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  • Title/Summary/Keyword: Embryonic developmental

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Morphological Characteristics of Pig Blastocysts Produced by Somatic Cell Nuclear Transfer

  • Y.M. Han;D.B. Koo;Park, Y.H.;Park, J.S.;Kim, H.N.;Y.K. Kang;W.K. Chang;Lee, K.K.
    • Proceedings of the KSAR Conference
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    • 2001.03a
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    • pp.68-68
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    • 2001
  • Blastocyst formation, consisting of the inner cell mass (ICM) and trophectoderm (TE), is the first differentiation process during embryonic development in mammals. It has been hypothesized that the proportion of ICM to TE in the blastocyst may be crucial for subsequent developmental competence of early embryos, which it may be expressed as a sensitive indicator for evaluating in vitro systems. In this study ICM/total cell ratio of nuclear transfer (NT) embryos was compared with IVF-derived and in vivo embryos. Somatic cell nuclei obtained from a fetus at Day 40 of gestation were transferred into the enucleated oocyte and then cultured in NCSU 23 medium for 6 days as previously described (Koo et al., Biol. Reprod. 2000; 63:986-992). ICM and TE cells of blastocysts were determined by using a differential staining method (Han et al., Biol. Reprod. 1999; 60:1110-1113). Development rate (9.8±2.5%, 23/225) to the blastocyst stage of NT embryos was lower than IVF embryos (23.8±2.7%, 53/223). Thus, a difference was detected in the in vitro developmental rate to blastocyst stage between NT and IVF-derived embryos (P<0.05). In the next experiment, we investigated ICM and TE nuclei to assess the quality of blastocysts that produced by NT, IVF and in vivo, respectively. NT blastocysts (27.6±8.3) showed a smaller total cell number than IVF-derived (42.6±17.4) and in vivo embryos (283.9±103.5) (P<0.05). Ratios of ICM/total cells in NT, IVF and in vivo blastocysts were 15.1± 18.6% (n=56), 12.3±9.2% (n=57) and 30.4±6.8% (n=40), respectively. Individual blastocysts for the ratio of ICM/total cells were assigned to 3 groups (I; <20%, II; 20 to 40% and III;>40%). As the results, most in vivo blastocysts (97.5%, 39/40) were distributed into group II while most NT (78.6%, 44/56) and IVF-derived blastocysts (82.5%, 47/57) were allocated to group I. Thus, our data show that NT or IVF-derived embryos have aberrant morphology during early development in vitro systems, suggesting that these anomalies may result in developmental failures of the NT embryos to term.

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Inhibition of Developmental Processes by Flavone in Caenorhabditis elegans and Its Application to the Pinewood Nematode, Bursaphelenchus xylophilus

  • Lee, Yong-Uk;Kawasaki, Ichiro;Lim, Yoongho;Oh, Wan-Suk;Paik, Young-Ki;Shim, Yhong-Hee
    • Molecules and Cells
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    • v.26 no.2
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    • pp.171-174
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    • 2008
  • Flavone (2-phenyl chromone) is a well-known plant flavonoid, but its bioactivity has been little explored. Treatment of Caenorhabditis elegans or C. brissage with flavones induced embryonic and larval lethality that was pronounced in early larval stages. This anti-nematodal effect was also observed in the pinewood nematode, B. xylophilus. LD50 values were approximately 100μM for both B. xylophilus and C. elegans. Our results indicate that flavone is an active nematicidal compound that should be further investigated with the aim of developing a potent drug against B. xylophilus.

Examination of Improved Tetracycline Inducible Gene Expression System In Vitro (새로운 Tetracycline 유도적 유전자 발현 System의 In Vitro 검정)

  • Kwon, Mo Sun;Kim, Teoan;Koo, Bon Chul
    • Reproductive and Developmental Biology
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    • v.37 no.3
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    • pp.109-115
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    • 2013
  • Until recently the most popular tetracycline-inducible gene expression system has been the one developed by Gossen and Bujard. In this study, we tested the latest version of same system and the results are summarized as follows: Compared with previous one, the difference of new system are minor changes of nucleotide sequences in transactivator and tetracycline response element (TRE) regions. Sensitivity to the doxycycline (a tetracycline derivative) was improved. Leakiness of GFP marker gene expression in non-inducible condition was significantly decreased. Higher expression of the marker gene was observed when the cells were fed with doxycycline-containing medium. Optimal insertion site of woodchuck posttranscriptional regulatory element (WPRE) sequence which was known to increase gene expression was different depending on the origin of cells. In chicken embryonic fibroblast, location of WPRE sequence at 3' end of TRE resulted in the highest GFP expression. In bovine embryonic fibroblasts, 3' end of transactivator was the best site for the GFP expression.

The Effect of P almulgunjatang gamibang on the Ovarian Functions in Female Mice (팔물군자탕(湯八物君子) 가미방(加味方)이 자성(雌性)생쥐의 생식능력에 미치는 영향)

  • Lee, Su-Keun;Kim, Dong-Chul
    • The Journal of Korean Obstetrics and Gynecology
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    • v.23 no.4
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    • pp.20-34
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    • 2010
  • Purpose: These experiments were undertaken to evaluate the effect of administration of Palmulgunjatang gamibang on ovarian functions in female mice. Methods: We administered the Palmulgunjatang gamibang to 6-week-old female CF-1 mice for 4, 8, or 12 days. After administration of Palmulgunjatang gamibang with different concentration, the female mice were injected PMSG and hCG for ovarian hyperstimulation. The mice were divided into 3 different groups for each experiment. To compare the differences, we set a control group treated with plain water at the same volume by the same way. Results: In case of 4-day, 8-day, 12-day administration of Palmulgunjatang gamibang, the mean number of total ovulated oocytes and the number of morphologically normal oocytes were increased compared with control group. We were also examined the embryonic developmental competence in vitro. In case of 4-day administration of Palmulgunjatang gamibang, the rates of blastocyst formation from 2-cell stages were higher than control group. Conclusion: From our results suggested that the medication of Palmulgunjatang gamibang has beneficial effect on reproductive functions of female mice via promotion of cell proliferation.

Differential Expression of TPX2 upon Differentiation of Human Embryonic Stem Cells

  • Noh, Hye-Min;Choi, Seong-Jun;Kim, Se-Hee;Kim, Kye-Seong;Kim, Jin-Kyeoung
    • Reproductive and Developmental Biology
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    • v.31 no.4
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    • pp.221-226
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    • 2007
  • Embryonic stem (ES) cells are known to have an infinite proliferation and pluripotency that are associated with complex processes. The objective of this study was to examine expression of genes differentially regulated during differentiation of human ES cells by suppression subtractive hybridization (SSH). Human ES cells were induced to differentiate into neural precursor cells via embryoid body. Neural precursor cells were isolated physically based on morphological criteria. Immunocytochemical analysis showed expression of pax6 in neural precursor cells, confirming that the isolated cells were neural precursor cells. Undifferentiated human ES cells and neural precursor cells were subject to the SSH. TPX2 (Targeting Protein for Xklp2 (Xenopus centrosomal kinesin-like protein 2)) was identified, cloned and analyzed during differentiation of human ES cells into neural lineages. Expression of TPX2 was gradually down-regulated in embryoid bodies and neural precursor cells relative to undifferentiated ES cells. Targeting Protein for Xklp2 has been shown to be involved in cell division by interaction with microtubule development in cancer cells. Taken together, result of this study suggests that TPX2 may be involved in proliferation and differentiation of human ES cells.

Effects of Cadmium, Copper, Chromium, Nickel, Silver, and Zinc on the Embryonic Development of the Sea Urchin, Strongylocentrotus intermedius (북쪽말똥성게 (Strongylocentrotus intermedius) 배아 (embryo)를 이용한 중금속에 대한 민감도 비교)

  • Ryu, Tae-Kwon;Hwang, In-Young;Lee, Taek-Kyun;Yoon, Jun-Heon;Lee, Chang-Hoon
    • Environmental Analysis Health and Toxicology
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    • v.25 no.1
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    • pp.19-26
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    • 2010
  • Discharged materials from the point or non-point source are released into the sea, and as the results, marine environment is directly affected. We must estimate the impacts of contaminants to marine pollution rapidly and accurately. Therefore, it is needed on early warning system for appreciating marine environmental impacts, and required a bioassay to evaluate abnormal changes. A bioassay test was developed to examine the effects of heavy metal contaminants on the early life stages of the marine annimals. We have studied the effects of metals on early development of a sea urchin species, Strongylocentrotus intermedius. S. intermedius embryos were tested with six metals (Cu, Ag, Zn, Cd, Cr, Ni) and showed the highest sensitivity to Cu as well as the lowest sensitivity to Cd. The order of biological impact for metals was Cu>Ag>Ni>Zn>Cr>Cd. In accordance with the results, sea urchins embryos can provide biological criteria for seawater quality assessment. The sensitivity of developmental bioassay whith S. intermedius is at intermediate level among marine organisms commonly used in aquatic bioassays. And this sea urchin can be routinely employed as a test organism for ecotoxicity assays.

Developmental Rate of Rabbit Parthenogenetic Embryos Derived Using Different Activating Protocols

  • Chrenek, P.;Makarevich, A.
    • Asian-Australasian Journal of Animal Sciences
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    • v.17 no.5
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    • pp.617-620
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    • 2004
  • The present study compares development of rabbit embryos generated using different oocyte activation protocols and reconstructed with embryonic or cumulus cells as nuclear donor. In vivo matured oocytes were collected from New Zealand White rabbits at 16 h after ovulation treatment and were activated at18 h of post-ovulation treatment. The following schemes of oocytes activation were tested: 1) single electric pulse (EP, 3.2 kV/cm, 3×20 μs, 0.3 M mannitol)+5 min culture in the presence of 5 mM Ionomycin, 2) single electric pulse (EP, 3.2 kV/cm, (×20 μs, 0.3 M mannitol)+1 h culture in the presence of 2 mM 6-DMAP, and 3) three electric pulses 30 min apart. Cleavage rate, percentage of expanded and hatched blastocysts as well as total cell number of blastomeres of parthenogenetic embryos were significantly higher using either EP+6-DMAP or 3×EP schemes, comparing with EP+Ionomycin. Development rate up to hatched blastocyst stage of cloned rabbit embryos using the EP+6-DMAP for activation of nuclei were 19% for embryonic cell nuclei and 36% for cumulus cell nuclei. The best activation protocol optimalized in this study was the combined treatment "P+6-DMAP" which may be potentially used for nuclear transfer protocol.

Molecular Characterization of Porcine DNA Methyltransferase I

  • Lee, Yu-Youn;Kang, Hye-Young;Min, Kwan-Sik
    • Reproductive and Developmental Biology
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    • v.34 no.4
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    • pp.283-288
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    • 2010
  • During normal early embryonic development in mammals, the global pattern of genomic DNA methylation undergoes marked. changes. The level of methylation is high in male and female gametes. Thus, we cloned the cDNA of the porcine DNA methyltransferase 1 (Dnmt1) gene to promote the efficiency of the generation of porcine clones. In this study, porcine Dnmt1 cDNA was sequenced, and Dnmt1 mRNA expression was detected by reverse transcription-polymerase reaction (RT-PCR) in porcine tissues during embryonic development. The porcine Dnmt1 cDNA sequence showed more homology with that of bovine than human, mouse, and rat. The complete sequence of porcine Dnmt1 cDNA was 4,774-bp long and consisted of an open reading frame encoding a protein of 1611 amino acids. The amino acid sequence of porcine DNMT1 showed significant homology with those of bovine (91%), human (88%), rat (76%), and mouse (75%) Dnmt1. The expression of porcine Dnmt1 mRNA was detected during porcine embryogenesis. The mRNA was detected at stages of porcine preimplantation development (1-cell, 2-cell, 4-cell, 8-cell, morula, and blastocyst stages). It was also abundantly expressed in tissues (lung, ovary, kidney and somatic cells). Further investigations are necessary to understand the complex links between methyltransferase 1 and the transcriptional activity in cloned porcine tissues.

Gene Expression Analysis of Gα13-/- Knockout Mouse Embryos Reveals Perturbations in Gα13 Signaling Related to Angiogenesis and Hypoxia

  • Park, Ji-Hwan;Choi, Sang-Dun
    • Genomics & Informatics
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    • v.9 no.4
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    • pp.161-172
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    • 2011
  • Angiogenesis is regulated by a large number of molecules and complex signaling mechanisms. The G protein Gα13 is a part of this signaling mechanism as an endothelial cell movement regulator. Gene expression analysis of Gα13 knockout mouse embryos was carried out to identify the role of Gα13 in angiogenesis signaling during embryonic development. Hypoxia-inducible response factors including those acting as regulators of angiogenesis were over expressed, while genes related to the cell cycle, DNA replication, protein modification and cell-cell dissociation were under expressed. Functional annotation and network analysis indicate that Gα13/ embryonic mice were exposed to hypoxic conditions. The present analysis of the time course highlighted the significantly high levels of disorder in the development of the cardiovascular system. The data suggested that hypoxia-inducible factors including those associated with angiogenesis and abnormalities related to endothelial cell division contributed to the developmental failure of Gα13 knockout mouse embryos.

A Maternal Transcription Factor, Junction Mediating and Regulatory Protein is Required for Preimplantation Development in the Mouse

  • Lin, Zi-Li;Li, Ying-Hua;Jin, Yong- Xun;Kim, Nam-Hyung
    • Development and Reproduction
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    • v.23 no.3
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    • pp.285-295
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    • 2019
  • Junction-mediating and regulatory protein (JMY) is a regulator of both transcription and actin filament assembly. The actin-regulatory activity of JMY is based on a cluster of three actin-binding Wiskott-Aldrich syndrome protein homology 2 (WH2) domains that nucleate actin filaments directly and promote nucleation of the Arp2/3 complex. In addition to these activities, we examined the activity of JMY generation in early embryo of mice carrying mutations in the JMY gene by CRISPR/Cas9 mediated genome engineering. We demonstrated that JMY protein shuttled expression between the cytoplasm and the nucleus. Knockout of exon 2, CA (central domain and Arp2/3-binding acidic domain) and NLS-2 (nuclear localization signal domain) on the JMY gene by CRISPR/Cas9 system was effective and markedly impeded embryonic development. Additionally, it impaired transcription and zygotic genome activation (ZGA)-related genes. These results suggest that JMY acts as a transcription factor, which is essential for the early embryonic development in mice.