Nuclear reprogramming in cell–free extracts

Methods for directly turning a somatic cell type into another type (a process referred to as transdifferentiation) would be beneficial for producing replacement cells for therapeutic applications. Adult stem cells have been shown to display a broader differentiation potential than anticipated and may contribute to tissues other than those in which they reside. In addition, novel transdifferentiation strategies are being developed. I report recent results on the functional reprogramming of a somatic cell using a nuclear and cytoplasmic extract derived from another somatic cell type. The reprogramming of 293T fibroblasts in an extract from T cells is evidenced by nuclear uptake and the assembly of transcription factors, induction of activity of a chromatin remodelling complex, changes in chromatin composition and activation of lymphoid cell–specific genes. The reprogrammed cells express T–cell–specific surface molecules and a complex regulatory function. Reprogramming cells in cell–free extracts may create possibilities for producing replacement cells for therapeutic applications. The system may also constitute a powerful tool to examine the mechanisms of nuclear reprogramming, at least as they occur in vitro .

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The Efficiency of Cell Fusion-Based Reprogramming Is Affected by the Somatic Cell Type and the In Vitro Age of Somatic Cells

Cellular Reprogramming ◽

10.1089/cell.2011.0002 ◽

2011 ◽

Vol 13 (4) ◽

pp. 331-344 ◽

Cited By ~ 12

Author(s):

Pollyanna Agnes Tat ◽

Huseyin Sumer ◽

Daniele Pralong ◽

Paul John Verma

Keyword(s):

Somatic Cell ◽

Cell Fusion ◽

Somatic Cells ◽

Cell Type ◽

Somatic Cell Type

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Adult stem cells and their trans-differentiation potential—perspectives and therapeutic applications

Journal of Molecular Medicine ◽

10.1007/s00109-008-0383-6 ◽

2008 ◽

Vol 86 (12) ◽

pp. 1301-1314 ◽

Cited By ~ 83

Author(s):

Sabine Hombach-Klonisch ◽

Soumya Panigrahi ◽

Iran Rashedi ◽

Anja Seifert ◽

Esteban Alberti ◽

...

Keyword(s):

Stem Cells ◽

Adult Stem Cells ◽

Differentiation Potential ◽

Therapeutic Applications

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Proliferation and differentiation potential of rat forebrain oligodendroglial progenitors both in vitro and in vivo

Development ◽

10.1242/dev.111.4.1061 ◽

1991 ◽

Vol 111 (4) ◽

pp. 1061-1080 ◽

Cited By ~ 7

Author(s):

R. Hardy ◽

R. Reynolds

Keyword(s):

Progenitor Cell ◽

Neonatal Rat ◽

Myelin Proteins ◽

Brdu Incorporation ◽

Cell Type ◽

Differentiation Potential ◽

Serum Free ◽

Rat Forebrain

We have followed the development of the O-2A progenitor cell from the neonatal rat forebrain, both in dissociated cell culture and in cryostat sections, using immunocytochemical techniques employing a panel of antibodies that recognise the cells at different stages of their development. This included the monoclonal antibody LB1, which binds to the surface ganglioside GD3 expressed on O-2A progenitor cells. In secondary cultures enriched for O-2A progenitors maintained in a serum-free chemically defined medium, a large proportion of the cells are primed to differentiate into oligodendroglia and go on to express the oligodendroglial specific surface glycolipid galactocerebroside (GC) and then the myelin proteins CNP and MBP. However, a significant proportion of immature bipolar GD3+ cells remained after 6 days in secondary culture. It appears that not all the O-2A progenitors in our cultures differentiate immediately and some cells remain in an undifferentiated state and divide to replenish progenitor numbers. We have also identified in our cultures a small apolar GD3- cell, which when isolated differentiated into a GD3+ bipolar O-2A progenitor cell. We have termed this cell type a preprogenitor. The differentiation of this cell type into O-2A progenitors may be the source of the immature GD3+ cells present at the later stages of our secondary cultures. The proliferative profile of the cultures was studied using 5′bromo-2-deoxyuridine (BrdU) incorporation as an index of mitosis. Only the immature, bipolar O-2A progenitors were seen to divide at any time in serum-free culture. Neither the more mature multipolar O-2A cells nor the oligodendroglia were seen to divide. The developmental profile of the O-2A cells in the rat forebrain in vivo showed a largely similar progression to that in culture, with a time lag of at least 6 days between GD3 expression and the onset of myelination. BrdU incorporation studies in vivo also showed that the GD3+ progenitor cell is mitotic whereas the GC(+)-expressing oligodendroglia is not. We have shown that there are several significant alterations in the timing of antigen expression in both O-2A progenitors and oligodendroglia in vitro compared to that seen in vivo.

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Osteogenic Potential of Multipotent Adult Progenitor Cells for Calvaria Bone Regeneration

Advances in Medicine ◽

10.1155/2016/2803081 ◽

2016 ◽

Vol 2016 ◽

pp. 1-11

Author(s):

Dong Joon Lee ◽

Yonsil Park ◽

Wei-Shou Hu ◽

Ching-Chang Ko

Keyword(s):

Bone Regeneration ◽

Progenitor Cells ◽

Adult Stem Cells ◽

Single Cells ◽

Osteogenic Potential ◽

Type I ◽

Differentiation Potential ◽

Multipotent Adult Progenitor Cells

Osteogenic cells derived from rat multipotent adult progenitor cells (rMAPCs) were investigated for their potential use in bone regeneration. rMAPCs are adult stem cells derived from bone marrow that have a high proliferation capacity and the differentiation potential to multiple lineages. They may also offer immunomodulatory properties favorable for applications for regenerative medicine. rMAPCs were cultivated as single cells or as 3D aggregates in osteogenic media for up to 38 days, and their differentiation to bone lineage was then assessed by immunostaining of osteocalcin and collagen type I and by mineralization assays. The capability of rMAPCs in facilitating bone regeneration was evaluatedin vivoby the direct implantation of multipotent adult progenitor cell (MAPC) aggregates in rat calvarial defects. Bone regeneration was examined radiographically, histologically, and histomorphometrically. Results showed that rMAPCs successfully differentiated into osteogenic lineage by demonstrating mineralized extracellular matrix formationin vitroand induced new bone formation by the effect of rMAPC aggregatesin vivo. These outcomes confirm that rMAPCs have a good osteogenic potential and provide insights into rMAPCs as a novel adult stem cell source for bone regeneration.

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Effect of somatic cell type on bovine embryonic development in co-culture

Theriogenology ◽

10.1016/0093-691x(91)90245-9 ◽

1991 ◽

Vol 35 (1) ◽

pp. 269 ◽

Cited By ~ 6

Author(s):

J.M. Scodras ◽

J.W. Pollard ◽

K.J. Betteridge

Keyword(s):

Embryonic Development ◽

Somatic Cell ◽

Cell Type ◽

Somatic Cell Type

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Metabolic programming of a Warburg effect-like phenotype in donor fibroblasts prior to somatic cell nuclear transfer

10.32469/10355/66732 ◽

2017 ◽

Author(s):

◽

Bethany Rae Mordhorst

Keyword(s):

Somatic Cell ◽

Somatic Cell Nuclear Transfer ◽

Nuclear Transfer ◽

Donor Cell ◽

Nuclear Reprogramming ◽

In Vitro Development ◽

Fetal Fibroblasts ◽

Pharmaceutical Agents ◽

Vitro Development

Gene edited pigs serve as excellent models for biomedicine and agriculture. Currently, the most efficient way to make a reliably-edited transgenic animal is through somatic cell nuclear transfer (SCNT) also known as cloning. This process involves using cells from a donor (which may have been gene edited) that are typically grown in culture and using their nuclear content to reconstruct a new zygote. To do this, the cell may be placed in the perivitelline space of an enucleated oocyte and activated artificially by a calcium-containing media and electrical pulse waves. While it is remarkable that this process works, it is highly inefficient. In pigs the success of transferred embryos becoming live born piglets is only 1-3%. The creation of more cloned pigs enables further study for the benefit of both A) biomedicine in the development of prognosis and treatments and B) agriculture, whether it be for disease resistance, feed efficiency, gas emissions, etc. Two decades of research has not drastically improved the cloning efficiency of most mammals. One of the main impediments to successful cloning is thought to be due to inefficient nuclear reprogramming and remodeling of the donor cell nucleus. In the following chapters we detail our efforts to improve nuclear reprogramming of porcine fetal fibroblasts by altering the metabolism to be more blastomere-like in nature. We used two methods to alter metabolism 1) pharmaceutical agents and 2) hypoxia. After treating donor cells both methods were used in nuclear transfer. Pharmaceutical agents did not improve in vitro development of gestational survival of clones. Hypoxia did improve in vitro development and we are currently awaiting results of gestation.

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3D printable Sodium alginate-Matrigel (SA-MA) hydrogel facilitated ectomesenchymal stem cells (EMSCs) neuron differentiation

Journal of Biomaterials Applications ◽

10.1177/0885328220961261 ◽

2020 ◽

Vol 35 (6) ◽

pp. 709-719 ◽

Cited By ~ 1

Author(s):

Yang Li ◽

Xia Cao ◽

Wenwen Deng ◽

Qingtong Yu ◽

Congyong Sun ◽

...

Keyword(s):

Stem Cells ◽

Sodium Alginate ◽

Neuronal Differentiation ◽

Adult Stem Cells ◽

Three Dimensional ◽

Cell Types ◽

Differentiation Potential ◽

Lineage Differentiation ◽

Cord Injury

Ectomesenchymal stem cells (EMSCs) are typical adult stem cells obtained from the cranial neural crest. They have the potential to differentiate into various cell types, such as osseous cells, neurons and glial cells. Three-dimensional (3 D) printing is a novel method to construct biological structures by rapid prototyping. Previously, our group reported on the stemness and multi-lineage differentiation potential of EMSCs on gels. However, the exploration of EMSCs in 3 D printing and then evaluation of the growth and neuronal differentiation of EMSCs on extruded 3 D printable hybrid hydrogels has not been reported. Therefore, the current study explored the novel hybrid Sodium alginate-Matrigel (SA-MA) hydrogel extruded 3 D printing to design an in vitro scaffold to promote the differentiation and growth of EMSCs. In addition, the physical properties of the hydrogel were characterized and its drug-releasing property determined. Notably, the results showed that the construct exhibited a sustain-released effect of growth factor BDNF in accordance with the Higuchi equation. Moreover, the cell survival rate on the 3 D printed scaffold was 88.22 ± 1.13% with higher neuronal differentiation efficiency compared with 2 D culture. Thus, SA-MA’s ability to enhanced EMSCs neuronal differentiation offers a new biomaterial for neurons regeneration in the treatment of spinal cord injury.

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Multipotent skin-derived precursors: adult neural crest-related precursors with therapeutic potential

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2006.2020 ◽

2007 ◽

Vol 363 (1489) ◽

pp. 185-198 ◽

Cited By ~ 96

Author(s):

Karl J.L Fernandes ◽

Jean G Toma ◽

Freda D Miller

Keyword(s):

Neural Crest ◽

Therapeutic Potential ◽

Precursor Cell ◽

Cell Type ◽

Differentiation Potential ◽

Therapeutic Implications ◽

Lineage Differentiation ◽

Mammalian Skin ◽

Surprising Finding

We previously made the surprising finding that cultures of multipotent precursors can be grown from the dermis of neonatal and adult mammalian skin. These skin-derived precursors (SKPs) display multi-lineage differentiation potential, producing both neural and mesodermal progeny in vitro , and are an apparently novel precursor cell type that is distinct from other known precursors within the skin. In this review, we begin by placing these findings within the context of the rapidly evolving stem cell field. We then describe our recent efforts focused on understanding the developmental biology of SKPs, discussing the idea that SKPs are neural crest-related precursors that (i) migrate into the skin during embryogenesis, (ii) persist within a specific dermal niche, and (iii) play a key role in the normal physiology, and potentially pathology, of the skin. We conclude by highlighting some of the therapeutic implications and unresolved questions raised by these studies.

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