造纖維母細胞生長因子對造牙骨質母細胞影響之研究

造纖維母細胞生長因子(Fibroblast growth factor-2,簡稱:FGF-2)擁有 許多生物活性的功能,在組織再生過程中扮演著重要訊息傳遞分子的角色。 造牙骨質母細胞(cementoblasts)是牙周組織的支持細胞,其主要功能包括調 控牙周組織的生長發育、維持與再生。但從文獻回顧中發現鮮少有研究探 討FGF-2對造牙骨質母細胞影響,故本研究以組織培養法,培養造牙骨質母 細胞(murine immortalized cementoblast cell line,簡稱 OCCM.30),探討 FGF-2對OCCM.30細胞的作用機轉,實驗利用 alamar blue 呈色法來評估 FGF-2對細胞存活率的影響;以傷口癒合法(wound healing assay)和轉移盤爬 行法(transwell migration assay)來觀察對細胞遷移(migration)能力的影響;並 以西方墨點法來檢測磷酸化態的AKT蛋白質表現量;另測定鹼性磷酸酶 (alkaline phosphatase)的活性來觀察細胞分化現象,且以茜草素紅(alizarin red)染色法進行礦化能力的分析。實驗結果顯示:FGF-2會促進OCCM.30 細胞的生長與增殖,此一現象會隨著濃度上升而有增加的趨勢(p<0.05); FGF-2亦會促進細胞遷移的能力(p<0.05);FGF-2會促進磷酸化態的AKT的 蛋白質表現量,且會隨著時間增加而增加(p<0.05)。然而,鹼性磷酸酶的活 性與鈣化結節形成的能力,與FGF-2的存在與否,並沒有顯著的變化(p>0.05)。 綜合實驗結果顯示,FGF-2會促進造牙骨質母細胞生長、遷移與增殖,並經 由活化磷酸化態的AKT訊息傳遞路徑而產生,但對誘發造牙骨質母細胞進 行分化及硬組織的生成能力影響不大,由此推斷FGF-2可能藉由促進細胞生長與遷移至傷口,而在早期牙周組織再生、傷口癒合的階段扮演重要的角 色。

Fibroblast growth factor (FGF)-2 has multifunctional biologic activity and is known as a signaling molecule that induces tissue regeneration. Cementoblasts not only function as support cells for periodontium but also control development, maintenance, and regeneration of periodontal tissues. Little is known about the effect of FGF-2 on cementoblasts. The aim of this study was to investigate FGF-2 on murine immortalized cementoblast cell line (OCCM.30) by measuring cell growth, proliferation, and differentiation. Cell viability was judged by using alamar blue reduction assay. Cell migration was evaluated by transwell and wound-healing assays. Western blot was employed to evaluate the expression of phosphorylated Akt (p-Akt). Cytodifferentiation was examined by alkaline phosphatase (ALP) assay and cytochemical staining of calcium by using alizarin red. Our results showed that FGF-2 was found to increase cell growth and proliferation in a dose-dependent manner (p<0.05). FGF-2 was also demonstrated to enhance cell migration (p<0.05). Cellproliferation-related proteins p-Akt were significantly elevated by stimulation with FGF-2 compared with untreated controls (p<0.05). However, ALP activity and calcified nodule formation had no significant change in the presence or absence of FGF-2 (p>0.05). Taken together, FGF-2 could increase cementoblast growth, migration, and proliferation but might suppress cytodifferentiation into hard tissue-forming cells. FGF-2 may play an important role in periodontal regeneration during the early phase of wound healing.