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    題名: 低氧狀態對於無血清培養 CD73+ 人類牙髓幹細胞的影響
    Effect of Hypoxia on CD73-Positive Human Dental Pulp Stem Cells in a Serum-free Culture System
    作者: 何福源
    Ho, Fu-Yuan
    貢獻者: 牙醫學系碩博士班
    李勝揚
    黃彥華
    關鍵詞: 牙髓幹細胞;低氧狀態;無血清培養基;增生;擴大
    Dental pulp stem cells;hypoxia;serum-free media;proliferation;expansion
    日期: 2013-01-23
    上傳時間: 2018-10-02 10:01:41 (UTC+8)
    摘要: 中文摘要
    從人類恆牙牙髓中取得的人類牙髓幹細胞 (Human dental pulp stem cells) 已被相信是一個很可靠且有潛力的前驅細胞 (Progenitor cells) 來源。若是能以無血清 (Serum-free) 的培養系統來培養及擴大這些前驅細胞，將會對於自體移植的再生醫學及臨床應用上會有很大的幫助。低氧氣環境 (Hypoxia) 已證實能促進幹細胞維持於未分化形態。然而，對於在低氧氧及無血清的狀態下培養人類牙髓幹細胞所造成的影響，尚未清楚。本研究中，從成人的齒顎矯正病患的牙齒中收集健康的牙髓，從這些牙髓組織培養出牙髓幹細胞，並以磁珠篩選系統 (Magnetic activated cell sorting system) 分離 CD73+ 及純化的人類牙髓幹細胞。藉由無血清的培養基來將這些牙髓幹細胞分別培養在正常氧氣狀態 (21% 氧氣濃度) 及低氧狀態 (5% 氧氣濃度) ，並且分析分析其細胞增生 (Proliferation assay)。研究結果證實低氧狀態下的 CD73+ DPSC 具有較高的細胞增生能力。此外，將低氧氣狀態下培養的 CD73+ DPSC 和正常氧氣狀態下培養的 CD73+ DPSC 比較，具有較佳的細胞型態 (Cell morphology) ，並且對於幹細胞特異的表面抗原 (Stem cell-specific markers) 例如 CD73 及 CD29 有較明顯的表現，及其分化的能力 (Differentiation capacity) 也較佳。此外，當在無血清的培養基中加入 IGF-1 生長因子 (Growth factor) 會大幅地增加人類牙髓幹細胞的幹細胞特性。這樣的結果顯示，IGF-1/IGF-1R 訊息傳遞路徑 (IGF-1/IGF-1R signal pathway) 對於維持人類牙髓幹細胞的幹細胞特性上扮演著重要的角色。總地來說，在我們的研究中指出，發現 IGF-1R 訊號及低氧氣培養，對於幹細胞的增生及幹細胞特性上扮演重要的角色。實驗結果顯示，提供一個更有效的培養環境，可以在體外培養及擴大成人恆牙牙髓幹細胞，更瞭解低氧氣狀態和 IGF-1 無血清狀態對人類牙髓幹細胞的影響，可望促進個體化的人類幹細胞療法的發展。
    Abstract
    Human dental pulp from permanent tooth is a potentially promising source of progenitor cells. Sustaining and amplifying progenitor cell populations with serum-free culture system would greatly benefit for autograft transplantation and clinical cell therapy. Hypoxia is known to promote the undifferentiated cell state in various stem cell populations. However, the effect of hypoxia combined with serum-free culture media on human dental pulp stem cells (DPSCs) has not been defined yet. In this article ,dental pulps from adult orthodontic patients were cultivated and the DPSCs were selected by magnetic activated cell sorting system (MACS) for experiments. The DPSCs were cultivated in serum-free culture medium under normoxic (21% O2) and hypoxic (5% O2) culture conditions for cell proliferation assay. A cell counting assay showed a significant increased proliferation of hypoxic-DPSCs. Additionally, the hypoxic-DPSCs showed a better cell morphology, higher expression level of stem cell–specific markers (CD73 and CD29), and differentiation capacity (adipogenesis and osterogenesis) while comparing with the normoxic-DPSCs. Importantly, supplementing IGF-1 or insulin in medium greatly increased the stem cell properties of DPSCs. Together with these results, highlight the important role of hypoxia and IGF-1R signaling in stemness maintenance of DPSCs. In summary, in this study, we found an important role of IGF-1R signaling and hypoxia in proliferation and maintaining stemness of DPSCs.
    描述: 碩士
    指導教授-李勝揚
    共同指導教授-黃彥華
    委員-鄭景暉
    委員-林泰元
    委員-黃惠美
    資料類型: thesis
    顯示於類別:[牙醫學系] 博碩士論文

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