利用生醫材料促進牙髓幹細胞神經分化

Neural Differentiation of DPSCs Promoted with Biomaterials

許芝維(Chih-Wei Hsu)

指導教授 ： 楊台鴻

共同指導教授 ： 陳敏慧(Min-Huey Chen)

國立臺灣大學/工學院/醫學工程學研究所/碩士(2014年)

https://doi.org/10.6342/NTU.2014.00581

摘要

本研究利用生醫材料去促進牙髓幹細胞的神經分化，牙髓幹細胞是種多潛性幹細胞可以分化成硬骨、軟骨、脂肪及神經細胞。由於其來源容易取得，擁有的倫理爭議、個體排斥問題較小等優點，再搭配生醫材料使其達到促進分化效果。
在牙髓幹細胞神經分化的文獻整理中可以得知在模擬神經幹細胞培養的環境下其幹性以及神經分化結果較一般環境以及加藥誘導來的更好，而且也有文獻指出當代表牙髓幹細胞的標記如STRO-1表現量越高對其分化的效率更佳。因此本實驗區分成兩部分，第一部分為將牙髓幹細胞培養在可貼附以及不可貼附的高分子材料上去評估在不同天期下牙髓幹細胞中幹細胞的比例改變並找出表現量最高的材料以及培養時間。第二部分為利用在的一部分所找出幹細胞比例最高的組別並培養至聚離胺酸以及由實驗室開發的類聚離胺酸材料後以定性及定量的方法去分析其神經相關的表現。
在第一部分實驗發現經過PVA懸浮後一天牙髓幹細胞STRO-1以及CD146的表現量最高，而在第二部分發現經過懸浮後再種回TCPS、PDL及N4時使用免疫染色其神經標記如Nestin、s-III tubulin及NFM都有表現其中以N4這個材料的表現量較佳。
在電生理應用測試方面可以發現經過PVA懸浮一天後再種回TCPS、PDL及N4這三種材料上培養十天後其大部分的表現量相較於TCPS的組別來的較高而且在PVA+Induction這個系統幾乎每一顆細胞都有表現。最後在做Patch Clamp測試則發現不論是在TCPS、PDL或N4這三種材料的細胞皆沒有Na離子通。

關鍵字

牙髓幹細胞 ； 生醫材料 ； 神經分化誘導 ； STRO-1 ； CD146

並列摘要

In this study, we use biomaterials to promote the differentiation of DPSCs into neural. DPSCs is a kind of multi-potent stem cells which can differentiate into bone , cartilage, fat and neural cells. Because it is easy to access the source without many ethic problems or rejection risks. However, in this assay we will use biomaterials to promotes neural differentiation in DPSCs.
Literature review of DPSCs on neural differentiation aspect we can be learned that to simulate the Neural Stem Cells culture environment DPSCs’ stemness and neural differentiation ability will be better than the normal and general induction environment. Therefore, there is another literature shows that when the STRO-1 marker( representatives as DPSCs stem cell marker) has higher expression that means it will have better differentiation ability. Therefore, in this study we will divided it into two parts, the first part is to culture DPSCs in polymer materials which can be attached or not and at a specific culture time to assess the highest proportion of stem cells. Based on part one, second part is to culture the high proportion of DPSCs into poly-D-lysine and poly-D-lysine like material which developed by the laboratory to qualitative and quantitative analysis the nerve -related performance.
In the part one experiment we found that when DPSCs during one day floated on PVA the expression of STRO-1 and CD146 was the highest. At the second part DPSCs reseeded on TCPS, PDL and N4 and from the immunofluorescence result we can found the neural marker such as Nestin, s-III tubulin and NFM has a performance on almost every materials and N4 was the best.
The neural function test of PVA one day on part one can be better than on TCPS. Moreover, when add induction medium on the PVA system almost every cell has excellent performance. Finally doing Patch Clamp found that there are no Sodium ion channel on all of the materials.

並列關鍵字

DPSCs ； Biomaterials ； Neural induction ； STRO-1 ； CD146

參考文獻

[2] Kim BC, Bae H, Kwon IK, Lee EJ, Park JH, Khademhosseini A, et al. Osteoblastic/cementoblastic and neural differentiation of dental stem cells and their applications to tissue engineering and regenerative medicine. Tissue engineering Part B, Reviews. 2012;18:235-44.

[3] Gronthos S, Brahim J, Li W, Fisher LW, Cherman N, Boyde A, et al. Stem cell properties of human dental pulp stem cells. Journal of dental research. 2002;81:531-5.

[4] Kiraly M, Porcsalmy B, Pataki A, Kadar K, Jelitai M, Molnar B, et al. Simultaneous PKC and cAMP activation induces differentiation of human dental pulp stem cells into functionally active neurons. Neurochemistry international. 2009;55:323-32.

[5] Xiao L, Tsutsui T. Characterization of human dental pulp cells-derived spheroids in serum-free medium: stem cells in the core. Journal of cellular biochemistry. 2013;114:2624-36.

[6] Barry FP, Murphy JM. Mesenchymal stem cells: clinical applications and biological characterization. The international journal of biochemistry & cell biology. 2004;36:568-84.