人類乳牙牙髓幹細胞於牙髓組織再生之應用

Dental pulp tissue regeneration by deciduous pulp stem cells

張懿丹(Yi-Tan Chang)

指導教授 ： 李伯訓

國立臺灣大學/醫學院/臨床牙醫學研究所/碩士(2008年)

https://doi.org/10.6342/NTU.2008.10133

摘要

牙齒常因牙周疾病、齲齒、撞傷或癌症而造成缺損，如果能發展出生物性的再生牙齒以取代缺損的牙齒將是臨床治療的一種新療法。本研究之目的是應用乳牙牙髓幹細胞，結合組織工程的方法達成牙髓組織的再生。首先，我們實驗利用酵素分解的方法，可以從人類脫落的乳牙分離出牙髓幹細胞，乳牙牙髓幹細胞能夠形成細胞群落(colony)，這顯示這些細胞有很好的增生及自我更新的能力，同時這些乳牙牙髓幹細胞會表現三種與幹性相關的轉錄因子: Oct4、Sox2和Nanog，以及多種幹細胞的表面標誌: STRO-1和CD146、CD29、CD44、CD90及CD105。在體外培養誘導下，乳牙牙髓幹細胞可以分化成骨母細胞、脂肪細胞或牙本質母細胞。在動物實驗中，我們將迷你豬乳牙的前牙作拔髓，後牙做斷髓處理，各分成四組做實驗，第一組不放任何細胞、第二組加入迷你豬乳牙牙髓幹細胞和培養液、第三組加入迷你豬乳牙牙髓幹細胞和lovastatin，第四組加入迷你豬乳牙牙髓幹細胞和odontoblast induction medium，觀察三個月後，以探討其牙齒組織是否會再生。結果發現會有再生性牙本質及collagen的產生。在後牙的組別，形成的再生性牙本質的量在各組並無差異；在前牙的組別，第三組加入lovastatin的牙齒，根管內產生了7.5 mm再生性牙本質，比第一組(4.0 mm)及第四組多(5.5 mm)，所以lovastatin可能會促進牙本質母細胞的分化。總括來說，本研究建立了一個新的研究牙髓幹細胞應用於牙齒再生治療的模式，我們發現從乳牙中可以分離出牙髓幹細胞，未來並可進一步應用來修復牙齒的缺損。

關鍵字

牙髓幹細胞 ； 乳牙 ； 間質幹細胞 ； 組織工程 ； 牙齒再生 ； 牙本質

並列摘要

Periodontal diseases、caries、trauma or cancer are common causes for tooth decay or tooth loss. A biological tooth substitute that could replace decayed tooth or lost tooth would provide a vital alternative to currently available clinical therapy. The aim of the study was to utilize deciduous dental pulp stem cells and tissue engineering method to regenerate injured dental pulp. By using enzyme digestion, dental pulp stem cells could be isolated from deciduous teeth. These cells have capacity to generate clongenic cell clusters in culture. They were also identified to be a population with high proliferation capability. They can express three stemness genes- Oct4, Sox2 and Nanog and surface markers which are known to express in stem cells, such as STRO-1, CD146, CD29, CD44, CD90 and CD105 were also expressed on these cells. Moreover, these deciduous dental pulp stem cells could differentiate into osteoblasts, adipocytes, or odontoblast. We also established an animal model with utilizing mini-pig to determine the functional property of dental pulp stem cell in vivo. Firstly, we performed pulpotomy for posterior teeth and pulpectomy for anterior teeth. And the experiments were divided into four groups. In the first group, nothing was put in the pulp chamber or canals. In the second group, deciduous dental pulp stem cells of mini-pig with cultured medium were transplanted into the pulp chamber and pulp canals. In the third group, deciduous dental pulp stem cells of mini-pig with lovastatin were transplanted. In the fourth group, deciduous dental pulp stem cells of mini-pig with odontoblast induction medium were transplanted. Then we determined the pulp reaction three month after transplantation. The result showed that reparative dentin and collagen were regenerated. The amount of regenerative dentin in each group of posterior teeth was roughly similar. However, in the third group of anterior teeth, in which lovastatin was added, 7.5 mm reparative dentin was regenerated. The regenerated amount was greater than those in the first and the fourth groups. Therefore, lovastatin might enhance odontoblast differentiation. In summary, we established a new model for investigating the functional reconstruction capability with dental pulp stem cells transplantation in vivo. We also revealed dental pulp stem cells from one deciduous tooth can be expanded in vitro which possibly can be utilized for teeth replacement therapy in the future.

並列關鍵字

dental pulp stem cell ； deciduous tooth ； mesenchymal stem cell ； tissue engineering ； tooth regeneration ； dentin

參考文獻

Aberg, T., Wozney, J., et al. (1997). "Expression patterns of bone morphogenetic proteins (Bmps) in the developing mouse tooth suggest roles in morphogenesis and cell differentiation." Dev Dyn 210(4): 383-96.

About, I., Bottero, M. J., et al. (2000). "Human dentin production in vitro." Exp Cell Res 258(1): 33-41.

Ahdjoudj, S., Lasmoles, F., et al. (2001). "Reciprocal control of osteoblast/chondroblast and osteoblast/adipocyte differentiation of multipotential clonal human marrow stromal F/STRO-1(+) cells." J Cell Biochem 81(1): 23-38.

Al-Talabani, N. G. and Smith, C. J. (1979). "A histological study of the effect of developmental stage of tooth-germ on transplantation to hamster cheek pouch." Arch Oral Biol 24(12): 933-7.

Banchs, F. and Trope, M. (2004). "Revascularization of immature permanent teeth with apical periodontitis: new treatment protocol?" J Endod 30(4): 196-200.