Derivation and propagation of human embryonic stem cell lines from frozen embryos in an animal product–free environment

Stephenson, Emma; Jacquet, Laureen; Miere, Cristian; Wood, Victoria; Kadeva, Neli; Cornwell, Glenda; Codognotto, Stefano; Dajani, Yaser; Braude, Peter; Ilić, Duško

doi:10.1038/nprot.2012.080

Cited by 69 publications

(78 citation statements)

References 39 publications

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“…Other investigators have reported that recombinant vitronectin 12 , a hydrogel 13 , and a LN-511 fragment 14 can serve as alternatives to Matrigel or feeder cells for culturing of hES cells. However, none of the cell culture substrata allows clonal survival of hES cells without the use of apoptosis inhibitors [15][16][17] or chemically undefined substances 18,19 , and the cells are often passaged in small clumps that can contain a significant proportion of differentiated cells. Since the initial number of cells is low during derivation of new ES cell lines, absence of cell culture conditions that enable clonal survival also complicates the derivation procedure especially from single blastomeres.…”

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confidence: 99%

Clonal culturing of human embryonic stem cells on laminin-521/E-cadherin matrix in defined and xeno-free environment

et al. 2014

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Lack of robust methods for establishment and expansion of pluripotent human embryonic stem (hES) cells still hampers development of cell therapy. Laminins (LN) are a family of highly cell-type specific basement membrane proteins important for cell adhesion, differentiation, migration and phenotype stability. Here we produce and isolate a human recombinant LN-521 isoform and develop a cell culture matrix containing LN-521 and E-cadherin, which both localize to stem cell niches in vivo. This matrix allows clonal derivation, clonal survival and long-term self-renewal of hES cells under completely chemically defined and xeno-free conditions without ROCK inhibitors. Neither LN-521 nor E-cadherin alone enable clonal survival of hES cells. The LN-521/E-cadherin matrix allows hES cell line derivation from blastocyst inner cell mass and single blastomere cells without a need to destroy the embryo. This method can facilitate the generation of hES cell lines for development of different cell types for regenerative medicine purposes.

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confidence: 99%

Clonal culturing of human embryonic stem cells on laminin-521/E-cadherin matrix in defined and xeno-free environment

et al. 2014

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“…This process which utilizes the direct reprogramming from one somatic lineage (fibroblasts) to another (either endothelial or smooth muscle cells) by skipping pluripotency may be a way to obtain safe and suitable cells of interest. Indeed, several protocols have been published to describe the generation of stem cell lines from human embryos, iPS cells and adult stem cells [24][25][26] , all of which however still raise concerning issues with regards to their use in clinic. It is now known that PiPSC do not develop teratomas in SCID mice, thus eliminating the concern of tumor formation.…”

Section: Discussionmentioning

confidence: 99%

Generation and Grafting of Tissue-engineered Vessels in a Mouse Model

Wong

Hong

Karamariti

et al. 2015

JoVE

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The construction of vascular conduits is a fundamental strategy for surgical repair of damaged and injured vessels resulting from cardiovascular diseases. The current protocol presents an efficient and reproducible strategy in which functional tissue engineered vessel grafts can be generated using partially induced pluripotent stem cell (PiPSC) from human fibroblasts. We designed a decellularized vessel scaffold bioreactor, which closely mimics the matrix protein structure and blood flow that exists within a native vessel, for seeding of PiPSC-endothelial cells or smooth muscle cells prior to grafting into mice. This approach was demonstrated to be advantageous because immune-deficient mice engrafted with the PiPSC-derived grafts presented with markedly increased survival rate 3 weeks after surgery. This protocol represents a valuable tool for regenerative medicine, tissue engineering and potentially patient-specific cell-therapy in the near future. Video LinkThe video component of this article can be found at

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“…This is related to safety concerns about the possibility of introduction of undesired proteins and DNA of animal origin in the cell preparation that may interfere with the properties of the cells or may cause adverse reactions. New techniques for extraction and propagation of stem cells in culture have been developed that exclude any products of xenotic origin [202][203][204][205], but virtually all of the initially established cell lines had been grown in the presence of heterologous protein and/or heterologous cells.…”

Section: 3mentioning

confidence: 99%

We heart cultured hearts. A comparative review of methodologies for targeted differentiation and maintenance of cardiomyocytes derived from pluripotent and multipotent stem cells

Arabadjiev¹,

Petkova²,

Chakarov³

et al. 2014

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Citation: Arabadjiev A, Petkova R, Chakarov S, Pankov R, Zhelev N. We heart cultured hearts. A comparative review of methodologies for targeted differentiation and maintenance of cardiomyocytes derived from pluripotent and multipotent stem cells. AbstractHuman cell lines, including disease cell lines are often superior to routine animal models for the purposes of rapid and safe assessment of the effects of different agents that may modulate myocardial functioning under physiological and pathological conditions. There are several currently existing methodologies for derivation of cardiomyocyte-like cells by targeted differentiation from pluripotent cells and by reprogramming/ transdifferentiation from other types of cells (multipotent progenitors, somatic cells, etc). The present paper reviews the potential sources of cells capable of differentiation along the cardiomyocyte lineage; the existing methodologies for targeted differentiation, outlining the specificities of each method; and the markers for differentiation along the mesodermal and the cardiogenic lineage. The yield of robustly beating cells expressing cardio-specific proteins derived by any of the existing methods, however, still rarely exceeds 70-90 %, even with the newly developed approaches for increasing the differentiation capacity. There still is significant variance in the results obtained by different research groups and even between different experiments carried out in the same laboratory, with the same type of cells and same general type of protocol. Derivation of new lines of human pluripotent and multipotent stem cells according to standardised protocols and in completely defined; xeno-free conditions may increase the reliability and reproducibility of research and speed up the development of potential clinical applications.

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Derivation and propagation of human embryonic stem cell lines from frozen embryos in an animal product–free environment

Cited by 69 publications

References 39 publications

Clonal culturing of human embryonic stem cells on laminin-521/E-cadherin matrix in defined and xeno-free environment

Clonal culturing of human embryonic stem cells on laminin-521/E-cadherin matrix in defined and xeno-free environment

Generation and Grafting of Tissue-engineered Vessels in a Mouse Model

We heart cultured hearts. A comparative review of methodologies for targeted differentiation and maintenance of cardiomyocytes derived from pluripotent and multipotent stem cells

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