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研究生:張雅雯
研究生(外文):Ya-Wen Chang
論文名稱:探討子宮內膜異位症病患中披衣菌感染的發生率及細胞附著的分子機轉
論文名稱(外文):Chlamydia trachomatis Infection and Cell Adhesion in Patient with Endometriosis
指導教授:楊維中楊維中引用關係
指導教授(外文):Wei-Chung Vivian Yang
學位類別:碩士
校院名稱:臺北醫學大學
系所名稱:生物醫學材料研究所
學門:工程學門
學類:生醫工程學類
論文種類:學術論文
論文出版年:2007
畢業學年度:95
語文別:中文
論文頁數:68
中文關鍵詞:砂眼披衣菌子宮內膜異位症細胞外間質細胞黏著分子
外文關鍵詞:Chlamydia trachomatisEndometriosisExtracellular matrixCell adhesion moleculesSPARC
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子宮內膜異位症(Endometriosis)是常見的慢性婦科疾病,好發於生殖年齡的婦女,其定義為在骨盆腹膜或子宮腔外發現子宮內膜的腺體(Gland)與基質細胞(Stroma cell),其臨床症狀最常見的為骨盆腔疼痛及不孕。至目前為止,其致病機轉仍不清楚,最被廣為接受的理論為經血逆流理論(Retrograde menstruation),也就是經血逆行到腹腔。砂眼披衣菌(Chlamydia trachomatis)是常見的細菌性性傳染病,常造成婦女骨盆腔疼痛及子宮、輸卵管沾黏,與子宮內膜異位症部分病徵相似,因此本論文研究第一部分在探討子宮內膜異位症患者的披衣菌感染盛行率,以患者腹腔液檢體進行披衣菌的ELISA及PCR檢測,發現患者呈現披衣菌陽性反應的百分比較對照組高。同時以子宮內膜異位症患者腹腔液檢測披衣菌的感染,陽性反應的百分比較傳統利用陰道檢測的方式來得高,推測子宮內膜異位症可能與披衣菌上行感染有關。
子宮內膜異位症雖為良性疾病,卻有沾粘、侵犯和轉移等類似癌症的病理特性,而細胞間的貼附和移動又與細胞外間質(Extracellular matrix, ECM)及細胞黏著分子(Cell adhesion molecules, CAM)有關,因此本論文研究第二部分探討子宮內膜異位症患者的原位及異位子宮內膜組織,與正常婦女的子宮內膜組織的細胞外間質與細胞黏著分子的表現有無差異,利用ECM與CAM微陣列(Microarray)實驗及基因表現分析子宮內膜組織,並進一步用西方墨點法做蛋白質表現分析後發現,一些細胞外間質及細胞黏著分子的表現的確在異位的子宮內膜組織與原位及正常婦女的子宮內膜組織比較有差異。此外,一種存在於細胞外間質的醣蛋白SPARC (Secreted protein acidic and rich in cysteine),於患者異位的子宮內膜組織中表現較正常婦女的子宮內膜組織明顯,而先前並無相關文獻探討SPARC在子宮內膜異位症患者的異位組織中的表現,因此,本研究將著重在SPARC在子宮內膜異位症形成的過程中可能扮演的角色作進一步探討。
Endometriosis is a common chronic gynecological disease affecting the female population during their reproductive life. The definition of endometriosis is presence of endometrial glands and stroma within the pelvic peritoneum and other extrauterine sites. The common clinical symptoms of endometriosis are pelvic pain and infertility. Until now, the pathogenesis of endometriosis remains poorly defined. The most widely accepted theory is Sampson''s theory of retrograde menstruation. Chlamydia trachomatis is the most common cause of bacterially-acquired sexually transmitted disease. It can induce pelvic pain, uterus and fallopian tubes adhesion formation. The symptoms caused by Chlamydia trachomatis infection are similar to endometriosis. For this reason, the study aims to investigate the prevalence of Chlamydia trachomatis in women with endometriosis. Peritoneal fluids, obtained from patient were assayed for Chlamydia trachomatis by ELISA and PCR. The percentage of positive was higher in peritoneal fluids from women with endometriosis than in the control. Our studies demonstrated that genital tract test for Chlamydia trachomatis may not sufficient to identify the infection of Chlamydia trachomatis in patients with endometriosis. Endometriosis maybe related to the upper infection of Chlamydia trachomatis.
Although endometriosis is a benign disease, its properties of adhesion, invasion and metastasis are similar to malignant tumor. Extracellular matrix (ECM) and cell adhesion molecules (CAM) are involved in cell adhesion and movement. Therefore, the second part of this study aims to analyze the differences in the expressions of ECM and CAM between eutopic and ectopic endometrium in patients with endometriosis and their expressions compared to women without endometriosis. Microarry and RT-PCR were used to analyze the mRNA expression pattern in endometrium. Western blot analyses were carried out to confirm the identified ECM and CAM expression in the endometrium. The results showed that SPARC (Secreted protein acidic and rich in cysteine), an extracellular glycoprotein, was expressed significantly in ectopic endometrium in patients with endometriosis compared to eutopic endometrium in the control women without endometriosis. The increased expression of SPARC in ectopic endometrium with endometriosis has not been reported in the literature. The possible roles of SPARC play in the progression of endometriosis are discussed.
目錄 I
圖表目錄 III
縮寫表 IV
摘要 1
ABSTRACT 3
一、前言 5
(一)、子宮內膜異位症(ENDOMETRIOSIS) 5
(二)、砂眼披衣菌(CHLAMYDIA TRACHOMATIS) 7
(三)、細胞外間質與細胞黏著分子(EXTRACELLULAR MATRIX AND CELL ADHESION MOLECULES) 9
(四)、微陣列(MICROARRAY) 10
二、研究目的(PURPOSES) 11
第一部分 砂眼披衣菌的感染 11
第二部分 細胞黏著分子之探討 12
三、第一部分 砂眼披衣菌的感染(CHLAMYDIA TRACHOMATIS INFECTION) 13
(一)、材料與方法 13
1. 樣本收集(Sample preparation) 13
2. 蛋白質分析(Protein analysis) 14
3. 核酸分析(Nucleic acid analysis) 15
4. 萃取子宮內膜組織RNA (Isolation of RNA from endometrium) 16
5. 反轉錄聚合酶鏈鎖反應 (Reverse-transcription polymerase chain reaction) 17
6. 統計分析方法(Statistical analysis methods) 18
(二)、結果 19
1. 披衣菌蛋白質於子宮內膜異位症患者腹腔液中的表現分析 19
2. 以PCR技術進行子宮內膜異位症患者腹腔液中披衣菌核酸分析表現 21
3. 以反轉錄聚合酶鏈鎖反應分析子宮內膜異位症患者內膜組織中披衣菌核酸表現 22
4. 以接受者操作特徵曲線設定以ELISA檢測子宮內膜異位症患者腹膜液中披衣菌脂多醣濃度之閾值 23
(三)、討論 24
(四)、結論 25
四、第二部分 黏著分子與細胞侵犯(ADHESION MOLECULAR AND CELL INVASION) 26
(一)、材料與方法 26
1. 樣本收集(Sample preparation) 26
2. 微陣列及基因表現分析(Microarray and gene expression analysis) 27
3. 反轉錄聚合酶鏈鎖反應(Reverse transcription-polymerase chain reaction) 28
4. 西方墨點法(Western blot) 29
5. 子宮內膜細胞初代培養(Primary culture of endometrial cells) 31
(二)、結果 33
1. 微陣列及基因表現分析 33
子宮內膜異位症病患的異位子宮內膜組織與正常子宮內膜組織比較 33
子宮內膜異位症病患的異位子宮內膜組織與原位子宮內膜組織比較 34
2. SPARC基因表現 36
3. 子宮內膜細胞初代培養結果 38
(三)、討論 39
五、未來研究方向 42
六、參考文獻 43
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9.Keegan, H., et al., Comparison of DNA extraction from cervical cells collected in PreservCyt solution for the amplification of Chlamydia trachomatis. Cytopathology, 2005. 16(2): p. 82-7.
10.van Susante, J.L.C., et al., Linkage of chondroitin-sulfate to type I collagen scaffolds stimulates the bioactivity of seeded chondrocytes in vitro. Biomaterials, 2001. 22(17): p. 2359-69.
11.Yan, Q., et al., Absence of SPARC in murine lens epithelium leads to increased deposition of laminin-1 in lens capsule. Invest Ophthalmol Vis Sci, 2005. 46(12): p. 4652-60.
12.Arnold, J.T., et al., Endometrial stromal cells regulate epithelial cell growth in vitro: a new co-culture model. Hum Reprod, 2001. 16(5): p. 836-45.
13.Wu, M.H., et al., Distinct regulation of cyclooxygenase-2 by interleukin-1beta in normal and endometriotic stromal cells. J Clin Endocrinol Metab, 2005. 90(1): p. 286-95.
14.Wu, M.H., et al., The differential expression of intercellular adhesion molecule-1 (ICAM-1) and regulation by interferon-gamma during the pathogenesis of endometriosis. Am J Reprod Immunol, 2004. 51(5): p. 373-80.
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17.Murphy-Ullrich, J.E., et al., SPARC mediates focal adhesion disassembly in endothelial cells through a follistatin-like region and the Ca(2+)-binding EF-hand. J Cell Biochem, 1995. 57(2): p. 341-50.
18.Said, N., I. Najwer, and K. Motamed, Secreted protein acidic and rich in cysteine (SPARC) inhibits integrin-mediated adhesion and growth factor-dependent survival signaling in ovarian cancer. Am J Pathol, 2007. 170(3): p. 1054-63.
19.Nomura, S., et al., Developmental expression of 2ar (osteopontin) and SPARC (osteonectin) RNA as revealed by in situ hybridization. J Cell Biol, 1988. 106(2): p. 441-50.
20.Reed, M.J., et al., Differential expression of SPARC and thrombospondin 1 in wound repair: immunolocalization and in situ hybridization. J Histochem Cytochem, 1993. 41(10): p. 1467-77.
21.Motamed, K. and E.H. Sage, Regulation of vascular morphogenesis by the matricellular protein SPARC. Kidney Int, 1997. 51(5): p. 1383-7.
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24.Koblizek, T.I., et al., Angiopoietin-1 induces sprouting angiogenesis in vitro. Curr Biol, 1998. 8(9): p. 529-32.
25.Yasunaga, C., Y. Nakashima, and K. Sueishi, A role of fibrinolytic activity in angiogenesis. Quantitative assay using in vitro method. Lab Invest, 1989. 61(6): p. 698-704.
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