| 摘要: | 二苯乙烯?是中藥材何首烏中的活性成分，具有降血脂、抗發炎等功效。本研究旨在探討二苯乙烯? (2,3,5,4’-tetrahydroxystilbene-2-O-beta-D-glucoside，簡稱 THSG) 刺激人類牙髓幹細胞（human dental pulp stem cells，簡稱 hDPSCs）產生外泌體在細胞活性、傷口癒合和成骨分化的影響，並觀察其於大鼠骨缺損中組織修復的效果。首先評估不同濃度 THSG 誘導的外泌體 (THSG-Exo) 與 hDPSCs 外泌體 (DMSO-Exo) 對於老鼠前類骨母細胞 (MC3T3) 與人類牙齦纖維母細胞 (HGF) 的細胞增殖影響。結果發現 MC3T3 與 HGF 分別在THSG-Exo (100 μM) 與 THSG-Exo (10 μM) 的條件下產生明顯的細胞增殖效果。傷口癒合試驗結果顯示 HGF 在 THSG-Exo 作用下，在第六天和第九天時缺口面積顯著縮小，且在第九天時缺口癒合超過百分之四十。骨分化實驗結果顯示 THSG-Exo 能讓 MC3T3 在第一週即誘發骨分化效果，並在第二週與第三週時觀察到更多鈣化結節。在大鼠上顎骨缺損的實驗模型中，THSG-Exo 讓骨缺損區域的骨容積、骨小樑厚度和骨小樑密度有顯著增加的情形。大鼠顎骨組織學切片的結果顯示 THSG-Exo 讓新生成骨區域的骨母細胞數量增加，也使得 OPN、PCNA、VEGF 的抗體表現增加。蛋白質體組成分析結果顯示，THSG-Exo 中內含多種骨分化、傷口癒合、組織修復相關蛋白質，例如 BMP-1、MMP-1 及 TSG-6。綜合以上，THSG 誘導 hDPSCs 產生的外泌體，不僅在體外實驗證實了有促使細胞增殖、促進傷口癒合與刺激成骨分化之能力，在體內實驗也證實有加速組織修復與骨再生的效果，合併蛋白質體組成分析結果證實 THSG-Exo 內含多種有利於組織修復的蛋白質。由此推論，THSG 能誘使 hDPSCs 產生更優質的外泌體，不論在質與量上皆有所提升，更有利於臨床上在傷口癒合、成骨再生、組織修復中的應用。
As an active component in Chinese herb He-Shou-Wu, 2,3,5,4'-tetrahydroxystilbene-2-O-beta-D-glucoside (THSG) is known to have antilipidemia & anti-inflammatory properties. The purpose of this study is to investigate the effects of THSG stimulated human dental pulp stem cells (hDPSCs) derived exosomes on cell viability and their applications in wound healing and osteogenic differentiation, as well as tissue repair in rat bone defects. Firstly, the study evaluated the effects of THSG-induced exosomes (THSG-Exo) and hDPSCs exosomes (DMSO-Exo) on the cell viability of mouse pre-osteoblasts (MC3T3) and human gingival fibroblasts (HGF). The results showed that 10 μM THSG-Exo led to a more significant cell proliferation effect on HGF, while 100 μM THSG-Exo had a more pronounced cell proliferation effect on MC3T3. Wound healing assay results demonstrated that under THSG-Exo treatment, HGF showed a significant reduction in wound area on the sixth and ninth days, with wound healing exceeding 40% on the ninth day. Osteogenic differentiation experiments revealed that THSG-Exo induced osteogenic effects in MC3T3 within the first week, with more calcified nodules observed in the second and third weeks. Micro-CT imaging analysis of rat bone defects showed that the group treated with THSG-Exo exhibited significant increases in bone volume, trabecular thickness, and trabecular density. Histological staining of rat maxillary bone sections revealed an increased number of osteoprogenitor cells in the newly formed bone in the THSG-Exo group, while the immunohistochemical staining results for OPN, PCNA, and VEGF showed an increased expression in the THSG-Exo group. Proteomic composition analysis indicated that THSG-Exo contained various proteins related to bone differentiation, wound healing, and tissue repair, e.g., BMP-1, MMP-1, TSG-6. In summary, THSG-induced exosomes derived from hDPSCs not only demonstrated the ability to promote cell proliferation, wound closure, and osteogenic differentiation in vitro experiments but also accelerated tissue repair and bone regeneration in vivo experiments. Combined with proteomic composition analysis results, THSG-Exo was found to contain various proteins beneficial for tissue repair. In conclusions, THSG can induce hDPSCs to produce higher quality exosomes, with improvements in both quality and quantity, making them more suitable for clinical applications in wound healing, osteogenic regeneration, and tissue repair. |
