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Study on Antioxidant Capacity of Paralichthys Olivaceus Egg Extracts in Oxidative Stress Exposed Porcine Oocyte Maturation Development

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Author(s)
Park, Hyo Jin
Issued Date
2021
URI
https://dcoll.jejunu.ac.kr/common/orgView/000000010414
Abstract
In vitro maturation and embryonic development of the egg, compared to the body, the oocyte is damaged by reactive oxygen species (ROS) generated inside the cell. In this study, we investigated the effect of treatment with paralichthys olivaceus egg extract (POEE) on the maturation and development of oocytes exposed to oxidative stress during in vitro maturation of pig oocytes. As a result of analyzing the components of POEE, the content of vitamin B9 (folic acid, FA) was higher than that of other components, and the IC50 of POEE was calculated using the radical scavenging ability of DPPH. To create a peroxidative environment in pig eggs, 200 μM of H₂O₂ was treated, and immature pig eggs were treated with 0, 600, 650, 700, and 750 ppm POEE concentrations. As a preliminary test, FAs were treated with 0, 0.1, 0.2, 0.3, 0.4 mM (0 FA, 0.1 FA, 0.2 FA, 0.3 FA, 0.4 FA). The rate of cleavage and blastocyst formation in pig embryos was significantly increased at 0.3 FA. In the 650 POEE group, the survival rate and in the 700 POEE group, the cleavage rate increased significantly. In addition, the 650 POEE group significantly increased glutathione (GSH) levels compared to the control group, and the 0.3 FA group significantly reduced reactive oxygen species (ROS) activity compared to the control group. The 650 POEE group and the 0.3 FA group prevented abnormal spindle organization and chromosomal misalignment compared to the control group, and also significantly increased the total number of cells in the blastocyst phase and decreased apoptotic cells compared to the control group. These results show that POEE and FA reduce oxidative stress in pig eggs and improve their quality and developmental potential.
난자의 시험관내 성숙 및 배아 발달은 체내에 비해 세포 내부에서 생성되는 활성산소 (ROS)에 의해 난모세포가 손상된다. 본 연구에서는 돼지 난모세포의 체외 성숙 동안 광어알 추출물 (paralichthys olivaceus egg extract; POEE)의 처리가 산화 스트레스에 노출 된 난자의 성숙과 발달에 미치는 영향을 조사하였다. POEE 의 성분을 분석한 결과, 비타민 B9(엽산, FA) 의 함량이 다른 성분보다 높았으며, DPPH 의 라디칼 소거능을 이용하여 POEE 의 IC50 을 계산하였다. 돼지 난자에 과산화 환경을 조성하기 위해 200μM 의 H₂O₂를 처리하였고 미성숙 돼지 난자에 0, 600, 650, 700, 750 ppm POEE 농도를 처리했다. 예비 시험으로 FA 를 0, 0.1, 0.2, 0.3, 0.4 mM (0 FA, 0.1 FA, 0.2 FA, 0.3 FA, 0.4 FA)으로 처리하였다. 돼지 배아의 난할 및 배반포 형성 속도는 0.3 FA 에서 유의하게 증가했다. 650 POEE 그룹에서는 생존율이, 700 POEE 그룹에서는 난할율이 유의적으로 증가했다. 또한, 650 POEE 그룹은 대조군에 비해 글루타티온(GSH) 수치를 유의하게 증가시켰고, 0.3 FA 그룹은 대조군에 비해 활성산소종 (ROS) 활성을 유의하게 감소시켰다. 650 POEE 그룹과 0.3 FA 그룹은 대조군에 비해 비정상적인 방추 조직 및 염색체 오정렬을 예방했으며, 또한 대조군에 비해 배반포기의 총 세포 수를 유의하게 증가시키고 세포 사멸 세포를 감소시켰다. 이러한 결과는 POEE 와 FA 가 돼지 난자의 산화 스트레스를 줄이고 품질과 발달 잠재력을 향상시킴을 보여준다.
Alternative Title
산화 스트레스에 노출 된 돼지 난모세포 성숙에서 광어알 추출물의 항산화 효과 연구
Affiliation
제주대학교 대학원
Department
대학원 생명공학부
Advisor
박세필
Awarded Date
2021. 8
Table Of Contents
ABSTRACT 1
1. INTRODUCTION 2
2. MATERIALS AND METHODS 5
2.1. Chemicals and reagents 5
2.2. Preparation of Paralichthys olivaceus eggs extract 5
2.3. Analysis of DPPH radical scavenging ability 5
2.4. IVM and peroxidized of porcine oocytes 6
2.5. Folic acid treatment 7
2.6. POEE treatment 7
2.7. Parthenogenetic activation and embryo culture 7
2.8. Measurement of intracellular ROS and GSH levels 8
2.9. Immunofluorescence 8
2.10. TUNEL assay and hoechst staining 9
2.11. Statistical analysis 9
3. RESULTS 10
3.1. Analysis of POE Extract (POEE) Components 10
3.2. Extraction of Paralichthys olivaceus egg and DPPH analysis of folic acid 12
3.3. Effect of POEE & Folic acid treatment on maturation of porcine oocytes 15
3.4. POEE and FA reduces the level of ROS and increase the level of GSH in peroxidized porcine oocytes in vitro 19
3.5. POEE & FA prevents aberrant spindle organization and chromosome misalignment in peroxidized porcine oocytes in vitro 21
3.6. POEE & FA improves the developmental capacity and quality of embryos derived from peroxidation porcine oocytes in vitro 23
4. DISCUSSION 25
REFERENCES 28
ABSTRACT IN KOREAN 36
ACKNOWLEDGEMENT 37
Degree
Master
Publisher
제주대학교 대학원
Citation
Park, Hyo Jin. (2021). Study on Antioxidant Capacity of Paralichthys Olivaceus Egg Extracts in Oxidative Stress Exposed Porcine Oocyte Maturation Development
Type
Dissertation
Appears in Collections:
Faculty of Biotechnology > ETC
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