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Title: | 檳榔鹼Arecoline經由TGF-β訊息傳遞路徑誘導人類頰黏膜纖維母細胞結締組織生長因子表現之研究 Arecoline-induced CCN2 expression in human buccal mucosal fibroblasts via TGF-β signaling pathway |
Authors: | Yi-Ting Deng 鄧伊婷 |
Advisor: | 郭彥彬 |
Keyword: | 口腔黏膜下纖維化症,檳榔鹼,結締組織生長因子,轉化生長因子β,薑黃素, Arecoline,CCN2,Curcumin,Oral submucous fibrosis,TGF-β, |
Publication Year : | 2012 |
Degree: | 博士 |
Abstract: | 結締組織生長因子(CTGF/CCN2) 在許多人類器官組織的纖維化病變中扮演了重要角色。口腔黏膜下纖維化症 (OSF)是一種不可逆的口腔黏膜纖維性病變。流行病學研究指出,嚼檳榔是口腔黏膜下纖維化症的最主要致病因子。本研究首先利用免疫組織化學染色方法,探討CCN2於20例OSF及10例正常口腔黏膜(NOM)中的表現。 結果發現在所有的 OSF 檢體中都有 CCN2的陽性染色,主要表現於纖維母細胞及內皮細胞,NOM 則幾乎沒有表現。以檳榔鹼 arecoline 刺激人類頰黏膜纖維母細胞(BMFs),可以誘導BMFs 中CCN2表現。Arecoline 刺激CCN2過度表現可能是嚼食檳榔誘導OSF發生的機轉之一。前處理 NF-κB專一性抑制劑Bay 117082、JNK專一性抑制劑 SP600125、p38 MAPK專一性抑制劑SB203580、抗氧化劑N-acetylcysteine (NAC) 都可以明顯的減少BMF中arecoline誘導CCN2的表現。因此Arecoline可能透過活性氧分子群 ( Reactive oxygen species,ROS),NF-kB,JNK及p38 MAPK 路徑誘導BMFs中CCN2的表現。薑黃素(curcumin)可以幾乎完全阻斷BMF中arecoline誘導的CCN2表現。此外TGF-β 中和抗體1D11、TβRI/ALK5的專一性抑制劑SB431542、Smad3的專一性抑制劑SIS3也可以抑制BMF中arecoline誘導的CCN2 表現。Arecoline可以誘導BMF中Smad2 和 Smad3 的磷酸化。可見得arecoline是經由TGF-β訊息傳導路徑誘導BMF的CCN2表現。此外NADPH氧化酶(NOX)的活化及活性氧物種被認為和許多人類器官組織的纖維化病變有關。我們利用泛NOX抑制劑diphenyleneiodonium chloride (DPI)、NOX4抑制劑plumbagin 及NAC前處理細胞可抑制BMF中arecoline誘導的CCN2 表現,但NOX2抑制劑 apocynin及muscarinic接受器拮抗劑 atropine 則沒有影響。可見得NOX4亦參與arecoline誘導BMF的CCN2訊息傳導路徑。本研究結果提供嚼食檳榔誘導OSF發生的新機轉及其調控方式。冀望未來可以藉由抑制CCN2來達到抑制或治療口腔黏膜下纖維化症。 Connective tissue growth factor (CCN2) is associated with the onset and progression of fibrosis in many human tissues. Areca nut (AN) chewing is the most important etiological factor in the pathogenesos of oral submucous fibrosis (OSF). We first immunohistochemically examined the expression of CCN2 protein in 20 cases of OSF and found positive CCN2 staining in fibroblasts and endothelial cells in all cases. Western blot analysis showed the arecoline, a main alkaloid found in AN, stimulated CCN2 synthesis in a dose- and time- dependent manner in buccal mucosal fibroblasts. Constitutive overexpression of CCN2 during AN chewing may enhance the fibrotic activity in OSF and play a role in the pathogenesis of OSF. Pretreatment with NF-κB inhibitor Bay 11-7082、JNK inhibitor SP600125、p38 MAPK inhibitor SB203580、antioxidatn N-acetyl-L-cycteine, but not ERK inhibitor PD98059, significantly reduced arecoline-induced CCN2 synthesis. Curcumin completely inhibited arecolined-induced CCN2 synthesis and the inhibition is dose-dependent. Arecoline also induced Smad 2 and Smad 3 activation. TGF-β neutralizing antibody、activin receptor-like kinase 5 (ALK5) inhibitor SB431542、Smad 3 inhibitor SIS3 significantly reduced arecoline-induced CCN2 synthesis. Furthermore, arecolined-induced CCN2 synthesis was inhibited by diphenyleneiodonium (DPI), a NADPH oxidase (NOX) inhibitor, NOX4 inhibitor plumbagin, but not muscarinic receptor antagonist atropine and NOX2 inhibitor apocynin. These results indicate that arecoline induced CCN2 expression in buccal mucosal fibroblasts via TGF-β signaling pathway. Curcumin may serve as a useful agent in controlling OSF. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/16143 |
Fulltext Rights: | 未授權 |
Appears in Collections: | 臨床牙醫學研究所 |
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