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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 林俊彬 | |
dc.contributor.author | Chih-Chun Chang | en |
dc.contributor.author | 張智鈞 | zh_TW |
dc.date.accessioned | 2021-06-17T07:04:08Z | - |
dc.date.available | 2019-08-26 | |
dc.date.copyright | 2019-08-26 | |
dc.date.issued | 2019 | |
dc.date.submitted | 2019-07-29 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/72708 | - |
dc.description.abstract | 活髓治療,係指在牙髓尚未發生不可逆牙髓炎前,將剩餘健康的牙髓組織保留下來的術式,目的是維持牙髓的活性,誘導形成牙本質橋之屏障阻擋外界的感染與刺激,更進一步促進牙髓組織的修復及再生。因此若活髓治療材料能攜帶抗發炎藥物控制牙髓的發炎狀態,且同時攜帶生長因子以促進牙髓組織的修復及再生,預期將可以達到良好的治療效果。然而目前並沒有活髓材料能夠有效地攜帶抗發炎藥物及生長因子應用於活髓治療。
本研究的目的為以含鋅介穩水泥,搭配聚麩胺酸-明膠(γ-PGA/ gelatin)作為攜帶藥物及生長因子之載體,導入抗發炎藥物Lysine acetylsalicylate(LAs)及生長因子Vascular endothelial growth factor(VEGF),發展兼具抗發炎效果和促進牙髓再生能力的新型活髓治療材料。 我們將此研究分為三大部分執行,第一部分為材料研發:在含鋅介穩水泥中,導入γ-PGA水膠,藉由電子顯微鏡/ X光繞射分析、硬化時間、抗壓強度等測試篩選出最適合臨床應用之理想成份。第二部分則藉由LPS誘導人類牙髓幹細胞進行發炎模式分析,偵測細胞激素TNF-α、IL-8釋放,評估抗發炎藥物對細胞發炎模式之抗發炎效用,並將攜帶LAs及VEGF之材料進行體外生物相容性測試(包括以alamar blue檢測細胞存活率、LDH檢測細胞毒性),藥物釋放測試,牙髓幹細胞硬骨分化能力之測試(包括ALP鹼性磷酸酶和鈣沉澱定性染色)。第三部分以大鼠作為模型進行動物實驗,利用micro–computed tomography ( µ-CT)與組織切片,分析材料於活體中之抗發炎及誘導牙髓牙本質組織再生之能力。 第一部分實驗結果顯示:加入1%或2%的γ-PGA水膠,並不影響含鋅界穩水泥之結晶性。各組初始硬化時間介於9~17分鐘,符合臨床操作所需之性質。且隨著介穩水泥含鋅量的增加,及添加γ-PGA濃度上升,硬化時間有縮短的趨勢。隨著含鋅量的下降,及添加γ-PGA濃度上升,材料的抗壓強度則有上升的趨勢。綜合以上測試結果,我們選擇2γ-5%ZnPSC和2γ-7%ZnPSC作為後續實驗組別。 第二部分實驗結果顯示:在以10 μg/ml LPS誘導6小時建立的牙髓幹細胞發炎模式下,添加LAs能有效降低TNF-α、IL-8的釋放量。在alamar blue的測試中顯示2γ-5%ZnPSC組別具有較佳的生物相容性,因此選擇此組進行後續實驗。VEGF和LAs於第一天內會快速釋放,且由鹼性磷酸酶或鈣沉澱染色結果顯示,VEGF和LAs的添加可以促進牙髓幹細胞(DPSCs)之硬骨分化之能力。 第三部分進行大鼠動物實驗,經由µ-CT和組織學切片分析結果顯示,添加VEGF的組別硬組織生成速率較快,周圍血管新生較明顯,且出現連續性牙本質橋的時間較早;而添加LAs的組別初期都沒有出現發炎反應。整體而言,5%ZnPSC +VEGF+ LAs組別在硬組織生成的速率、連續性、厚度及牙髓發炎狀態,都是表現最優異的組別。 綜合以上結果,本研究所研發攜帶生長因子VEGF及抗發炎藥LAs的含鋅介穩水泥具有優異的性質,且在動物實驗中展現良好的生物活性,相當具有潛力作為活髓治療的材料。 | zh_TW |
dc.description.abstract | The goals of vital pulp therapy are mainly to treat reversible pulpal injury, maintain pulp vitality, and establish a barrier to protect pulp from the invasion of bacteria and stimulations from the environment. Inducing repair and regeneration process of the pulp is the ultimate purpose of vital pulp therapy. A material which carrying anti-inflammatory drug and growth factor is excepted to have promising treatment outcome for vital pulp therapy due to the ability to control inflammatory status and induce repair and regeneration process of the pulp. However, nowadays there is no material that can carry growth factor and anti-inflammatory drug efficiently for vital pulp therapy.
The aim of this study was to develop a zinc partial stabilized cement /γ- PGA hydrogel to carry vascular endothelial growth factor (VEGF) as growth factor and lysine acetylsalicylate (LAs) as anti-inflammatory drug for vital pulp therapy. This study was designed as three parts. In the first part, SEM/XRD analysis, setting time, compressive strength, pH value were used to investigate the physical-chemical and mechanical properties of ZnPSC/γ-PGA hydrogel. In the second part, DPSCs inflammatory model, alamar blue, LDH, UV-VIS, ALP assay, Alizarin red staining, were used to investigate the anti-inflammatory effect, biocompatibility, cytotoxicity, drug releasing pattern, mineralization-inducing potential of the material carrying VEGF and LAs. In the third part, animal experiment with rat model was performed, µ-CT and histological analysis were utilized to evaluate the anti-inflammatory efficacy and the ability of inducing pulpal repair and regeneration. Part I : The results showed that adding 1% or 2% γ-PGA hydrogel didn’t affect the crystallinity of ZnPSC. All tested groups’ initial setting time were between 9~17 minutes, which were optimal for clinical use. As the zinc and γ-PGA content increased, the setting time tended to decrease. As the zinc content decreased and the concentration of γ-PGA increased, the compressive strength of the material tended to increase. According to the results, 2γ-5%ZnPSC and 2γ-7%ZnPSC groups were selected for further experiment. Part II : The results showed that LAs reduced TNF-α, IL-8 releasing in the inflammatory model of DPSCs induced by10μg/ml LPS. 2γ-5%ZnPSC showed better cell viability than other groups in alamar blue assay, thus was chosen for further experiment. VEGF and LAs both showed burst release in drug releasing pattern. The results of ALP assay and Alizarin red staining indicated that VEGF and LAs promoted osteogenic differentiation of DPSCs. Part III : Animal study in rat model were analyzed by µ-CT and histological section. The results showed that in the groups with VEGF added, hard tissue and continuous dentin bridge formation was faster and the peripheral angiogenesis was more obvious than other groups. In the groups with LAs added, there were no inflammatory reaction in the early stage . Overall, 5% ZnPSC + VEGF + LAs group showed the best results in hard tissue formation rate, continuity and thickness of dentin bridge formation, and the inflammatory status of the pulp. According to the results of this study, the developed zinc partial stabilized cement /γ- PGA hydrogel carrying VEGF as growth factor and LAs as anti-inflammatory drug has great potential as a material for vital pulp therapy. | en |
dc.description.provenance | Made available in DSpace on 2021-06-17T07:04:08Z (GMT). No. of bitstreams: 1 ntu-108-R05422003-1.pdf: 4714547 bytes, checksum: 212272c536589efad0d868c26aeaa974 (MD5) Previous issue date: 2019 | en |
dc.description.tableofcontents | 致謝 I
中文摘要 II Abstract IV 目錄 VI 圖目錄 X 表目錄 XIII 第一章 前言 1 第二章 文獻回顧 3 2.1牙髓牙本質複合體(pulp-dentin complex) 3 2.2保存活髓(vital pulp)於臨床牙科治療之重要性 4 2.3活髓治療 5 2.4活髓保存材料於活髓治療的重要性 5 2.5氫氧化鈣 6 2.6 Mineral trioxide aggregate (MTA) 6 2.7本研究團隊研發之含鋅介穩水泥 7 2.8生長因子:血管內皮生長因子(vascular endothelial growth factor, VEGF) 8 2.9 抗發炎藥物: Lysine acetylsalicylate (LAs) 9 2.10聚麩胺酸 (Poly-γ-glutamic acid, γ-PGA) 10 2.11明 膠 (Gelatin) 10 第三章 動機與目的 12 第四章 材料與方法 14 4.1材料製備 14 4.1.1含鋅介穩水泥材料之製備 14 4.1.2電子顯微鏡觀察含鋅介穩水泥粉末型態 14 4.1.3 X光繞射評估分析含鋅介穩水泥粉末 14 4.1.4含鋅介穩水泥混合聚麩胺酸-明膠之製備 15 4.1.5 X光繞射評估含鋅介穩水泥混合聚麩胺酸-明膠之材料性質15 4.2硬化時間測試 15 4.2.1測試樣本製備 15 4.2.2測試方法 15 4.3抗壓強度測試 16 4.3.1測試樣本置備 16 4.3.2測試方法 16 4.4 酸鹼值測試 16 4.4.1測試樣本製備 16 4.4.2測試方法 16 4.5 離子釋放濃度測試 17 4.5.1感應耦合電漿質譜分析儀(ICP-MS)校正 17 4.5.2測試樣本製備 17 4.5.3測試方法 17 4.6 體外生物活性測試 17 4.6.1 測試樣本製備 17 4.6.2 測試方法 17 4.7 抗發炎能力測試 18 4.7.1 LPS誘導細胞發炎模式之建立 18 4.7.2 LPS誘導細胞發炎偵測細胞激素的釋放量 18 4.7.3 評估攜帶生長因子與抗發炎藥物之含鋅介穩水泥對細胞發炎的治療效果 20 4.8 材料生物相容性測試 22 4.8.1 人類牙髓幹細胞之初級培養(Primary culture) 22 4.8.2 萃取液製備 22 4.8.3 細胞存活率實驗(Alamar Blue cell viability assay) 23 4.8.4 細胞毒性測試(LDH release assay) 23 4.9 生長因子及抗發炎藥物釋放測試 24 4.9.1 測試樣本製備 24 4.9.2 測試方法 24 4.10生物礦化能力分析 25 4.10.1鹼性磷酸酶(Alkaline phosphatase, ALPase)定性染色分析 25 4.10.2鈣沉澱定性染色 25 4.11 動物實驗 26 4.11.1實驗動物之選擇與照顧 26 4.11.2實驗步驟 26 4.11.3動物犧牲 27 4.11.4 μ-CT 照射 27 4.11.5組織學切片標本備置 27 4.11.6 μ-CT及組織學評分標準 28 第五章 結果 29 5.1 材料粉末分析 29 5.1.1 X光繞射分析 29 5.1.2電子顯微鏡觀察 29 5.2水合產物分析 29 5.2.1 X光繞射分析 29 5.3硬化時間測試 30 5.4抗壓強度測試 30 5.5離子釋放濃度測試 30 5.6酸鹼值測試 31 5.7體外生物活性測試 31 5.8抗發炎藥物細胞發炎模式測試 32 5.8.1 LPS誘導細胞發炎偵測細胞激素的釋放量 32 5.8.2評估攜帶生長因子與抗發炎藥物之含鋅介穩水泥對細胞發炎的治療效果 32 5.9材料生物相容性測試 33 5.9.1細胞存活率實驗(Alamar Blue cell viability assay) 33 5.9.2細胞毒性測試(LDH release assay) 33 5.10生長因子及抗發炎藥物釋放測試 33 5.11 生物礦化能力分析 34 5.11.1鹼性磷酸酶(Alkaline phosphatase, ALPase)定性染色分析 34 5.11.2鈣沉澱定性染色 34 5.12 動物實驗μ-CT照射影像分析 34 5.12.1動物實驗兩週μ-CT照射影像 34 5.12.2動物實驗一個月μ-CT照射影像 36 5.12.3動物實驗兩個月μ-CT照射影像 38 5.13 動物實驗組織學切片觀察 40 5.13.1動物實驗兩週組織學切片觀察 40 5.13.2動物實驗一個月組織學切片觀察 41 5.13.3動物實驗兩個月組織學切片觀察 42 第六章 討論 45 6.1鋅含量和γ-PGA 水膠的添加對介穩水泥硬化時間的影響 45 6.2添加鋅和γ-PGA 水膠對介穩水泥抗壓強度的影響 46 6.3抗發炎藥物LAs對牙髓幹細胞發炎模式之影響 47 6.4探討加入含VEGF及LAs之γ-PGA 水膠對含鋅介穩水泥生物相容性的影響 48 6.5探討加入含VEGF及LAs之γ-PGA 水膠對含鋅介穩水泥生物礦化能力的影響 49 6.6探討加入含VEGF及LAs之γ-PGA 水膠對含鋅介穩水泥於活體之整體表現 49 第七章 結論 51 參考文獻 52 附圖 59 附表 111 | |
dc.language.iso | zh-TW | |
dc.title | 研發攜帶生長因子與抗發炎藥物之含鋅介穩水泥應用於活髓治療 | zh_TW |
dc.title | Development of zinc partial stabilized cement carrying growth factor and anti-inflammatory drug for vital pulp therapy | en |
dc.type | Thesis | |
dc.date.schoolyear | 107-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 章浩宏,林弘萍,廖運炫,王姻麟 | |
dc.subject.keyword | 含鋅介穩水泥,活髓治療,抗發炎,牙髓組織修復及再生,動物實驗, | zh_TW |
dc.subject.keyword | zinc partial stabilized cement,vital pulp therapy,anti-inflammatory,pulp regeneration,animal study, | en |
dc.relation.page | 113 | |
dc.identifier.doi | 10.6342/NTU201902093 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2019-07-29 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 臨床牙醫學研究所 | zh_TW |
顯示於系所單位: | 臨床牙醫學研究所 |
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