scholarly journals Morphokinetics of cloned mouse embryos treated with epigenetic drugs and blastocyst prediction

Reproduction ◽  
2016 ◽  
Vol 151 (3) ◽  
pp. 203-214 ◽  
Author(s):  
Anna Mallol ◽  
Laia Piqué ◽  
Josep Santaló ◽  
Elena Ibáñez
Keyword(s):  
Nuclear Transfer ◽  
Embryonic Stem ◽  
Time Lapse ◽  
Developmental Potential ◽  
Time Lapse Microscopy ◽  
Epigenetic Effects ◽  
Psammaplin A ◽  
Esc Derivation ◽  
Microscopy Analysis ◽  
Selection Of

Time-lapse monitoring of somatic cell nuclear transfer (SCNT) embryos may help to predict developmental success and increase birth and embryonic stem cells (ESC) derivation rates. Here, the development of ICSI fertilized embryos and of SCNT embryos, non-treated or treated with either psammaplin A (PsA) or vitamin C (VitC), was monitored, and the ESC derivation rates from the resulting blastocysts were determined. Blastocyst rates were similar among PsA-treated and VitC-treated SCNT embryos and ICSI embryos, but lower for non-treated SCNT embryos. ESC derivation rates were higher in treated SCNT embryos than in non-treated or ICSI embryos. Time-lapse microscopy analysis showed that non-treated SCNT embryos had a delayed development from the second division until compaction, lower number of blastomeres at compaction and longer compaction and cavitation durations compared with ICSI ones. Treatment of SCNT embryos with PsA further increased this delay whereas treatment with VitC slightly reduced it, suggesting that both treatments act through different mechanisms, not necessarily related to their epigenetic effects. Despite these differences, the time of completion of the third division, alone or combined with the duration of compaction and/or the presence of fragmentation, had a strong predictive value for blastocyst formation in all groups. In contrast, we failed to predict ESC derivation success from embryo morphokinetics. Time-lapse technology allows the selection of SCNT embryos with higher developmental potential and could help to increase cloning outcomes. Nonetheless, further studies are needed to find reliable markers for full-term development and ESC derivation success.

scholarly journals Exact Bayesian lineage tree-based inference identifies Nanog negative autoregulation in mouse embryonic stem cells

2016 ◽  
Author(s):  
Justin Feigelman ◽  
Stefan Ganscha ◽  
Simon Hastreiter ◽  
Michael Schwarzfischer ◽  
Adam Filipczyk ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Model Selection ◽  
Embryonic Stem ◽  
Time Lapse ◽  
Cell Populations ◽  
Time Lapse Microscopy ◽  
Stochastic Inference ◽  
Proliferating Cell ◽  
Selection Of

AbstractThe autoregulatory motif of Nanog, a heterogeneously expressed core pluripotency factor in mouse embryonic stem cells, remains debated. Although recent time-lapse microscopy data provide the unparalleled ability to monitor Nanog expression at the single-cell level, the extraction of mechanistic knowledge is precluded by the lack of inference techniques suitable for noisy, incomplete and heterogeneous data obtained from proliferating cell populations.This work identifies Nanog’s autoregulatory motif from quantified time-lapse fluorescence line-age trees with STILT (Stochastic Inference on Lineage Trees), a novel particle-filter based algorithm for exact Bayesian parameter inference and model selection of stochastic models. We first verify STILT’s ability to accurately infer parameters and select the correct autoregulatory motif from simulated data. We then apply STILT to time-lapse microscopy movies of a fluorescent Nanog fusion protein reporter and reject the possibility of positive autoregulation. Finally, we use STILT for experimental design, perform in silico overexpression simulations, and experimentally validate model predictions via exogenous Nanog overexpression. We finally conclude that the protein expression dynamics and overexpression experiments strongly suggest a weak negative feedback from the protein on the DNA activation rate.We find that a simple autoregulatory mechanism can explain the observed heterogeneous Nanog dynamics. This finding has implications on the understanding of the core pluripotency network, such as supporting the ability of mESC populations to diversify their proteomic profile to respond to a spectrum of differentiation cues. Beyond this application STILT constitutes a generally applicable fully Bayesian approach for model selection of gene regulatory models on the basis of time-lapse imaging data of proliferating cell populations. STILT is freely available at: http://www.imsb.ethz.ch/research/claassen/Software/stilt—stochastic-inference-on-lineage-trees.html

scholarly journals Nuclear transfer reprogramming does not improve the low developmental potency of embryonic stem cells induced by long-term culture

Reproduction ◽  
2006 ◽  
Vol 132 (2) ◽  
pp. 257-263 ◽  
Author(s):  
T Amano ◽  
M Gertsenstein ◽  
A Nagy ◽  
H Kurihara ◽  
H Suzuki
Keyword(s):  
Cell Lines ◽  
Nuclear Transfer ◽  
Embryoid Body ◽  
Es Cells ◽  
Embryonic Stem ◽  
Developmental Potential ◽  
Developmental Capacity ◽  
Developmental Rates

Epigenetic states of embryonic stem (ES) cells are easily altered by long-term cultivation and lose their developmental potential. To rescue this reduced developmental capacity, nuclear transfer (NT) of ES cells was carried out, and original ES and ES cells from cloned blastocysts (ntES) cells established after NT were compared with in vitro differentiation ability and developmental potential by embryoid body formation and tetraploid aggregation respectively. In the establishment of ntES cell lines, the oocytes fused with the ES cell were activated, and further cultured to cloned blastocysts. When in vitro differentiation ability was examined between original and ntES cell lines derived from ES cells with extensive passages (ES-ep), the day of appearance of simple embryoid body, cystic embryoid body, and spontaneous beating was almost similar. The developmental rates of ES-ep cells, that aggregated with tetraploid embryos to term, ranged from 3 to 6%. Moreover, the majority of live pups died soon after birth. In the ntES cell lines derived from ES-ep cells, developmental rates ranged from 0 to 5%. Those pups also died soon after birth, similar to the ES-ep-derived pups. These results suggest that profound epigenetic modifications of ES cells were retained in the re-established cell lines by NT.

scholarly journals Establishment of bovine expanded potential stem cells

2021 ◽  
Vol 118 (15) ◽  
pp. e2018505118
Author(s):  
Lixia Zhao ◽  
Xuefei Gao ◽  
Yuxuan Zheng ◽  
Zixin Wang ◽  
Gaoping Zhao ◽  
...  
Keyword(s):  
Stem Cells ◽  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Mammalian Species ◽  
Embryonic Stem ◽  
Preimplantation Embryos ◽  
Adult Individual ◽  
Developmental Potential ◽  
Cell Lineages

Embryonic stem cells (ESCs) and induced pluripotent stem cells have the potential to differentiate to all cell types of an adult individual and are useful for studying development and for translational research. However, extrapolation of mouse and human ESC knowledge to deriving stable ESC lines of domestic ungulates and large livestock species has been challenging. In contrast to ESCs that are usually established from the blastocyst, mouse expanded potential stem cells (EPSCs) are derived from four-cell and eight-cell embryos. We have recently used the EPSC approach and established stem cells from porcine and human preimplantation embryos. EPSCs are molecularly similar across species and have broader developmental potential to generate embryonic and extraembryonic cell lineages. We further explore the EPSC technology for mammalian species refractory to the standard ESC approaches and report here the successful establishment of bovine EPSCs (bEPSCs) from preimplantation embryos of both wild-type and somatic cell nuclear transfer. bEPSCs express high levels of pluripotency genes, propagate robustly in feeder-free culture, and are genetically stable in long-term culture. bEPSCs have enriched transcriptomic features of early preimplantation embryos and differentiate in vitro to cells of the three somatic germ layers and, in chimeras, contribute to both the embryonic (fetal) and extraembryonic cell lineages. Importantly, precise gene editing is efficiently achieved in bEPSCs, and genetically modified bEPSCs can be used as donors in somatic cell nuclear transfer. bEPSCs therefore hold the potential to substantially advance biotechnology and agriculture.

scholarly journals Large-scale time-lapse microscopy of Oct4 expression in human embryonic stem cell colonies

2016 ◽  
Vol 17 (1) ◽  
pp. 122-129 ◽  
Author(s):  
Kiran Bhadriraju ◽  
Michael Halter ◽  
Julien Amelot ◽  
Peter Bajcsy ◽  
Joe Chalfoun ◽  
...  
Keyword(s):  
Stem Cell ◽  
Embryonic Stem Cell ◽  
Human Embryonic Stem Cell ◽  
Large Scale ◽  
Embryonic Stem ◽  
Time Lapse ◽  
Oct4 Expression ◽  
Time Lapse Microscopy

Dynamic Persistence of Antibiotic-Stressed Mycobacteria

Science ◽  
2013 ◽  
Vol 339 (6115) ◽  
pp. 91-95 ◽  
Author(s):  
Yuichi Wakamoto ◽  
Neeraj Dhar ◽  
Remy Chait ◽  
Katrin Schneider ◽  
François Signorino-Gelo ◽  
...  
Keyword(s):  
Mycobacterium Smegmatis ◽  
Bacterial Population ◽  
Drug Exposure ◽  
Single Cells ◽  
Time Lapse ◽  
Dynamic State ◽  
Epigenetic Effects ◽  
Catalase Peroxidase ◽  
Apparent Stability ◽  
Selection Of

Exposure of an isogenic bacterial population to a cidal antibiotic typically fails to eliminate a small fraction of refractory cells. Historically, fractional killing has been attributed to infrequently dividing or nondividing “persisters.” Using microfluidic cultures and time-lapse microscopy, we found thatMycobacterium smegmatispersists by dividing in the presence of the drug isoniazid (INH). Although persistence in these studies was characterized by stable numbers of cells, this apparent stability was actually a dynamic state of balanced division and death. Single cells expressed catalase-peroxidase (KatG), which activates INH, in stochastic pulses that were negatively correlated with cell survival. These behaviors may reflect epigenetic effects, because KatG pulsing and death were correlated between sibling cells. Selection of lineages characterized by infrequent KatG pulsing could allow nonresponsive adaptation during prolonged drug exposure.

Experience of using time-lapse microscopy in IVF and ICSI programs

2020 ◽  
pp. 47-50
Author(s):  
N. V. Saraeva ◽  
N. V. Spiridonova ◽  
M. T. Tugushev ◽  
O. V. Shurygina ◽  
A. I. Sinitsyna
Keyword(s):  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Selection Procedure ◽  
Time Lapse ◽  
Single Embryo Transfer ◽  
Main Group ◽  
Clinical Pregnancy ◽  
Control Group ◽  
Time Lapse Microscopy

In order to increase the pregnancy rate in the assisted reproductive technology, the selection of one embryo with the highest implantation potential it is very important. Time-lapse microscopy (TLM) is a tool for selecting quality embryos for transfer. This study aimed to assess the benefits of single-embryo transfer of autologous oocytes performed on day 5 of embryo incubation in a TLM-equipped system in IVF and ICSI programs. Single-embryo transfer following incubation in a TLM-equipped incubator was performed in 282 patients, who formed the main group; the control group consisted of 461 patients undergoing single-embryo transfer following a traditional culture and embryo selection procedure. We assessed the quality of transferred embryos, the rates of clinical pregnancy and delivery. The groups did not differ in the ratio of IVF and ICSI cycles, average age, and infertility factor. The proportion of excellent quality embryos for transfer was 77.0% in the main group and 65.1% in the control group (p = 0.001). In the subgroup with receiving eight and less oocytes we noted the tendency of receiving more quality embryos in the main group (р = 0.052). In the subgroup of nine and more oocytes the quality of the transferred embryos did not differ between two groups. The clinical pregnancy rate was 60.2% in the main group and 52.9% in the control group (p = 0.057). The delivery rate was 45.0% in the main group and 39.9% in the control group (p > 0.050).

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