Selection of Targeted Mutants from a Library of Randomly Mutagenized ES Cells

Organ or tissue transplantation is the preferred treatment for numerous diseases but is hindered by immunologic barriers. Genetically matched pluripotent embryonic stem cells generated via nuclear transfer (ntES cells) or parthenogenesis (pES cells) are possible sources of histocompatible cells and tissues. We have developed two ways of isolating pES cells that carry the full complement of major histocompatibility complex (MHC) antigens of the oocyte donors. One method entails activation of oocytes after blockade of karyokinesis in meiosis II, followed by selection of predominantly homozygous pES cells that have undergone recombination in their MHC antigen region to restore the heterozygous maternal MHC genotype (parthenote recombinant, or prES cells). The second method involves activation of immature oocytes after blockade of karyokinesis of meiosis I, followed by selection of predominantly heterozygous pES lines that retain the MHC genotype of the oocyte donor (parthenote clone recombinant, or pcrES cells). The cells are pluripotent by several criteria: teratoma formation, in vitro differentiation into hematopoietic elements, and high-level skin chimerism in blastocyst chimeras. Breeding of 8 founder females and examination of over 700 progeny failed to demonstrate germ line transmission of the pES cells. Injection of over 50 tetraploid embryos with these lines and embryo transfer have failed to support full gestational development. However, differentiated tissues from these pluripotent ES cells engraft when transplanted into genetically matched immunocompetent recipients, demonstrating that selected pES cells can serve as a source of histocompatible tissues for transplantation.

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Selection of ventricular‐like cardiomyocytes from ES cells in vitro

The FASEB Journal ◽

10.1096/fj.00-0002com ◽

2000 ◽

Vol 14 (15) ◽

pp. 2540-2548 ◽

Cited By ~ 210

Author(s):

M. MÜller ◽

B. K. Fleischmann ◽

S. Selbert ◽

G. J. Ji ◽

E. Endl ◽

...

Keyword(s):

Es Cells ◽

Selection Of

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Fully human agonist antibodies to TrkB using autocrine cell-based selection from a combinatorial antibody library

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1806660115 ◽

2018 ◽

Vol 115 (30) ◽

pp. E7023-E7032 ◽

Cited By ~ 13

Author(s):

Spyros Merkouris ◽

Yves-Alain Barde ◽

Kate E. Binley ◽

Nicholas D. Allen ◽

Alexey V. Stepanov ◽

...

Keyword(s):

Es Cells ◽

Drug Candidate ◽

Antibody Library ◽

Screening Assay ◽

Physiological Characterization ◽

Human Neurons ◽

Hydrophobic Molecule ◽

Human Es Cells ◽

Selection Of

The diverse physiological roles of the neurotrophin family have long prompted exploration of their potential as therapeutic agents for nerve injury and neurodegenerative diseases. To date, clinical trials of one family member, brain-derived neurotrophic factor (BDNF), have disappointingly failed to meet desired endpoints. Contributing to these failures is the fact that BDNF is pharmaceutically a nonideal biologic drug candidate. It is a highly charged, yet is a net hydrophobic molecule with a low molecular weight that confers a short t1/2 in man. To circumvent these shortcomings of BDNF as a drug candidate, we have employed a function-based cellular screening assay to select activating antibodies of the BDNF receptor TrkB from a combinatorial human short-chain variable fragment antibody library. We report here the successful selection of several potent TrkB agonist antibodies and detailed biochemical and physiological characterization of one such antibody, ZEB85. By using a human TrkB reporter cell line and BDNF-responsive GABAergic neurons derived from human ES cells, we demonstrate that ZEB85 is a full agonist of TrkB, comparable in potency to BDNF toward human neurons in activation of TrkB phosphorylation, canonical signal transduction, and mRNA transcriptional regulation.

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Localization of Liv2 as an Immature Hepatocyte Marker in EB Outgrowth

The Scientific World JOURNAL ◽

10.1100/tsw.2009.18 ◽

2009 ◽

Vol 9 ◽

pp. 190-199 ◽

Cited By ~ 5

Author(s):

Ikkei Takashimizu ◽

Yoshiki Tanaka ◽

Susumu Yoshie ◽

Yoshiya Kano ◽

Hinako Ichikawa ◽

...

Keyword(s):

Cell Membrane ◽

Es Cells ◽

Embryonic Stem ◽

Embryoid Bodies ◽

Surface Marker ◽

Neuronal Marker ◽

Mouse Es Cells ◽

Undifferentiated State ◽

Hanging Drop Method ◽

Selection Of

The objective of this study was to establish Liv2, a surface marker of mouse immature hepatocytes (hepatoblasts), as a selection tool for embryonic stem (ES) cell–derived immature hepatocytes by acquiring basic data on Liv2 in normal mouse embryos and by confirming Liv2 expression in mouse ES-derived cells. The estimated molecular weight of Liv2 was 4045 kDa, and immunoreactivity was definitively detected in the cell membrane of fetal hepatocytes on embryonic day (E) 9.5, declined gradually until E12.5, and subsequently became undetectable. Liv2 was localized on and close to the cell membrane. Embryoid bodies (EB) were formed from mouse ES cells whose undifferentiated state was confirmed with immunostaining of Nanog by the hanging drop method. A few Liv2-positive cells occurred as a cluster in EB outgrowth on day 7, but only some of these were albumin (ALB)-positive on day 13. These cells had the same pattern of immunoreactivity, i.e., localization on the cell membrane, as immature hepatocytes in the developing liver, although there were other types of cells with a different pattern of immunoreactivity that were seen only as a granular pattern in the cytoplasm and without ALB or the neuronal marker nestin. These results suggest that Liv2 may be useful as a surface marker for immature hepatocytes derived from ES cells. This application would allow for the sole selection of immature hepatocytes and provide a useful tool for regenerative medicine.

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The Process of Selection of Erythromycin-Resistant Mitochondria by Erythromycin in Paramecium

Journal of Cell Science ◽

10.1242/jcs.14.3.475 ◽

1974 ◽

Vol 14 (3) ◽

pp. 475-497

Author(s):

R. PERASSO

Keyword(s):

Es Cells ◽

Single Cells ◽

Mitochondrial Genomes ◽

Wild Type ◽

Erythromycin Resistance ◽

Mitochondrial Mutations ◽

Cellular Division ◽

Thin Sections ◽

Mitochondrial Population ◽

Selection Of

Mitochondrial mutations conferring erythromycin resistance (ER) are available in Paramecium and it is possible to obtain (by conjugation and cytoplasmic exchange) exconjugant cells containing a majority of wild-type erythromycin-sensitive (ES) mitochondria and a minority of ER ones. In the presence of erythromycin, such ‘mixed’ cells progressively become resistant. This process of acquisition of resistance has been studied cytologically (on thin sections of single cells) and genetically (by evaluating, on the basis of previous data, the proportion of ER/ES mitochondrial genomes at various times). While at early stages of the process of transformation the whole mitochondrial population appears rather homogeneous, at later stages, (i.e. when the cell has resumed growth in the antibiotic-containing medium) one finds, side by side, both resistant-looking mitochondria (structurally normal) and sensitive-looking ones, showing the typical alterations induced in Es cells by erythromycin. Conversely, a progressive decrease in the number of ES genomes can be demonstrated. The complete genetical and cytological transformation from erythromycin sensitivity to erythromycin resistance can occur after less than three fissions in erythromycin-containing medium. The results indicate that intensive selective multiplication of ER mitochondria occurs, under the pressure of erythromycin, virtually in the absence of cellular division. The possibility of dissociating mitochondrial division from cell division emphasizes the extent of mitochondrial autonomy.

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Cognitive gadgets and cognitive priors

Behavioral and Brain Sciences ◽

10.1017/s0140525x19000992 ◽

2019 ◽

Vol 42 ◽

Author(s):

Gian Domenico Iannetti ◽

Giorgio Vallortigara

Keyword(s):

Cognitive Neuroscience ◽

Selection Of

Abstract Some of the foundations of Heyes’ radical reasoning seem to be based on a fractional selection of available evidence. Using an ethological perspective, we argue against Heyes’ rapid dismissal of innate cognitive instincts. Heyes’ use of fMRI studies of literacy to claim that culture assembles pieces of mental technology seems an example of incorrect reverse inferences and overlap theories pervasive in cognitive neuroscience.

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Note on Selection of Coordinate Systems for Geodynamics

International Astronomical Union Colloquium ◽

10.1017/s0252921100051174 ◽

1975 ◽

Vol 26 ◽

pp. 395-407

Author(s):

S. Henriksen

Keyword(s):

Sea Level ◽

The Other ◽

Coordinate Systems ◽

Mean Sea Level ◽

The Earth ◽

The One ◽

Static Systems ◽

Selection Of

The first question to be answered, in seeking coordinate systems for geodynamics, is: what is geodynamics? The answer is, of course, that geodynamics is that part of geophysics which is concerned with movements of the Earth, as opposed to geostatics which is the physics of the stationary Earth. But as far as we know, there is no stationary Earth – epur sic monere. So geodynamics is actually coextensive with geophysics, and coordinate systems suitable for the one should be suitable for the other. At the present time, there are not many coordinate systems, if any, that can be identified with a static Earth. Certainly the only coordinate of aeronomic (atmospheric) interest is the height, and this is usually either as geodynamic height or as pressure. In oceanology, the most important coordinate is depth, and this, like heights in the atmosphere, is expressed as metric depth from mean sea level, as geodynamic depth, or as pressure. Only for the earth do we find “static” systems in use, ana even here there is real question as to whether the systems are dynamic or static. So it would seem that our answer to the question, of what kind, of coordinate systems are we seeking, must be that we are looking for the same systems as are used in geophysics, and these systems are dynamic in nature already – that is, their definition involvestime.

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