- 學位論文
透過與真皮乳頭細胞共培養促進角質形成細胞之誘導毛髮再生能力並形成具3D結構的毛囊微組織應用於毛囊重建之研究
Promoting the hair-inductive activity of keratinocytes by co-culturing with dermal papilla cells and form 3D dermal papilla microtissues for hair follicle reconstruction
摘要
禿髮在現今社會普遍的問題，影響到越來越多年齡層的男女，因為直接影響外觀，對於患者的心理負擔更是嚴重。目前禿髮的標準治療使用像是藥物治療、光治療或植髮等，然而這些治療的成本都較高，但是卻有治療效果不穩定性及藥物副作用等可能的問題。近期隨著組織工程的發展與趨勢有越來越多學者、團隊使用組織工程的方式來進行毛囊再生之研究。毛囊再生研究與屬於特化間葉組織的真皮乳頭細胞息息相關，但真皮乳頭細胞會隨著繼代培養次數上升而降低其誘導毛髮生長的能力。毛髮發育是透過表皮—間葉之間的相互作用產生，若能使毛囊細胞與表皮細胞共同培養聚集形成立體球狀構造，以促進彼此的交互作用，使毛囊保持其毛髮誘導的能力。 因此本研究的目的在於提供一種簡易的方法用來製備及培養真皮乳頭細胞和表皮細胞形成的微球組織，有可以達到高產量、大小均一的特點，且觀察其是否能在生物體內的環境下達到毛髮新生的效果。本研究選用聚二甲基矽氧烷（Polydimethylsiloxane, PDMS）作為材料，由於其本身良好的生物相容性及細胞不易貼附的特性，將真皮乳頭細胞培養在已塗覆PDMS的96孔盤內便能使其自行聚集形成微球組織，再加上表皮細胞在外層包覆並透過免疫螢光染色確保其仍保有促進毛囊新生能力。為了提供適當的生長骨架，以膠原蛋白（collagen type I）與細胞微球組織混合使膠原蛋白包覆著微球成膠，並植入裸鼠體內觀察其發育形態，以證明本實驗設計之真皮乳頭細胞微球能促進毛囊的生成及毛髮再生，並在未來有應用於毛囊重建之組織工程的潛力。
並列摘要
Alopecia is a disturbing problem effecting men and women of all ages. Because the syndrome directly affects the appearance, cause physiological impacts on patient. The treatment for alopecia involves either hair transplant, pharmacotherapy or phototherapy. But these treatment options are fraught with problems of cost, side effects, and treatment effect is unstable. Recently, with the development and trend of tissue engineering, more and more scholars study on hair follicle regeneration. Cell-based therapies have focused on the dermal papilla cell. However, many obstacles exist, inducing ability and maintenance of dermal papilla productivity after several passages of culture. The natural hair development process depends on a series of interactions between dermal papilla cells and epidermal cells. Thus, the DP cells and epidermal cells are cultured together to form a three-dimensional spheroid structure for promote cells interaction and retain the ability to induce hair regeneration. Therefore, the aim of this study was to culture dermal papilla cells and epidermal cells to form microsphere tissues, which are high-throughput and can be produced in uniform-size manner, and to investigate whether these microsphere tissues could induce hair regeneration. Polydimethylsiloxane (PDMS) was selected as the material. Due to its good biocompatibility and low cell adhesion, dermal papilla cells were cultured in 96-well discs coated with PDMS to form micro spheroids. Moreover, epidermal cells were added to form “core-shell” structure, and immunofluorescence staining was used to ensure that they still retained the ability to promote hair follicle regeneration. On the other hand, in order to provide an appropriate scaffold, we choose collagen type I as biomaterial scaffolds. Microspheres were collected and mixed with collagen solution to form a hydrogel and transplanted into nude mice, then observe their morphology to prove that this experimental design can induce hair regeneration and has potential application in hair follicle reconstruction in the future.