後期糖化終產物經由TGF-β訊息傳遞路徑誘導人類纖維母細胞結締組織生長因子生成

Abstract

糖尿病病人通常合併多種器官纖維化的嚴重併發症, 例如: 腎間質纖維化, 心臟肌肉組織纖維化、肺纖維化等。口腔也不例外。糖尿病患者的牙齦組織無論是否有牙周病都亦有較緻密之膠原蛋白累積。牙齦纖維化對於病患的口腔衛生、美觀、咀嚼會造成影響，經由手術方式雖然可以獲得症狀緩解，但是再發生率卻很高。在牙齦纖維化的組織中常可偵測到結締組織生長因子(connective tissue growth factor, CTGF/CCN2)的上升，且其表現量與牙齦纖維化的程度呈正相關。但糖尿病導致的牙齦纖維化其相關機制目前並不清楚。 長期高血糖會增加糖化終產物(Advanced Glycation End Products, 簡稱為AGEs)的產生。AGEs被報告和腎、肺、心肌纖維化的發生有密切相關性。我們發現人類牙齦纖維母細胞(Human gingival fibroblast，HGF)在AGE的刺激下，可隨濃度及時間誘導CCN2表現。使用 TGF-β1 中和抗體、ALK5 抑制劑SB431542、Smad3 抑制劑SIS3可以有效降低AGE誘導的 CCN2蛋白表現。ELISA分析發現AGE可使細胞培養液中活化態的 TGF-β1表現量增加。顯示AGE的成纖維化特性是經由 TGF-β1 訊息路徑調控。此外兒茶素(epigallocatechin gallate, EGCG) 可明顯抑制HGF中由AGEs所誘導釋出活化態的 TGF-β1 及CCN2蛋白表現量，且抑制效果具有濃度依賴性。EGCG可能可以成為預防或治療糖尿病患者牙齦纖維化的潛力藥物。

Fibrosis of severe organs (such as renal glomeruli sclerosis, myocardial fibrosis, or lung fibrosis) is one of the most common complications in patients suffering from diabetes mellitus (DM). Moreover, dense collagen fibers was found in gingival tissue of DM patients with or without periodontal disease. Gingival fibrosis may impede oral hygiene maintenance, and affect the aesthetic and mastication function of the patient. Although surgical excision may alleviate the hyperplastic situation temporarily, the recurrence rate of gingival hyperplasia is so high to be solved with one surgery. Studies have already showed increased amount of connective tissue growth factor (CTGF/CCN2) in the fibrotic gingival tissue; and the amount of CTGF was also found to be positively related to the severity of gingival fibrosis. Yet, the exact mechanism of DM-induced gingival fibrosis was not clear. Due to the persisting hyperglycemic status, DM patients have more advanced glycation end product (AGEs) in their body. AGEs was reported to be closely related to fibrosis of kidney, lung and cardiac muscle. In this study, we found out that AGEs could stimulate the amount of connective tissue growth factor (CTGF/CCN2) protein expression in human gingival fibroblast (HGF), in a time- and dose-dependent manner. The expression of AGE-induced CCN2 was significantly suppressed by pretreating HGF with SIS3 (Smad3 inhibitor), SB431542 (ALK5 inhibitor) and TGF-β neutralizing antibody. Furthermore, by using TGF-β ELISA for quantification, we found that the expression of active form TGF-β was increased after adding AGE-BSA to HGF. This implied that AGE-induced CCN2 expression in HGF is controlled by TGF-β/smad signaling pathway. Lastly, we also discovered that epigallocatechin gallate (EGCG) could inhibit AGE-induced active form TGF-β and CCN2 protein in a dose dependent manner. Thus, EGCG may be able to prevent or treat gingival fibrosis in DM patient.