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研究生:陳南妮
研究生(外文):NanNi Chen
論文名稱:一氧化氮合成在大鼠及小鼠著床旁期的胚胎中分布情形
論文名稱(外文):Distribution of Nitric Oxide Synthases in the Peri-implantation Embryos of Rats and Mice
指導教授:劉紹東
指導教授(外文):S. T. Liu
學位類別:碩士
校院名稱:高雄醫學大學
系所名稱:醫學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2001
畢業學年度:89
語文別:中文
論文頁數:56
中文關鍵詞:大鼠小鼠著床旁期滋養層細胞一氧化氮一氧化氮合成
外文關鍵詞:RatMousePeri-implantationTrophoblastNitric Oxide( NO )Nitric Oxide Synthases( NOS )
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囓齒類動物的胚胎,若要能成功的進行著床,是需要胚胎與子宮之間進行一連串的交互作用後,才能達成的;且胚胎需發育至囊胚期,而子宮則需達到一可接受著床的狀態。著床的過程可分為三個階段:接觸、附著及鑽入。在這三個著床的階段中,可發現囊胚滋養細胞與子宮內膜之間特有的互動情形;例如,在接觸的階段中,胚胎在被包於透明帶的情況下,會誘發鄰近的子宮內膜產生局部性的蛻膜化反應及血管通透性增加;接著在附著的位置上會產生子宮內膜上皮細胞凋亡的現象,而這些凋亡的上皮細胞會被胚胎滋養層細胞以類似巨噬細胞的吞噬作用清除;並且在鑽入時胚胎滋養層細胞會分泌消化細胞間基質的酵素,以調節胚胎的鑽入及子宮組織的重塑。然而,著床反應的詳細調節機轉,至目前仍未清楚確立。
一氧化氮已被發現在數種生理機制中擔任重要的調節者:其可調節血管活性、平滑肌收縮、神經訊息的傳遞和結締組織的重塑。一氧化氮在細胞內是經由一氧化氮合成的催化而產生。而一氧化氮對於胚胎發育過程之影響,目前多偏重於著床後期的研究,但在著床過程中,胚胎卻已表現出許多與一氧化氮功能有關的現象。因此,本實驗的目的即欲探討大鼠及小鼠在著床旁期的胚胎內三種一氧化氮合成( endothelial NOS, inducible NOS and neuronal NOS )的分布情形,及比較大鼠及小鼠之間一氧化氮合成分布的差異。本實驗是利用冷凍切片方式處理,再以免疫組織化學染色法來觀察在大鼠及小鼠的胚胎中三種一氧化氮合成的表現。實驗結果發現,一氧化氮合成確實會出現在大鼠及小鼠著床過程中的胚胎內。在大鼠方面,除了在懷孕第五天、第六天、第八天沒有endothelial NOS的表現外,其餘二種一氧化氮合成均可觀察到表現於著床旁期之大鼠胚胎中;在小鼠方面,三種一氧化氮合成都出現在小鼠的胚胎中。雖然個別一氧化氮合成在著床旁期大小鼠胚胎上所出現的區域有些微的差異,但發現一氧化氮合成大多共同分布在由滋養層所分化出來的細胞內,如,胎盤外錐體、胚體外內胚層、胚體外外胚層及滋養層巨細胞等。
因為滋養層細胞在著床過程中實際負責與子宮細胞的接觸,而且將分化形成胚外膜及部份的胎盤,因此,我們推論,滋養層細胞內一氧化氮合成的產物(即一氧化氮),對於著床過程的進行應扮演著一特殊及重要的角色。
關鍵字:大鼠,小鼠,著床旁期,滋養層細胞,一氧化氮,一氧化氮合成

Successful implantation of rodent embryos is strictly dependent on the synchrony and coordinated interactions between a uterus that is “receptive” and an embryo that has reached the “blastocyst stage” of development.
Implantation has been divided into three progressive processes, i.e., apposition, attachment and penetration, according to the intimacy between blastocyst trophoblast and endometrial epithelium. Not only morphological approach, characteristic trophoblast-uterine interactions can be found in all these stages of implantation. For example, localized decidualization and increase of endometrial vascular permeability appear during apposition, phagocitize apoptic endometrial epithelium as embryos attach uterus at sites of attachment, and production of matrix metalloproteases ( MMPs ) as embryos move into the endometrium during penetration. However, mechanisms in regulation of implantation have not yet been clearly defined.
Nitric oxide ( NO ) has been come to prominence recently as a major mediator in numerous biological processes and is responsible for altering vasoactivity, smooth muscle contraction, neurotransmission and connective tissue remodeling. NO is a biological free radical and is produced by cytoplasmic nitric oxide synthases ( NOS ). It has also been shown that cytokines, prostaglandins, MMPs and hormones all may play a role, alone or in combination, in altering NO concentration.
Sicnce the correlations between the processes of implantation and the functions of NO, we, therefore, designed the following experiments to test the existence and to localize the distribution of three isoforms of NOS ( i.e. endothelial NOS, inducible NOS, and neuronal NOS ) in embryos of rats and mice during the processes of implantation.
Samples of embryo were collected from females of day 5, 6, 7and 8 of pregnancy, and processed by methods of frozen section and by immunocytochemistry.
Results showed that nitric oxide synthases were present in peri-implantation embryos of rats and mice. In rat, exceptive the endothelial NOS was not found in day 5, 6 and 8 of pregnancy, the other two NOS were present in the implanting embryos. In mice, all three NOS isoforms were present in the embryos of day 5, 6, 7 and 8 of pregnancy.
Although minor differences exist in the distribution among three NOS between rats and mice, we found that the most detected NOS were localized in the trophoblast-derived cells; i.e. ectoplacental cone, extraembryonic endoderm, extraembryonic ectoderm and trophoblast giant cells.
Since the trophoblast cells are responsible for contacting uterine cells, differentiating extraembryonic membranes, and establishment of placenta, presence of NOS in three cells suggests that the product of NOS is involved and regulate the interactions between embryos and uterus during the processes of implantation.
Keywords: rat, mouse, peri-implantation, trophoblast, nitric oxide ( NO ), nitric oxide synthases ( NOS )

中文摘要…………………………………………1-2
英文摘要…………………………………………3-5
緒論………………………………………………6-14
研究材料與方法
動物準備……………………………15
組織的收集…………………………16
免疫組織化學染色法………………17-18
結果………………………………………………19-22
討論………………………………..……………23-27
參考文獻…………………………………………28-33
圖一………………………………………………34
表一………………………………………………35
表二………………………………………………36
圖二………………………………………………37-38
圖三………………………………………………39-40
圖四………………………………………………41-42
圖五………………………………………………43-44
圖六………………………………………………45-46
圖七………………………………………………47-48
圖八………………………………………………49-50
圖九………………………………………………51-52
圖十………………………………………………53-54
圖十一……………………………………………55-56

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