纖維素原膠支架混合富血小板纖維素進行豬齒芽細胞之組織工程再生

Abstract

牙齒有咬碎及研磨食物的主要功能，更有助人類發音與支撐臉部輪廓的作用。但牙齒會因牙周疾病、齲齒、撞傷或癌症而造成缺損，如果能利用再生醫學所產生組織工程的牙齒以取代缺損的牙齒將是臨床治療的一大進步。本研究之目的乃是利用組織工程的方法達成整顆牙齒的再生。牙齒發育主要是有許多形態原與生長因子參與上皮及間葉組織交互作用的模式形成。新型態的血小板聚合物-富血小板纖維素(Platelet-rich fibrin；PRF)能夠提供生長因子 (Growth factor) 和形態原 (Morphogens)，包括了Platelet-derived growth factor（PDGF）、Bone morphogenetic protein (BMP)、Epidermal growth factor（EGF）、Transforming growth factor-beta (TGF-β)、及Vascular endothelial growth factor (VEGF)等，這些生長因子及形態原和齒胚胎發育及生長之所需部份是相同的，故本實驗試著將PRF導入生牙作用以期可協同牙胚細胞和支架促進牙齒的再生。分離1.5月齡迷你豬的未萌發牙胚組織，細胞培養後使細胞量

Loss of teeth has many causes including physical trauma, gum disease and tooth decay. A biological tooth substitute that could replace lost teeth would provide a vital alternative to currently available clinical treatments. The purpose of this project is to generate a complete tooth crown by tissue engineering. Platelet-rich fibrin(PRP) is a new type of platelet concentrate which can apply a lot of growth factors and morphogens in osteogenesis, including platelet-derived growth factor（PDGF）、bone morphogenetic protein (BMP)、epidermal growth factor（EGF）、transforming growth factor-beta (TGF-β)、and vascular endothelial growth factor (VEGF). Partial of those growth factors and morphogens which involve in tooth development are match with odontogenesis. The aim of this project is to combined the dental bud cells (DBC)、fibrinogen gel and PRF to regenerate a complete tooth. The dental bud cells of molar tooth were removed from 1.5 month-old miniature pigs before eruption. When the DBC yield reached 1×106 cell/ml, the DBC were isolated, re-suspended, and then mixed the cell with fibrinogen gel and PRF. The DBC/fibrinogen gel/PRF constructs were implanted in xenograft into under dermis of nude mice’s pelvic dorsum. After xenogreat implantation for 16 weeks, there were evaluated by macrography and microsections stained with hematoxylin and eosin. We could see mineralized nodule, dentinogenic-like cells and vascular. In the swine experimental groups, one of the 9 pigs generated root, pulp, enamel, ameloblast, dentin, odontoblast, dentin tubular, cementum, blood vessels, periodontal ligament, and the other one pig erupted the regeneration tooth, and generated whole crown, root, pulp, enamel, dentin, odontoblast, dentin tubular, cementum, blood vessels, periodontal ligament. The immunohistochemical analysis results showed ameloblast expressed CK14, the dentin, dentin tubular and odontoblast expressed DMP-1, cementum and alveolar bone expressed OPN and the vascular endothelial cell of pulp expressed VEGF, too. We demonstrated that the porcine dental bud cells combined with fibrinogen gel and PRF which can grow regenerated tooth by tissue engineering when porcine model with autogenic cell transplantation.