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標題: | 毛囊之真皮乳頭細胞在乙烯-乙烯醇共聚合薄膜上自我聚集成立體微組織 Self-assembly of dermal papilla cells into three-dimensional microtissues on poly (ethylene-co-vinyl-alcohol) membrane |
作者: | Chiao-Yun Lee 李巧芸 |
指導教授: | 楊台鴻(Tai-Horng Young) |
關鍵字: | 毛囊,真皮乳頭細胞,毛囊再生,多細胞球,微組織,乙烯-乙烯醇共聚合薄膜,細胞遷移, hair follicle (HF),dermal papilla (DP) cells,HF regeneration,multicellular spheroids,microtissues,poly (ethylene-co-vinyl alcohol) (EVAL),cell migration, |
出版年 : | 2007 |
學位: | 碩士 |
摘要: | 很多疾病會造成毛囊缺失,而利用毛囊再生來治療人類毛囊缺失,一直是很多研究者的夢想。目前研究顯示,將毛囊真皮乳頭細胞適當地移植到表皮下方,可以使毛囊再生。然而,真皮乳頭細胞必須在聚集成多細胞球(亦稱作微組織)的情況下,才有誘導毛囊新生的功能。在先前的文獻中,大多是利用離心的方式使細胞聚集,但此方法效率低,無法在短時間內大量獲得微組織。也就是說,目前缺乏在體外直接將真皮乳頭細胞大量地培養成微組織的方法。因此我們嘗試找出適當之高分子基材,一方面能利於真皮乳頭細胞增生,一方面可以有效率地促進細胞之聚集並保有毛囊誘導的功能。
本研究中,我們發現乙烯-乙烯醇共聚合薄膜(EVAL)能促進真皮乳頭細胞形成立體微組織。首先,我們的研究顯示真皮乳頭細胞在EVAL薄膜上會有細胞增殖的現象。當真皮乳頭細胞在EVAL薄膜上達到一定的細胞數量並培養三天後,細胞會自行聚集形成許多結構緊密的微組織。而在微組織中真皮乳頭細胞約有96%的存活率並保有其原本細胞的特性。我們更進一步發現,真皮乳頭細胞能在EVAL薄膜上自行聚集成微組織的關鍵是局部的高細胞密度,而非整體的細胞數量或密度所影響。同時,我們也發現真皮乳頭細胞在EVAL薄膜上的貼附性較差。因此,我們推測微組織的形成,是由於真皮乳頭細胞在EVAL薄膜上貼附能力較差,導致細胞更容易遷移,進而縮短細胞之間的距離並增加細胞間互相碰撞的頻率。同樣地,當局部細胞密度愈高時,細胞碰撞的機會也會增加,微組織也更容易形成。 利用此方法大量生產的毛囊真皮乳頭微組織不僅能應用在毛囊再生,也能作為治療毛囊缺失之藥物的篩檢平台。在此,我們提出毛囊重建的三個步驟:先在體外大量增生真皮乳頭細胞、再將真皮乳頭細胞培養成微組織、最後將微組織移植到活體內誘導毛囊再生。此外,這種多細胞自行聚集成為立體微組織的模型,對於胚胎時期毛囊發育與細胞生物學的研究也有很大的幫助。 Many diseases cause gradual destruction and loss of normal hair follicles (HFs). Regeneration of HF in post-natal period can be achieved by transplanting cultured HF dermal papilla (DP) cells under the epidermis. However, the ability of DP cells to guide the transdifferentiation of epidermis into follicular structures is only preserved when they are kept in multicellular spheroids or microtissues, an intercellular organization similar to that in vivo. Up to date, there have been no efficient methods to produce DP microtissues on a large scale. In this research, we attempt to find the feasible polymer substrates that facilitate DP cell abundant expansion and self-assembly into three-dimensional multicellular spheroids. We demonstrate that DP cells spontaneously grow into microtissues on poly (ethylene-co-vinyl alcohol) (EVAL) membrane. First we show that EVAL membrane is able to support the proliferation of DP cells. Seeded above a critical cell number on EVAL membrane, DP cells spontaneously form dense microtissues after 3 day in culture. Averagely, the cell viability of each microtissue is about 96%. And the differentiation markers of DP cells are still preserved in DP microtissues. We further demonstrate that the local cell density on a smaller area, rather than the overall cell number or the overall cell density, is key to the self-assembly of DP cells into microtissues on EVAL membrane. We suggest that the declined cell attachment to EVAL membrane may facilitate cell migration and the decreased intercellular distance may increase the frequency of intercellular collision. Similarly, a higher cell density also increases the frequency of intercellular collision and there by promotes the formation of DP microtissues. With further development, this system can produce DP microtissues efficiently for clinical applications in hair follicle regeneration and also for pharmaceutical testing. Here, we propose a three-step approach for HF reconstruction: large expansion of DP cells in vitro, cultivating DP cells into microtissues and transplantation of DP micritissues in vivo. In addition, the self-assembly of DP cells into three-dimensional microtissues can be a useful model for the study of hair follicle morphogenesis and DP physiology. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/29568 |
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顯示於系所單位: | 醫學工程學研究所 |
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