臺灣博碩士論文加值系統

團隊近年的研究發現IL-1β為二氫比啶（dihydropyridine）促進牙齦過度增生(dihydropyridine induced gingival overgrowth,　DIGO)的主要的促發炎細胞激素。在DIGO細胞中，IL-1β刺激後會增加IL-6和睪固酮受器(androgen receptor,AR)之mRNA表現。而分別抑制IL-6或抑制AR皆會降低總膠原蛋白(total collagen)的生成量，並降低結締組織生長因子(Connective tissue growth factor,CTGF)或第一型膠原蛋白(type I collagen)的mRNA表現。故IL-6與AR是影響牙齦增生機轉中的重要因素。本研究欲探討牙齦增生細胞之IL-6/gp130/STAT3訊息傳遞鍊與AR之關聯。實驗方法將健康的牙齦纖維母細胞與DIGO細胞，培養在charcoal stripped serum中，分別以不同濃度與不同刺激時間的IL-6刺激，進行西方墨點法與雷射共軛焦顯微鏡檢測STAT3的磷酸化與AR的蛋白質表現與位置。結果顯示健康的牙齦纖維母細胞與DIGO細胞STAT3磷酸化皆於15分鐘內達到高峰，且DIGO細胞受IL-6刺激產生的STAT3磷酸化作用較強。AR受IL-6刺激後的反應在本刺激條件下未產生顯著差異。需進行更進一步的實驗探討STAT3與AR之間的交互作用。

Our previous studies found that IL-1βis the main cytokine which induces Dihydropyridine Induced Gingival Overgrowth (DIGO) in patient with hypertension. Besides, there were more mRNA expressions of IL-6 and androgen receptor(AR) after stimulation of IL-1β in DIGO cells than healthy cells. Therefore, we supposed that IL-6 and AR are main factors of dihydropyridine induced gingival overgrowth. The propose of this study is to find the relationship of IL-6/STAT3 signaling pathway and AR in DIGO cells. Two types of gingival fibroblast, DIGO cells and Healthy cells, were cultured in charcoal stripped serum and treated with different time and dose of IL-6 cytokine. The result of Western blot and Laser confocal microscopy shows that IL-6 induced STAT3 phosphorylation was occurred in 15 minutes and the phosphorylation peak is between 3 and 15 minutes. The STAT3 phosphorylation is more significant in DIGO cells than healthy cells. However, the change of AR is not significant in this study. We have design further experiments to analysis the relationship between AR and IL-6/STAT3 signaling pathway.

目錄
中文摘要 ………………………………………….....……………. 1
英文摘要 ………………………………………….....……………. 2

第一章 研究動機與目的…………………………..……………. 3
第二章 文獻回顧
第一節 Dihydropyridine引發之牙齦增生之文獻回顧…..….. 4
第二節 Nifedipine及IL-1β刺激於牙齦纖維母細胞之研究 5
第三節.IL-6在慢性牙周炎中調控發炎反應之角色…………....6
第四節.IL-6 與 gp130/JAK2/STAT3訊息傳遞路徑………….. 7
第三章 研究材料與方法
第一節 細胞培養材料…………………………………………..8
第二節 實驗藥品………………………………………………. 9
第三節 細胞來源……………………………………………… 10
第四節 蛋白質濃度定量……………………………………….11
第五節 西方墨點法(Western blot)……………………………..12
第六節 雷射共軛焦顯微鏡…………………………………….14


第四章 實驗結果
第一節 IL-6刺激不同時間STAT3的磷酸化與AR的相關性..15
第二節 Anit-IL-6 Antibody對內生性IL-6的中和反應…....16
第三節 不同濃度IL-6對STAT3磷酸化與AR的影響…….…17
第四節 雷射共軛焦顯微鏡之結果……………..…………….…18
第五章 討論與未來展望
第一節 IL-6引發STAT3之磷酸化………….…………..…….19
第二節 IL-6與AR之相關性………………….………….…...20
第三節 Anti IL-6抗體中和內生性IL-6中和反應……….…...21
第六章 結論…………………………………………………….…. 24
參考資料
附錄
表1 牙齦纖維母細胞來源……….……..……...................30
圖1-1~1-3 西方墨點法之結果………….…..…….......................31.
圖2-1~2-6 雷射共軛焦顯微鏡之結果….……………………….34
圖3 實驗假說圖…………………………………………...40
圖4 未來實驗設計模式圖…………………………………41

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