Noninvasive imaging of embryonic stem cell cultures by multiphoton microscopy reveals the significance of collagen hydrogel preparation parameters

2016 ◽  
Vol 8 (2) ◽  
pp. 280-294
Author(s):  
Xuye Lang ◽  
Matthew Spousta ◽  
Yu Jer Hwang ◽  
Julia G. Lyubovitsky
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Embryonic Stem Cells ◽  
Embryonic Stem Cell ◽  
Cell Cultures ◽  
Noninvasive Imaging ◽  
Multiphoton Microscopy ◽  
Embryonic Stem ◽  
Collagen Hydrogel

Imaging discovers implications for impaired collagen and other biomaterials' functions during culturing embryonic stem cells in and on them.

scholarly journals Similar Population of CD133+ and DDX4+ VSEL-Like Stem Cells Sorted from Human Embryonic Stem Cell, Ovarian, and Ovarian Cancer Ascites Cell Cultures: The Real Embryonic Stem Cells?

Cells ◽  
2019 ◽  
Vol 8 (7) ◽  
pp. 706 ◽  
Author(s):  
Irma Virant-Klun ◽  
Petra Skerl ◽  
Srdjan Novakovic ◽  
Eda Vrtacnik-Bokal ◽  
Spela Smrkolj
Keyword(s):  
Ovarian Cancer ◽  
Stem Cells ◽  
Stem Cell ◽  
Embryonic Stem Cells ◽  
Embryonic Stem Cell ◽  
Cell Cultures ◽  
Human Embryonic Stem Cell ◽  
Embryonic Stem ◽  
Recurrent Ovarian Cancer

A population of small stem cells with diameters of up to 5 μm resembling very small embryonic-like stem cells (VSELs) were sorted from human embryonic stem cell (hESC) cultures using magnetic-activated cell sorting (MACS) based on the expression of a stem-cell-related marker prominin-1 (CD133). These VSEL-like stem cells had nuclei that almost filled the whole cell volume and expressed stem-cell-related markers (CD133, SSEA-4) and markers of germinal lineage (DDX4/VASA, PRDM14). They were comparable to similar populations of small stem cells sorted from cell cultures of normal ovaries and were the predominant cells in ascites of recurrent ovarian cancer. The sorted populations of CD133+ VSEL-like stem cells were quiescent in vitro, except for ascites, and were highly activated after exposure to valproic acid and follicle-stimulating hormone (FSH), indicating a new tool to study these cells in vitro. These VSEL-like stem cells spontaneously formed clusters resembling tumour-like structures or grew into larger, oocyte-like cells and were differentiated in vitro into adipogenic, osteogenic and neural lineages after sorting. We propose the population of VSEL-like stem cells from hESC cultures as potential original embryonic stem cells, which are present in the human embryo, persist in adult human ovaries from the embryonic period of life and are involved in cancer manifestation.

Effect of NANOG overexpression on porcine embryonic development and pluripotent embryonic stem cell formation in vitro

Zygote ◽  
2021 ◽  
pp. 1-6
Author(s):  
Gerelchimeg Bou ◽  
Shimeng Guo ◽  
Jia Guo ◽  
Zhuang Chai ◽  
Jianchao Zhao ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Embryonic Stem Cells ◽  
Embryonic Stem Cell ◽  
Embryonic Stem Cell Line ◽  
Embryonic Stem ◽  
Cell Clones ◽  
Morula Stage ◽  
Pluripotent Embryonic Stem Cell

Summary The efficiency of establishing pig pluripotent embryonic stem cell clones from blastocysts is still low. The transcription factor Nanog plays an important role in maintaining the pluripotency of mouse and human embryonic stem cells. Adequate activation of Nanog has been reported to increase the efficiency of establishing mouse embryonic stem cells from 3.5 day embryos. In mouse, Nanog starts to be strongly expressed as early as the morula stage, whereas in porcine NANOG starts to be strongly expressed by the late blastocyst stage. Therefore, here we investigated both the effect of expressing NANOG on porcine embryos early from the morula stage and the efficiency of porcine pluripotent embryonic stem cell clone formation. Compared with intact porcine embryos, NANOG overexpression induced a lower blastocyst rate, and did not show any advantages for embryo development and pluripotent embryonic stem cell line formation. These results indicated that, although NANOG is important pluripotent factor, NANOG overexpression is unnecessary for the initial formation of porcine pluripotent embryonic stem cell clones in vitro.

scholarly journals Induced Pluripotent Stem Cells: Hope in the Treatment of Diseases, including Muscular Dystrophies

2020 ◽  
Vol 21 (15) ◽  
pp. 5467
Author(s):  
Daniela Gois Beghini ◽  
Samuel Iwao Horita ◽  
Cynthia Machado Cascabulho ◽  
Luiz Anastácio Alves ◽  
Andrea Henriques-Pons
Keyword(s):  
Stem Cells ◽  
Clinical Trials ◽  
Stem Cell ◽  
Embryonic Stem Cells ◽  
Embryonic Stem Cell ◽  
Embryonic Stem ◽  
Ips Cells ◽  
High Plasticity ◽  
Cell Based Therapy ◽  
Induced Pluripotent

Induced pluripotent stem (iPS) cells are laboratory-produced cells that combine the biological advantages of somatic adult and stem cells for cell-based therapy. The reprogramming of cells, such as fibroblasts, to an embryonic stem cell-like state is done by the ectopic expression of transcription factors responsible for generating embryonic stem cell properties. These primary factors are octamer-binding transcription factor 4 (Oct3/4), sex-determining region Y-box 2 (Sox2), Krüppel-like factor 4 (Klf4), and the proto-oncogene protein homolog of avian myelocytomatosis (c-Myc). The somatic cells can be easily obtained from the patient who will be subjected to cellular therapy and be reprogrammed to acquire the necessary high plasticity of embryonic stem cells. These cells have no ethical limitations involved, as in the case of embryonic stem cells, and display minimal immunological rejection risks after transplant. Currently, several clinical trials are in progress, most of them in phase I or II. Still, some inherent risks, such as chromosomal instability, insertional tumors, and teratoma formation, must be overcome to reach full clinical translation. However, with the clinical trials and extensive basic research studying the biology of these cells, a promising future for human cell-based therapies using iPS cells seems to be increasingly clear and close.

scholarly journals The action behind the words: embryonic stem cell research marches on

2006 ◽  
Vol 174 (6) ◽  
pp. 743-746 ◽  
Author(s):  
Mitch Leslie
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Embryonic Stem Cells ◽  
Embryonic Stem Cell ◽  
Stem Cell Research ◽  
Embryonic Stem ◽  
Embryonic Stem Cell Research ◽  
Medical Resource ◽  
Stem Cell Science

Talk of policy has dominated talk of science for those interested in embryonic stem cell science. But research is continuing, and the advances are making clear why embryonic stem cells are such an important scientific and medical resource.

scholarly journals Controlling Embryonic Stem Cell Growth and Differentiation by Automation

2012 ◽  
Vol 17 (9) ◽  
pp. 1171-1179 ◽  
Author(s):  
Michael P. Kowalski ◽  
Amy Yoder ◽  
Li Liu ◽  
Laura Pajak
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Drug Discovery ◽  
Embryonic Stem Cells ◽  
Embryonic Stem Cell ◽  
High Throughput ◽  
Embryoid Body ◽  
Embryonic Stem ◽  
Basic Research ◽  
Complex Biological Process

Despite significant use in basic research, embryonic stem cells have just begun to be used in the drug discovery process. Barriers to the adoption of embryonic stem cells in drug discovery include the difficulty in growing cells and inconsistent differentiation to the desired cellular phenotype. Embryonic stem cell cultures require consistent and frequent handling to maintain the cells in a pluripotent state. In addition, the preferred hanging drop method of embryoid body (EB) differentiation is not amenable to high-throughput methods, and suspension cultures of EBs show a high degree of variability. Murine embryonic stem cells passaged on an automated platform maintained ≥90% viability and pluripotency. We also developed a method of EB formation using 384-well microplates that form a single EB per well, with excellent uniformity across EBs. This format facilitated high-throughput differentiation and enabled screens to optimize directed differentiation into a desired cell type. Using this approach, we identified conditions that enhanced cardiomyocyte differentiation sevenfold. This optimized differentiation method showed excellent consistency for such a complex biological process. This automated approach to embryonic stem cell handling and differentiation can provide the high and consistent yields of differentiated cell types required for basic research, compound screens, and toxicity studies.

scholarly journals Characterization of a negative retinoic acid response element in the murine Oct4 promoter.

1994 ◽  
Vol 14 (2) ◽  
pp. 1122-1136 ◽  
Author(s):  
J Schoorlemmer ◽  
A van Puijenbroek ◽  
M van Den Eijnden ◽  
L Jonk ◽  
C Pals ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Retinoic Acid ◽  
Embryonic Stem Cells ◽  
Embryonic Stem Cell ◽  
Embryonic Stem ◽  
Negative Regulation ◽  
Hormone Response ◽  
Response Element ◽  
Hormone Response Element

Expression of Oct4 in embryonic stem cells is controlled by a distal upstream stem cell-specific enhancer that is deactivated during retinoic acid (RA)-induced differentiation by an indirect mechanism not involving binding of RA receptors (H. Okazawa, K. Okamoto, F. Ishino, T. Ishino-Kaneko, S. Takeda, Y. Toyoda, M. Muramatsu, and H. Hamada, EMBO J. 10:2997-3005, 1991). Here we report that in RA-treated P19 embryonal carcinoma cells the Oct4 promoter is also subject to negative regulation by RA. The minimal Oct4 promoter sequence mediating repression consists of a promoter-proximal sequence containing a GC-rich SP1 consensus-like sequence and several hormone response element half-sites that can be arranged into direct repeats with different spacing. The GC box binds a nuclear factor that is invariably present in undifferentiated and RA-treated differentiated P19 cells. By contrast, the hormone response element-containing sequence binds factors that are induced following RA treatment. Mutational analysis and competition experiments show that the functional entity binding the RA-induced factor is a direct repeat sequence with a spacing of one nucleotide, previously shown to be a binding site for COUP transcription factors (COUP-TFs). Cotransfected orphan receptors COUP-TF1, ARP-1, and EAR-2 were able to repress the activity of Oct4 promoter-driven reporters in P19 EC cells, albeit with different efficiencies. Furthermore, the negative transcriptional effect of COUP-TFs is dominant over the activating effect of the Oct4 embryonic stem cell-specific enhancer. These results show that negative regulation of Oct4 expression during RA-induced differentiation of embryonic stem cells is controlled by two different mechanisms, including deactivation of the embryonic stem cell-specific enhancer and promoter silencing by orphan nuclear hormone receptors.

P–797 A novel method for establishing human embryonic stem cells independent of feeder cells

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
B Cai
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Embryonic Stem Cells ◽  
Embryonic Stem Cell ◽  
Success Rate ◽  
Cell Lines ◽  
Human Embryonic Stem Cells ◽  
Embryonic Stem ◽  
Feeder Cells ◽  
Novel Method

Abstract Study question Is there a efficient establishing method of human embryonic stem cells directly from the human blastocysts independent of feeder cells? Summary answer We established a novel method of generating human embryonic stem cells directly from human blastocysts independent of feeder layer cells. What is known already Establishing embryonic stem cells lines mainly needed to coculture ICM clumps with feeder cells (like mouse or human fibroblasts) ,this brought in potential heterogeneous pollution.Although there had be some reports about generating human ESCs independent of feeder cells,but the efficiency was low and conditioned medium were unstable and also had the biological contamination. Study design, size, duration We used ten day5/6 donated human blastocysts from our reproductive center ,most of them were genetically diseased embryos with abnormal PGT diagnosis.After establishing ESCs procedure , all the cell lines were identified with pluripotency and differentiation potential tests.The success rate of system was calculated and compared with the conventional methods. Participants/materials, setting, methods In brief, ICM clumps were separated mechanically by using a micromanipulation system,and then transferred to a 30ul mTESR plus culture media drop pretreated with the geltrex (1:100 dilution) matrix and oxygen concentration was 5%. When cells attached and migrated,we also used laser to destroy the remaining trophoblast cells.About 10 days,the typical ES clone can be mechanically passaged and cells can be cultured in normal oxygen concentrations after passage 2. . Main results and the role of chance Using this method we had successfully established nine embryonic stem cell lines from donated human blastocysts ,the success rate was 90% (9/10). Each cell lines had passed the evaluation test of embryonic stem cell. When compared with the conventional feeder cells dependent method,our novol methods not only eliminated the pollution from heterogeneous cells,but also had higher success rate (90% vs 25%). Limitations, reasons for caution Due to the scarcity of donated human blastocysts, this experiment was a single-center experiment with small samples. Wider implications of the findings: We speculated that the batch differences of culture dishes, matrix and culture medium might affect the establish efficiency , and how to carry out a high level of quality control work might be the key factor to keep the system stable. Trial registration number basic research

Cloning Animals, Cells, and Genes

2017 ◽  
pp. 129-150
Author(s):  
Scott Gilbert
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Embryonic Stem Cells ◽  
Embryonic Stem Cell ◽  
New Technology ◽  
Embryonic Stem ◽  
The Body ◽  
Animal Cloning ◽  
Medical Task

Every technology has its history, and this chapter looks at the technologies of animal cloning and how they changed when it became apparent that one could perform some of the medical task of cloning with embryonic stem cells. These stem cells were difficult to obtain (and morally worrisome to many) and a new technology, induced pluripotential stem cells, enabled researchers to transform nearly any cell of the body into an embryonic stem cell. This has brought new worries concerning the ability to manipulate these cells to enhance a person’s capabilities.

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