臺灣博碩士論文加值系統

近幾年的研究指出，約有12~15%已婚夫婦患有不孕症，近一半的病例包含男性因素。仍然有25%男性不孕症的病因不清楚，基因遺傳變異為導致的可能因素。實驗室藉由cDNA微陣列分析，發現於不孕症的睪丸組織中，SEPTIN12基因表現量明顯下降。SEPTIN12/Septin12專一性表現於人類與小鼠睪丸組織中。建立Septin12基因剔除嵌合小鼠，發現於生殖的表現性中，如睪丸重量、精子數目、精子活動力皆有下降情形。於男性不孕個案中帶有SEPTIN12 c.474 G>A，具有精子形態異常的表現性，如圓形頭部、彎曲的頸部與尾巴，精子頭部也會出現DNA損壞。於個案中帶有SEPTIN12 c.589G>A (D197N)，具有精子環節結構缺損的情形，並且缺乏SEPT12的表現性。根據先前研究，說明Septin12基因會參與哺乳類的精子分化過程。然而，SEPT12失去功能性時，造成精子結構異常的機制尚待釐清。我們假設SEPT12會組合成複合體，共同參與精子的分化，所以藉由酵母菌雜交法，鑑別出與SEPT12結合的蛋白質。
於本篇研究中，初步針對鑑別的SEPT12結合蛋白質進行表現分析，主要選擇核孔蛋白NDC1進行後續研究。首先，證實NDC1表現於人類睪丸組織與成熟精子，SEPT12與NDC1於精子頸部的表現位置具有相似性。於精子的分化過程，內生性NDC1表現於精細胞核膜的周圍，並參與精子頭部與頸部的結構生成。於男性生殖細胞株模式中，說明SEPT12與NDC1具有結合性，當大量表現NDC1，影響SEPT12功能性絲狀結構的組成。於動物模式中，發現SEPT12(D197N)突變小鼠，影響NDC1於精子結構的表現位置。於臨床檢體方面，NDC1於SEPTIN12 c.474 A/A基因型精子的表現情形中，發現會不完整圍繞於精細胞核膜位置。這些研究有助於我們了解SEPT12/NDC1複合體參與精子分化過程的角色。

In recent years, it is calculated that 12~15% of couples are infertile. The male factors cause of infertility approach about half of the cases. Genetic variants are related to male infertility. Our previous studies indicated that SEPTIN12/Septin12 is expressed specifically in human and mice testis tissue. The reproductive phenotypes of Septin12+/- chimera mice included lower testis weighs, decreased sperm count, decreased sperm motility and increased abnormal sperm morphology. Infertility men carrying SEPTIN12 c.474 G>A variants were presented with abnormal morphology of spermatozoa, including round head, bent neck and bent tail. In addition, the sperm head shows severe DNA damage. Another, infertility men carrying SEPTIN12 c.589G>A (D197N) presented with defective annulus, and SEPT12 signal was loss from the annulus. However, the mechanisms of dysfunctional SEPT12 induced abnormal morphology of spermatozoa are still unclear. We screened SEPT12-interacted proteins from yeast 2-hybrid system. In this study, expression and location of SEPT12-interactors have been confirmed in testis and mature spermatozoa. We choose nucleoporin NDC1 for further analysis. In mature human spermatozoa, SEPT12 colocolizes with NDC1 at the neck region. During spermiogenesis, we found that endogenous nucleoporin NDC1 located at nuclear membrane and participated in sperm head and neck formation. Over-expression of NDC1 repressed the formation of SEPT12 filament structure in male germ cell lines. In animal model, mutated SEPT12(D197N) mice reveals the abnormal localization of NDC1 in mature sperm. In clinical aspect, NDC1 is located at broken nuclear membrane of spermatozoa from case with SEPTIN12 c.474 G>A. These results provides us that spermatogenic roles of SEPT12/NDC1 complexes in mammalian spermiogenesis.

中文摘要 III
Abstract IV
表目錄 VII
圖目錄 VIII
第一章、緒論 1
第二章、實驗材料與方法 10
第三章、實驗結果 20
第四章、討論 25
第五章、結論 29
參考文獻 30

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