Gangliosides are glycolipids in which oligosaccharide is combined with sialic acids. Our previous studies have suggested an interplay between ganglioside GD1a/GT1b and meiotic maturation capacity in porcine oocyte maturation. Furthermore, ganglioside GD1a and GT1b are known for its antioxidant activity, but it is still unclear whether possible antioxidant role of GD1a and GT1b is involved in porcine embryos development competence during in vitro culture (IVC). Here, the effects of ganglioside GD1a and GT1b on the embryonic developmental competence during in vitro culture of porcine were investigated. The effects of ganglioside GD1a and GT1b on the expression of ST3GAL2 were confirmed during embryos development (2-cell, 4-cell, 8-cell and blastocyst) using immunofluorescent staining (IF). As a result, the fluorescent expression of ST3GAl2 was higher in embryos at 4-8 cells stage than blastocysts. Blastocyst development rate significantly increased in only 0.1 μM GD1a and GT1b treated groups compared with control group. To investigate the cellular apoptosis, we analyzed TUNEL assay. In case of only 0.1 μM GD1a and GT1b treated groups, the total number of cells in blastocyst compared with control group, but there was no significant difference in the rate of apoptotic cells. We identified the intracellular ROS levels using DCF-DA staining. According to the result, ROS production significantly decreased in blastocysts derived from the 0.1 μM GD1a and GT1b treated groups. These results suggest that ganglioside GD1a and GT1b improve the developmental competence of porcine embryos via reduction of intracellular ROS during preimplantation stage.

Among fatty acid families, the polyunsaturated fatty acids were demonstrated to be mediators in various reproductive processes as precursor of steroid hormone (via cholesterol) and prostaglandins (via arachidonic acid), and in the last decade, major research was focused on the effects of omega-6 and especially omega-3 fatty acid. Eicosapentaenoic acid, the longest members of omega-3 fatty acid family, can be produced by a series of desaturation and elongation reactions from shorter member such as α-Linolenic acid. However, very few studies have provided detailed descriptions of Eicosapentaenoic acid effects and mechanisms of action in mammalian oocytes. The purpose of this study was to evaluate the effect of Eicosapentaenoic acid supplementation on in vitro maturation and developmental potential of porcine oocytes. Various concentrations of Eicosapentaenoic acid was added into in vitro maturation medium, and we evaluated the degree of cumulus expansion, nuclear maturation rate, blastocysts quality, and levels of prostaglandin E2, 17β-estradiol, progesterone in the spent medium. High doses (100 mM) of Eicosapentaenoic acid supplementation significantly inhibited cumulus expansion and oocyte nuclear maturation, and prostaglandin E2 synthesis also significantly decreased compared with other groups (p < 0.05). Supplementation of 50 mM Eicosapentaenoic acid showed higher quality blastocysts in terms of high cell numbers and low apoptosis when compared with other groups (p < 0.05), and synthesis ratio of E2/P4 also significantly increased compared with control group (p < 0.05). However, Supplementation of 100 mM Eicosapentaenoic acid showed high apoptosis when compared with other groups (p < 0.05), and synthesis ratio of 17b-estradiol/progesterone also significantly decreased compared with control group (p < 0.05). Our results indicated that supplementation with appropriate levels of Eicosapentaenoic acid beneficially affects the change of hormone synthesis for controlling oocyte maturation, leading to improved embryo quality. However, high doses of Eicosapentaenoic acid treatment results in detrimental effects.

Omega-3 α-linolenic acid and omega-6 linoleic acid are essential fatty acids for health maintenance of human and animals because they are not synthesized in vivo. The purpose of this study was to evaluate the effect of α-linolenic acid and linoleic acid supplementation on in vitro maturation and developmental potential of porcine oocytes. Various concentrations of α-linolenic acid and linoleic acid were added into in vitro maturation medium, and we evaluated the degree of cumulus expansion, oocyte nuclear-maturation rate, blastocyst rate, blastocyst quality, and levels of prostaglandin E2, 17b-estradiol, and progesterone in the spent medium. High doses (100 μM) of α-linolenic acid and linoleic acid supplementation significantly inhibited cumulus expansion and oocyte nuclear maturation, and prostaglandin E2 synthesis also significantly decreased compared with other groups (p < 0.05). Supplementation of 50 μM α-linolenic acid and 10 μM linoleic acid showed higher quality blastocysts in terms of high cell numbers and low apoptosis when compared with other groups (p < 0.05), and synthesis ratio of 17b-estradiol / progesterone also significantly increased compared with control group (3.59 ± 0.22 vs. 2.97 ± 0.22, 3.4 ± 0.28 vs. 2.81 ± 0.19, respectively; p < 0.05). Our results indicated that supplementation with appropriate levels of α-linolenic acid and linoleic acid beneficially affects the change of hormone synthesis (in particular, an appropriate increase in the 17b-estradiol / progesterone synthesis ratio) for controlling oocyte maturation, leading to improved embryo quality. However, high doses of α-linolenic acid and linoleic acid treatment results in detrimental effects.

Growth differentiation factor8 (GDF8) is a member of the transforming growth factor-β that has been identified as a strong physiological regulator. Overall of the current studies, the GDF8 is detected in oviduct fluid and uterus which led us to suggest that the GDF8 may effect on preimplantation embryonic development and act paracrine role to correlate with successful late-blastocyst implantation in in vivo. The purpose of this study is the effects of GDF8 on porcine parthenogenesis (PA) embryo development during in vitro culture (IVC). We were investigated the effect of GDF8 supplement during PA embryo IVC by cleavage and blastocyst formation rate and patterning analysis. Data were analyzed by on way ANOVA, followed by Tukey’s range test. Respectively 0.2, 2 and 20 ng/mL of GDF8 were added during IVC followed experiment design as control, 0.2, 2, and 20 GDF8 supplement groups. After 48h of embryo culture time, no significant difference was observed on cleavage rate from the different concentration (0, 0.2, 2, and 20 ng/ml) of GDF8 supplement groups (65.7%, 66.0%, 66.3%, and 65.8%, respectively). After 120h of embryo culture time, the 0.2 and 2 group showed significantly (p<0.05) higher blastocyst formation rate than control (40.4% and 36.4% VS 40.4%, respectively). In embryo developmental pattern analysis, the 0.2 ng/ml GDF8 supplement groups showed significantly higher (p<0.05) 2-3 cell cleavage- and early blastocyst pattern compared with control (12.0% and 10.4% VS 6.6% and 6.2%, respectively). However there are no significantly different pattern was observed in other groups. In conclusion, the 0.2 ng/ml of GDF8 supplementation during porcine PA embryo IVC significantly changed embryonic developmental patterns. However there are further studies are required such as analysis of blastocyst total number, specific gene transcription pattern, and ICM/TE rate to make clarify and support the conclusion.

Embryonic compaction is essential for normal preimplantation development in mammals. The present study was to investigate the effects of compaction patterns on developmental competence of pig embryos. The proportion of blastocyst formation derived from compacted morula was higher than those of compacting and pre-compacting morula (P<0.01). Nuclei numbers of inner cell mass (ICM), trophectoderm (TE), and total of blastocysts derived from compacted group were also superior to those of compacting and pre-compacting groups (P<0.05). Then, compaction patterns, developmental ability and structural integrity were compared between mono- and poly-spermic embryos. The rate of compacted morula in mono-spermic embryos was higher than that of poly-spermic embryos (P<0.05). Especially, the rate of blastocyst formation derived from compacted embryos in mono-spermic embryo group was higher than that of poly-spermic embryo group (P<0.05), although no difference was detected between the two groups in the structural integrity. Finally, we confirmed that beta-catenin was differentially expressed according to compaction patterns in morula and blastocyst stage embryos. In conclusion, our results suggest that the compaction patterns during preimplantation development play a direct role in developmetal competence and quality of pig embryos.

Meckel’s cartilage is one of the earliest structure to appear in a mandible derived from the first branchial arch and serves as the primordium for the formation of mandible, malleus, incus, and sphenomandibular ligament. However, its direct role and the mechanism in mandibular development are not well elucidated. To address this issue, we observed morphological and histological changes and gene expression patterns in the Meckel's cartilage of a developng mouse, from E13.5 to E18.5 embryos, using skeletal preparation samples and routinely prepared slide sections for light rnicro scopic observation in various sectional planes. The following methods were performed: H&E staining for general histo logical observation, Von Kossa staining for detection of rnineralization, TRAP actiγity staining for localization of osteoclastic cells, immunohistochernistry for MMP- 1 and - 9 for evaluation of enzymatic activity of osteoclastic cells, and in situ hybridization for detection of collagen type 1. II, and X mRNA expression, respectively. At E13.5 Meckel's cartilage appeared as a V- shaped rod fused at the rnidline and thin rnineralized mandibular buccal plate was formed lateral to, and at some distance from, Meckel's cartilage in an intramembranous ossification mode With the progression of tooth development, the Meckel’s in cartilage adjacent incisors revealed hypertrophic chondrocyte differentiation with rnineralization of the chondroid matrix The Meckel’s cartilage was replaced with bone by - osteoclasts, showing strong immunoreactivity for MMP-1 and - 9 from E16.5. With time, this bony replacement of Meckel’s cartilage in an endochondral ossification mode was extended up to the rnid- portion of the molar sockets till E18 5 The bony replacement of rnineralized hypertrophic chondrocyte zone expressing X collagen mRNA contributed to the formation of thick mandibular lingual plate. These findings suggest that mandibular formation and development is closely related with not only Meckel's cartilage, but also with the developing tooth, and that certain influence from the developing tooth may play a role in deterrnining the fate of Meckel’s cartilage during mandibular development.

키틴생합성저해제인 diflubenzuron을 톱다리개미허리노린재(Riptortus clavatus Thunberg)에 미량국소처리하였을 때 충태 및 영기에 따른 약제의 감수성차이와, 종령약충 처리후 우화율 및 우화성충에 미치는 영향을 조사하였다. Diflubenzuron은 살람효과를 보였으며 산란후 12시간내의 알은 산란후 48~60시간의 알보다 감수성이 높았으나, 알의 나이에 관계없이 처리된 알의 배는 정상적으로 발육하였다. 영기별 감수성은 영기가 진행될수록 낮아져 1령약충이 2령에 비하여 1.5배, 3령약충에 비하여 18.2배, 4령약충에 비하여 39.4배, 5령약충에 비하여 42.4배 높은 감수성을 나타내었다. 종령약충에 처리하였을 때 우화율과 우화성충의 체중, 수명 및 산란율은 현저히 감소하였다.

Telomeres are the end of chromosomes and consist of a tandem repeat sequence of (TTAGGG)n and associated proteins. Telomerase is a ribonucleoprotein which act as a template for the synthesis of telomeric DNA. Telomeres are essential for chromosome stability and are related with cell senescence, apoptosis and cancer. Even though telomeres and telomerase have been studied extensively, very little is known about telomere dynamics in embryonic cells. This study was carried out to analyze the telomeres distribution and telomerase activity of chicken cells during embryonic and developmental stages. The target cells for analysing were sperms, ovulated ova, early embryonic cells and the cells from brain, heart, liver, kidney and germinal tissue in fetus. Telomeres distribution on target cells was analyzed by Q-FISH (Quantitation-Fluorescence in situ Hybridization) techniques using a chicken telomere repeat probe. Telomerase activity was performed by TRAP assay (Telomeric repeat Amplification Protocol) with target DNA. In results, the telomeres of chicken were found at the ends of all chromosomes. In addition, chicken had interstitial telomeres on chromosomes 1, 2 and 3. Telomerase activity was highly detectable in early embryonic cells, germinal tissues and kidney cells. Whereas telomerase activity was gradually down-regulated when the organs, including brain, heart, and liver, were developed from embryos. In the distribution of telomeric DNA on the embryonic and developmental stages, most of the cells was gradually decreased in telomere quantity during ontogenesis.

Even though success in birth of live offspring from nuclear transfer(NT) using somatic cells in many species, detailed information on processes or mechanisms of development are not well known. Cytoplasm of bovine oocyte has been known to support the development of nuclear transferred embryos using nuclear donor cells from different species. Therefore, interspecies NT might be used to find answers of some questions in basic aspect of nuclear transfer In this study, we examined the developmental potential of reconstructed embryos when bovine oocyte as a cytoplasm recipient and mouse embryonic fibroblast as a nuclear donor were used. The nuclear transfer units were aliocated in Group 1 (murine block media and normal media) and Group 2. (bovine block media and normal media). NT units were not blocked at 2-cell stage regardless of types of medium. On mouse media, poor development of interspecies NT units was observed compared to bovine media. However, as NT units cultured in bovine normal medium, embryos developed over 8-cell stage. Further studies performed to increase the developmental rate in condition of antioxidant treatment. Despite low development, bovine-murine interspecies nuclear transferred embryos could develop to blastocysts and they showed that blastocyts rate of antioxidant group was superior to those of non-antioxidant group. Next, we investigated gene expression pattern which is carried out for zygotic activation. The Xist gene is expressed in female mouse embryo after zygotic activation of 4-cell stage. But interspecies nuclear transferred embryos do not express Xist gene at 4-cell stage. As a result, it is suggested that the bovine cytoplasm controls the early preimplantation development in interspecies NT However, the development of later stages might require genomic control from transferred donor nucleus. Therefore, even though the involvement of several other factors such as mitochondrial incompatibility, effective development of embryos produced by interspecies NT requires proper genomic activation of donor nucleus after overcoming the cytoplasmic control of recipient oocytes.

부화 및 착상에서 prostaglandins의 역할에 대한 연구는 많으나 배아단계에 따라 포배의 팽창과 부화에 미치는 영향에 대한 연구는 미미하다. 따라서 본 실험에서는 다양한 농도의 prostaglandins를 발생단계에 따라 처리하거나, indomethacin과 prostaglandins의 양 변화를 통해 팽창과 부화에 미치는 영향을 알아보고, 배아 단계별 PG 와 PG 의 생합성양 변화양상을 알아보았다. 100M이 상 농도의 PG 처리시 거의 모든