells via temporal modulation of ETV2 with modified mRNA

Opportunities and Challenges in the Delivery of mRNA-Based Vaccines

Vascular Tissue Engineering: Advanced Techniques and Gene Editing in Stem Cells for Graft Generation

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