Effect of Fibroblast growth facter-7 on in vitro Maturation of Porcine Cumulus-Oocyte Complexes and Subsequent Developmental Potential after Parthenogenetic Activation
Effect of Fibroblast growth facter-7 on in vitro Maturation of Porcine Cumulus-Oocyte Complexes and Subsequent Developmental Potential after Parthenogenetic Activation
- 발행기관 충북대학교
- 지도교수 현상환
- 발행년도 2023
- 제출일 2023-07-10
- 학위수여년월 2023. 8
- 학위명 석사
- 학과 및 전공 일반대학원 수의학과(원)
- 원문페이지 44
- 실제URI http://www.dcollection.net/handler/chungbuk/200000771956
- UCI I804:43009-000000058563
- 본문언어 영어
- 저작권 충북대학교 논문은 저작권에 의해 보호됩니다
초록/요약
In vitro generation of porcine embryos is an indispensable method in the realms of both agriculture and biomedicine. Nonetheless, the extant procedures encounter substantial obstacles pertaining to both the caliber and efficacy of the produced embryos, necessitating extensive research to in vitro maturation (IVM), the seminal commencement phase. One potentially fruitful approach may lie in refining the composition of the media and supplements utilized for oocyte maturation. Fibroblast growth factor-7 (FGF7), alternatively termed keratinocyte growth factor, is a theca-derived cytokine integral to folliculogenesis. This study's objective was to examine the ramifications of supplementing FGF7 during the IVM phase. To determine the location of FGF7 and its receptor in porcine ovaries, immunohistochemistry was executed based on follicle size categories (1–2, 3–6, and 7–9 mm). Regardless of follicle size, it was determined that FGF7 was expressed in theca and granulosa cells (GCs), while the FGF7 receptor was only expressed in the GCs of the larger follicles. During the IVM process, the maturation medium was supplied with various concentrations of FGF7, aiming to mature porcine cumulus-oocyte complexes (COCs). The results indicated a significant augmentation in the nuclear maturation rate only within the group treated with 10 ng/mL of FGF7. Post-IVM, the diameter of the oocytes exhibited a significant expansion in all groups that received FGF7 supplementation. Additionally, all FGF7-supplemented groups exhibited a substantial elevation in intracellular glutathione levels, coupled with a noticeable reduction in reactive oxygen species levels. With respect to gene expressions related to apoptosis, FGF7 treatment elicited a downregulation of pro-apoptotic genes and an upregulation of anti-apoptotic genes. The expression of genes associated with antioxidants underwent a significant enhancement. In terms of the FGF7 signaling pathway-associated genes, there was a considerable elevation in the mRNA expression of ERK1, ERK2, c-kit, and KITLG. Remarkably, the group of 10 ng/mL of FGF7 demonstrated an appreciable uptick in the blastocyst formation rate during embryonic development post-parthenogenetic activation. In conclusion, the supplementation of FGF7 during IVM substantially augments the quality of matured oocytes and facilitates the subsequent development of parthenogenetically activated embryos. These results provide a new perspectives on improving the in vitro maturation of porcine oocytes. Keywords: cytokine, cumulus-oocyte complexes, embryonic development, fibroblast growth factor 7, parthenogenetic activation, porcine embryos
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Ⅰ. INTRODUCTION 2
Ⅱ. MATERIALS AND METHODS 4
1. Fluorescent immunohistochemical analysis of the ovary 4
2. Oocyte collection and in vitro maturation 5
3. Evaluation of oocyte diameter 6
4. Evaluation of nuclear maturation 8
5. Measurement of intracellular glutathione GSH and ROS levels 8
6. Quantitative reverse transcription polymerase chain reaction 8
7. Parthenogenetic activation 12
8. Developmental competence evaluation and total blastocyst cell count 12
9. Statistical analysis 12
Ⅲ. RESULTS 13
1. FGF7 and its receptor localized in porcine ovarian follicular 14
2. Effect of FGF7 supplementation during IVM on oocyte nuclear maturation 16
3. Effect of FGF7 supplementation during IVM on oocyte diameter 18
4. Effect of FGF7 supplementation during IVM on intracellular GSH and ROS
levels 20
5. Effect of FGF7 supplementation on gene expression levels in CCs and oocytes
during IVM 23
6. Effect of FGF7 supplementation in IVM media on developmental potential
after PA 27
Ⅳ. DISCUSSION 29
Ⅴ. REFERENCES 34