臺灣博碩士論文加值系統

黃體素 ( Progesterone, P4 ) 是一種類固醇激素。在懷孕期間、女性月經週期和著床皆有重要的角色。而P4必須與黃體素接受器 ( Progesterone receptor, PR ) 結合才能引發作用，若P4與黃體素接受器拮抗劑 ( Mifepristone, RU486 ) 結合會造成P4無法發揮其功能。在先前的P4研究較著重於對子宮內膜的影響而不是胚胎發育的影響，在本研究中我們將探討P4在小鼠胚胎發育和著床期間之影響，故利用不同濃度RU486 ( 2x10-9、2x10-7、2x10-5M ) 共同培養，去探討P4作用被抑制後對小鼠囊胚之胚胎發育與著床後之影響。藉由免疫螢光染色檢測PR在胚胎不同階段的表現，由結果發現PR在受精卵、著床前胚胎到囊胚期都被偵測到。利用放射免疫測定法 ( Radioimmunoassay ) 偵測到在懷孕期間P4會存在於小鼠的血清、輸卵管與子宮腔內。囊胚共同培養人類臍帶血清 (human Cord Serum, hCS )中給予RU486 ( 2x10-5M ) 24小時與給予RU486 ( 2x10-5M ) 48小時後對囊胚內細胞數有明顯減少( P <0.001 ) ; 在著床後的實驗中，而給予RU486 48小時後，8天的胚胎發育，胚胎可以到達後期階段有減少的趨勢 ( P <0.05/ P <0.001 )。我們的研究數據明顯表示P4經由胚胎內PR的結合去參與著床前與著床後胚胎發育的生理功能。

Progesterone, a steroid hormone plays an important role in gestation, female menstrual cycle and implantation. Must be P4 and progesterone receptor combination can trigger action. If P4 and PR antagonist-RU486 binding will cause P4 can not its function. Previous studies have focused more on the effect of progesterone on the endometrium of uterus rather than on embryo development. In this study, we investigated the roles of progesterone in mouse embryo development and implantation. We examined the effects of RU486 (2x10-9 to 2x10-5M) on mouse embryonic cell growth and blastocyst outgrowth in vitro. Expression of PR on embryo was examined by immunofluorescence staining. We found that expression of PR was detected on the stages of preimplantation embryo from zygotes to blastocysts. The presence of progesterone in reproductive tract was determined by Radioimmunoassay (RIA). The progesterone level could be found in the serum, fallopian tube and uterine cavity. Our results showed that RU486-pretreated blastocysts with human cord serum (hCS) or treatment 48 hours a significant reduction the number of cells (P <0.001 ). The RU486-pretreated 48 hours or 8 days blastocysts were allowed to implant in vitro, significantly fewer embryos were able to reach a later stage of embryo development (P <0.05/ P <0.001 ). Our results suggested that the expression
of PR on preimplantation embryos was involved in the physiological roles in the embryo development in the time of or after implantation.

目錄
摘要 I
Abstract II
謝誌 IV
圖目錄 VII
表目錄 IX
第一章 前言 1
第二章 文獻回顧 2
2.1 影響誘發排卵因子 2
2.1.1 妊娠馬血清促性腺素 2
2.1.2 人類絨毛膜促性腺激素 2
2.2 小鼠胚胎 3
2.2.1 胚胎發育過程 3
2.2.2 著床前胚胎型態 5
2.2.3 著床後胚胎型態 7
2.3黃體素 10
2.3.1黃體素結構來源 10
2.3.2黃體素接受器 11
2.3.3黃體素應用 12
2.3.4黃體素的生理角色 13
2.4 Mifepristone 15
第三章 材料與方法 16
3.1 實驗動物 16
3.1.1 實驗動物之飼養 16
3.1.2 囊胚之取得 16
3.2 P4之製備 19
3.3 RU486之製備 19
3.4 P4腔內濃度測試 19
3.4.1 P4腔內濃度步驟 19
3.5 免疫螢光染色 19
3.5.1 免疫螢光染色步驟 20
3.6 囊胚體外八天培養 23
3.6.1 囊胚體外八天培養步驟 23
3.7 區別染色 26
3.7.1 兔抗鼠血清製備 26
3.7.2 人類臍帶血清製備 26
3.7.3 區別染色之步驟 27
3.8 囊胚貼附後細胞增生染色 31
3.8.1 囊胚貼附後細胞增生染色步驟 31
第四章 統計分析 34
第五章 結果 35
5.1 P4在小鼠腔內濃度測試 35
5.1.1 小鼠血清、輸卵管、子宮 35
5.2 PR在小鼠著床前後胚胎之表現 39
5.3 P4對小鼠囊胚發育之影響 42
5.3.1 囊胚體外八天培養 42
5.3.2 囊胚內細胞團及滋養外胚層細胞 45
5.3.3 囊胚細胞增生染色 53
第六章 討論 56
第七章 結論 59
參考文獻 60
作者簡介 71

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