مجال
التميز
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تميز دراسي و بحثي
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البحوث المنشورة
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البحث (1):
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عنوان البحث:
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A
thyroid hormone receptor/KLF9 axis in human hepatocytes and pluripotent stem
cells
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رابط إلى البحث:
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تاريخ النشر:
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02/2015
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موجز عن البحث:
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Biological
processes require close cooperation of multiple transcription factors that
integrate different signals. Thyroid hormone receptors (TRs) induce
Krüppel-like factor 9 (KLF9) to regulate neurogenesis. Here, we show that
triiodothyronine (T3) also works through TR to induce KLF9 in HepG2 liver
cells, mouse liver, and mouse and human primary hepatocytes and sought to
understand TR/KLF9 network function in the hepatocyte lineage and stem cells.
Knockdown experiments reveal that KLF9 regulates hundreds of HepG2 target
genes and modulates T3 response. Together, T3 and KLF9 target genes influence
pathways implicated in stem cell self-renewal and differentiation, including
Notch signaling, and we verify that T3 and KLF9 cooperate to regulate key
Notch pathway genes and work independently to regulate others. T3 also
induces KLF9 in human embryonic stem cells (hESCs) and human induced
pluripotent stem cells (hiPSC) and this effect persists during
differentiation to definitive endoderm and hiPSC-derived hepatocytes.
Microarray analysis reveals that T3 regulates hundreds of hESC and hiPSC
target genes that cluster into many of the same pathways implicated in TR and
KLF9 regulation in HepG2 cells. KLF9 knockdown confirms that TR and KLF9
cooperate to regulate Notch pathway genes in hESC and hiPSC, albeit in a
partly cell-specific manner. Broader analysis of T3 responsive hESC/hiPSC
genes suggests that TRs regulate multiple early steps in ESC differentiation.
We propose that TRs cooperate with KLF9 to regulate hepatocyte proliferation
and differentiation and early stages of organogenesis and that TRs exert
widespread and important influences on ESC biology.
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البحث (2):
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عنوان البحث:
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Discordant
Growth of Monozygotic Twins Starts at the Blastocyst Stage: A Case Study
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تاريخ النشر:
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12/2015
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موجز عن البحث:
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Discordant
growth is a common complication of monochorionic/diamniotic pregnancies; in
approximately 50% of cases, the cause is unknown. The case presented here
suggests that discordant growth of monozygotic twins could start during
preimplantation development. Two inner cell masses (ICMs) within the same
blastocyst may originate in uneven splitting of a single “parental”
ICM, or the two ICMs may be formed independently de novo. We studied the
transcriptomes of two morphologically distinct ICMs within a single
blastocyst using high-resolution RNA sequencing. The data indicated that the
two ICM were at different stages of development; one was in the earliest
stages of lineage commitment, while the other had already differentiated into
epiblast and primitive endoderm. IGF1-mediated signaling is likely to play a
key role in ICM growth and to be the major driver behind these differences.
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البحث (3):
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عنوان البحث:
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Developmental
clock compromises human twin model created by embryo splitting
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تاريخ النشر:
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12/2015
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موجز عن البحث:
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STUDY
QUESTION:
Is the
quality of the human embryos generated by twinning in vitro comparable to the
quality of the embryos created by fertilization?
SUMMARY
ANSWER:
Our
data suggest that the human twin embryos created in vitro are unsuitable not
only for clinical use but also for research purposes.
WHAT IS
KNOWN ALREADY:
Pregnancy
from in vitro generated monozygotic twins by embryo splitting or twinning
leads to live birth of healthy animals. Similar strategies, however, have
been less successful in primates. Recent reports suggest that the splitting
of human embryos might result in viable, morphologically adequate
blastocysts, although the qualitative analyses of the embryos created in such
a way have been very limited.
STUDY
DESIGN, SIZE, DURATION:
This
study was a comparative analysis of embryos generated by twinning in vitro
and the embryos created by in vitro fertilization
PARTICIPANTS/MATERIALS,
SETTING, METHOD:
We
analysed morphokinetics and developmental competence of 176 twin embryos
created by splitting of 88 human embryos from either early (2-5 blastomeres,
n = 43) or late (6-10 blastomeres, n = 45) cleavage stages. We compared the
data with morphometrics of embryos created by in vitro fertilization and
resulting in pregnancy and live birth upon single blastocyst transfer (n =
42).
MAIN
RESULTS AND THE ROLE OF CHANCE:
The
morphokinetic data suggested that the human preimplantation development is
subjected to a strict temporal control. Due to a ‘developmental clock’, the
size of twin embryos was proportionate to the number of cells used for their
creation. Furthermore, the first fate decision was somewhat delayed; the
inner cell mass (ICM) became distinguishable later in the twin than in the normal
blastocysts obtained through fertilization. If an ICM was present at all, it
was small and of poor quality. The majority of the cells in the twin embryos
expressed ICM and trophectoderm markers simultaneously.
LIMITATIONS,
REASONS FOR CAUTION:
We created
monozygotic twins by blastomere separation from cleavage stage embryos.
Embryo twinning by blastocyst bisection may circumvent limitations set by the
developmental clock.
WIDER
IMPLICATIONS OF THE FINDINGS:
Taken
together, our data suggest that the human twin embryos created in vitro are
unsuitable not only for clinical use but also for research purposes.
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البحث (4):
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عنوان البحث:
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MicroRNAs in spent blastocyst culture
medium are derived from trophectoderm cells and can be explored for human embryo
reproductive competence assessment
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تاريخ النشر:
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01/2016
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موجز عن البحث:
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OBJECTIVE:
To assess whether extracellular microRNAs
(miRNAs) can be accurately profiled from spent blastocyst culture media (SBM)
and used as embryonic biomarkers.
DESIGN:
Prospective cohort study.
SETTING:
Private and academic in vitro fertilization
centers.
PATIENT(S):
Inner cell mass-free trophectoderm (TE)
samples and their relative SBM from five good-quality human blastocysts.
INTERVENTION(S):
Protocol for miRNA purification and
analysis based on quantitative polymerase chain reaction set and validated on
human embryonic stem cells (hESCs) and on SBM with and without biological
variability.
MAIN OUTCOMES MEASURE(S):
Analysis of miRNAs in culture media in
relation with TE cells and comparison of miRNA profiles between implanted and
unimplanted euploid blastocysts.
RESULT(S):
Culture media from embryos in the cleavage,
morula, and blastocyst stages were collected to investigate the presence of
miRNAs. The SBM were prospectively collected from euploid implanted (n = 25)
and unimplanted blastocysts (n = 28) for comparison. We hypothesized that
human embryos secrete miRNAs in culture media that can be used as biomarkers.
The comparative analysis of TE and SBM samples revealed that 96.6% (57 of 59;
95 CI, 88.3-99.6) of the miRNAs detected in the SBM were expressed from TE
cells, suggesting a TE origin. The culture media collected from cleavage and
morula stage embryos showed a pattern similar to blanks, suggesting that
miRNAs profiling from spent culture media applies only for blastocysts.
MicroRNAs analysis of SBM from euploid implanted and unimplanted blastocysts
highlighted two miRNAs (miR-20a, miR-30c) that showed increased
concentrations in the former and were predicted in silico to be involved in
23 implantation-related pathways.
CONCLUSION(S):
MicroRNAs secreted from human blastocysts
in culture media can be profiled with high reproducibility, and this approach
can be further explored for non-invasive embryo selection.
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البحث (5):
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عنوان البحث:
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Potential of human twin embryos generated by
embryo splitting in assisted reproduction and research
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تاريخ النشر:
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10/2016
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موجز عن البحث:
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BACKGROUND:
Embryo splitting or twinning has been
widely used in veterinary medicine over 20 years to generate monozygotic
twins with desirable genetic characteristics. The first human embryo
splitting, reported in 1993, triggered fierce ethical debate on human embryo
cloning. Since Dolly the sheep was born in 1997, the international community
has acknowledged the complexity of the moral arguments related to this
research and has expressed concerns about the potential for reproductive
cloning in humans. A number of countries have formulated bans either through
laws, decrees or official statements. However, in general, these laws
specifically define cloning as an embryo that is generated via nuclear
transfer (NT) and do not mention embryo splitting. Only the UK includes under
cloning both embryo splitting and NT in the same legislation. On the
contrary, the Ethics Committee of the American Society for Reproductive
Medicine does not have a major ethical objection to transferring two or more
artificially created embryos with the same genome with the aim of producing a
single pregnancy, stating that ‘since embryo splitting has the potential to
improve the efficacy of IVF treatments for infertility, research to
investigate the technique is ethically acceptable’.
OBJECTIVE AND RATIONALE:
Embryo splitting has been introduced
successfully to the veterinary medicine several decades ago and today is a
part of standard practice. We present here an overview of embryo splitting
experiments in humans and non-human primates and discuss the potential of
this technology in assisted reproduction and research.
SEARCH METHODS:
A comprehensive literature search was
carried out using PUBMED and Google Scholar databases to identify studies on
embryo splitting in humans and non-human primates. ‘Embryo splitting’ and
’embryo twinning’ were used as the keywords, alone or in combination with
other search phrases relevant to the topics of biology of preimplantation
embryos.
OUTCOMES:
A very limited number of studies have been
conducted in humans and non-human primates. The published material,
especially the studies with human embryos, is controversial. Some reports
suggest that twinning technology will find clinical use in reproductive
medicine in the future, whereas others conclude the opposite that human twin
embryos created in vitro are unsuitable not only for clinical, but also for
research, purposes.
WIDER IMPLICATIONS:
The blastomere biopsy technique of embryo
splitting seems to be unsuitable for either clinical or research purposes;
however, embryo bisection, a preferable method of cloning in veterinary
medicine, has not yet been tested on human embryos.
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البحث (6):
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عنوان البحث:
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Human Embryos Created by Embryo Splitting
Secrete Significantly Lower Levels of miRNA-30c
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رابط إلى البحث:
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تاريخ النشر:
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12/2016
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موجز عن البحث:
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Studies reporting term pregnancy and the
production of genetically identical offspring from isolated blastomeres of
early stage embryos have been carried out in small and large animals.
However, very little is known about the effects of embryo splitting on the
development and reproductive competency of human embryos. In this study, we
investigated the effects of embryo splitting on profile of microRNAs (miRNAs)
detected in their spent blastocyst medium (SBM) by comparative analysis of
miRNA profiles in SBM of human twin embryos created by blastomere biopsy and
SBM of blastocysts that resulted in a healthy pregnancy and live birth
following embryo transfer. The profile of miRNA secretion in in vitro culture
media consistently distinguishes twin from control embryos. We found that six
miRNAs are significantly more abundant in SBM from twin embryos, while nine
are significantly more abundant in SBM from euploid implanted blastocysts.
These nine include miRNA-30c, a previously reported marker of blastocyst
implantation potential. Furthermore, 22.9% of miRNAs secreted by twin embryos
were never detected in SBM from normal reproductively competent blastocysts,
or from trophectoderm (TE) samples from normal blastocysts donated for the
research. The miRNA profile, unique to twin blastocysts, might be a result of
differential lineage commitment in these embryos.
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المؤتمرات العلمية:
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المؤتمر (1):
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عنوان المؤتمر:
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31st Annual
Meeting of the European Society of Human Reproduction and Embryology
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تاريخ الإنعقاد:
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17/06/2015
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مكان
الإنعقاد:
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Lisbon, Portugal
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طبيعة المشاركة:
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Poster
presentation
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عنوان المشاركة:
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Comparative
analysis of twin blastocysts derived from human embryo splitting at cleavage
stage
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ملخص المشاركة:
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STUDY QUESTION: Embryo splitting has a potential to reduce
number of the embryos used for research. Our aim was to investigate safety of
embryo splitting technique by comparing morphokinetics and linage commitment
of monozygotic twin blastocysts (derived from split embryos) with normal
embryos that with laser ablation of zona pellucida on Day 3 and cultured to
D5/D6.
METHODS: Cleavage stage embryos (n=88), donated for
research and underwent splitting, were grouped into two groups based on the
number of blastomeres that survived post thawing process. Group 1 consisted
embryos with 2-5 blastomeres (n=43) and Group 2 consisted embryos with 6-10
blastomeres (n=45). Matching cleavage stage embryos (n=42) that resulted in
positive clinical pregnancy following single embryo transfer were used as a
control. Half numbers of the blastomeres from the donor embryos were biopsied
and transferred into the empty recipient zona pellucida (Fig. 1). Resulting
twin embryos were cultured in the EmbryoScope® for morphokinetic studies.
Further, we investigated the expression of early lineage-specific gene
markers (NANOG,CDX2 and GATA2) in the blastocysts derived from split embryos
using immunostainining.
RESULTS: Splitting human pre-implantation embryos
from the Group 2 resulted, in general, in more viable and better quality
blastocysts compared to the embryos from the Group 1 (Fig. 2). Blastocysts
developed from split embryos were smaller than the control blastocysts (Fig.
3). In majority of split embryos, ICM was poorly developed or
indistinguishable, containing cells with a dual expression of TE and ICM
markers (Fig. 4).
CONCLUSION: The data suggest that the blastocysts
derived from split embryos are inferior in comparison with normal matching
blastocysts and as such might not be suitable as a research tool.
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