苯妥英經由乙型轉化生長因子訊息傳導路徑誘導人口腔上皮細胞LOXL2之表現

Phenytoin Induced LOXL2 Expression Through TGF-β signaling in Human Oral Epithelial Cells

胡家瑄(Jia-Xuan Hu)

指導教授 ： 郭彥彬

國立臺灣大學/醫學院/臨床牙醫學研究所/碩士(2021年)

https://doi.org/10.6342/NTU202100987

摘要

苯妥英引起之牙齦增生(Phenytoin-induced gingival overgrowth, PIGO)在癲癇患者中是常見的副作用，也是藥物引起牙齦增生中最具有纖維化特徵的一種，PIGO的產生會讓口腔衛生維持困難、影響美觀、咀嚼功能等問題造成患者的社交及進食困難。近期研究發現PIGO的組織中在結締組織及上皮有離氨基氧化酶樣蛋白2(Lysyl Oxidase-like protein 2, LOXL2) 大量表現，且LOXL2也因為具有穩定細胞外間質以及上皮間質轉化(Epithelial-mesenchymal transition, EMT)特性，而被認為可能是器官纖維化的治療標的。本研究目的擬探討Phenytoin是否可誘導LOXL2以及相關機轉。本研究中發現隨著Phenytoin處理的濃度及時間增加可使人類纖維母細胞(HGF)、人類口腔上皮細胞CA9-22、OECM-1的LOXL2的表現量也隨之增加，且Phenytoin可使CA9-22、OECM-1的上皮標誌因子中E-cad下降，間質標誌因子Vimentin及轉化因子Slug/Twist上升，而有EMT的現象。 LOX的抑制劑BAPN可以抑制這個現象。使用TGF-β路徑的抑制劑（TGF-β中和抗體、SIS3、SB431542）處理HGF、CA9-22、OECM-1發現可以抑制它們LOXL2的產生。薑黃素(Curcumin)可以降低Phenytoin造成的LOXL2產生量，並具有劑量依賴效應。當Curcumin濃度到達10μM時開始顯著抑制HGF中LOXL2的生成，CA9-22及OECM-1則是5μM時就可以發現顯著抑制。從這些結果中我們推測Phenytoin在口腔上皮細胞中可以經由TGF-β路徑誘導LOXL2促使EMT的產生。薑黃素可以抑制LOXL2的產生，可發展為未來治療PIGO的新策略。

關鍵字

苯妥英引起之牙齦增生 ； 離氨基氧化酶樣蛋白2 ； 乙型轉化生長因子 ； 上皮間質轉化 ； 薑黃素 ； 人類牙齦纖維母細胞 ； 口腔上皮細胞CA9-22/ OECM-1

並列摘要

Phenytoin-induced gingival overgrowth (PIGO) is a common side effect of epilepsy who was treated with Phenytoin and as the most fibrotic histologic pattern in drug-induced gingival overgrowth. PIGO will impair oral hygiene maintenance, esthetic and chewing function that further become social and dining barriers. Lysyl Oxidase-like protein 2(LOXL2) has been found to induce epithelial-mesenchymal transition (EMT) and plays important role in various organ fibrosis. In PIGO, LOXL2 is overexpressed in connective tissue and epithelium. The aim of this study was to investigate whether phenytoin could induce LOXL2 expression and its mechanism. We found phenytoin increased LOXL2 expression in human gingival fibroblast and oral epithelial cell lines (CA9-22, OECM-1) in a dose- and time-dependent manner. Phenytoin increased LOXL2 expression in HGF and oral epithelial cell lines could be inhibited by pretreatment with neutralized TGF-β antibody, SIS3 and SB431542. Phenytoin can induce expression of EMT markers such as vimentin, slug and twist and suppress E-cadherin expression in OECM-1/CA9-22. These effects can be inhibited by LOX inhibitor BAPN indicated the enzyme activity of LOXL2 is required for phenytoin-induced EMT. These results indicated that phenytoin induced EMT through TGF-β/LOXL2 pathway. Furthermore, Curcumin decreased phenytoin induced LOXL2 production in a dose dependent manner in HGF and oral epithelial cell lines (CA9-22, OECM-1). At concentration of 5μM and 10μM, curcumin significantly inhibited LOXL2 expression in oral epithelial cells (CA9-22, OECM-1) and HGF, respectively. Curcumin could be a potential medication for prevention and treatment of PIGO by inhibit LOXL2.

並列關鍵字

Phenytoin-induced gingival overgrowth ； LOXL2 ； TGF-β ； EMT ； curcumin ； HGF ； oral epithelial cell(CA9-22, OECM-1)

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