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Development of zinc partial stabilized cement carrying growth factor and anti-inflammatory drug for vital pulp therapy
zinc partial stabilized cement,vital pulp therapy,anti-inflammatory,pulp regeneration,animal study,
|Publication Year :||2019|
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。
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.
|Appears in Collections:||臨床牙醫學研究所|
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