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The Investigation of Porous PLGA Scaffolds Controlled Release System for Inducing Mouse Embryonic Stem Cells to Vascular Endothelial Cells
|關鍵字:||多孔PLGA釋放控制支架;Control release scaffold of porous PLGA;小鼠胚胎幹細胞;血管內皮細胞;條件培養液;控制釋放;Mouse embryonic stem cell;Endothelial cell;Conditioned medium;Control release||出版社:||生醫工程研究所||引用:|| A. Callegari, S. Bollini, L. Iop, A. Chiavegato, G. Torregrossa, M. Pozzobon, G. Gerosa, P. De Coppi, N. Elvassore, and S. Sartore, “Neovascularization induced by porous collagen scaffold implanted on intact and cryoinjured rat hearts,” Biomaterials, vol. 28, no. 36, pp. 5449-5461, 2007.  C. Wu, Y. Zhang, Y. Zhou, W. Fan, and Y. Xiao, “A comparative study of mesoporous glass/silk and non-mesoporous glass/silk scaffolds: Physiochemistry and in vivo osteogenesis,” Acta Biomaterialia, vol. 7, no. 5, pp. 2229-2236, 2011.  X. Liu, M. N. Rahaman, and Q. Fu, “Bone regeneration in strong porous bioactive glass scaffolds with an oriented microstructure implanted in rat calvarial defects,” Acta Biomaterialia, vol. 9, no. 1, pp. 4889-4898, 2013.  李宜書, “淺談組織工程,” 物理雙月刊, vol. 24, no. 3, pp. 430-435, 2001.  L. M. 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本研究利用以生物相容性材料PLGA與丙酮形成4種不同重量百分比(w/w) 1：3、1：4、1：5、1：6之溶液比例，加上2種之鹽顆粒之粒徑分別為13.6±2.8 μm與404.4±20.6 μm並以鹽析法製備出8種含水率相異之多孔PLGA支架，再於支架中包覆血管內皮細胞培養液以誘導小鼠胚胎幹細胞分化。因不同支架對水的通透性不同進而影響控制釋放速率。其中1：3 Porous I與1：6 Porous II PLGA支架，分別是含水率最小與最大之支架結構，含水率差異高達213%。1：3 Porous I支架對包含於內皮細胞條件培養液中之生長因子VEGF之控制釋放速度為1：6 Porous II支架之2倍。西方墨點法與ELISA之分析結果亦驗證小鼠胚胎幹細胞開始分化成血管內皮細胞之時間點亦與控制釋放速率之差異相符。當小鼠胚胎幹細胞開始分化時，支架之VEGF濃度僅是0.06±0.01-0.14±0.03 ng/mL，相較於現有文獻之20-50 ng/mL，本研究之方法可減少大量之VEGF之使用。且不須隨時間之增加再添加生長因子，而誘導時間亦可縮短1/2，再藉由不同支架之設計可精確控制幹細胞分化之時間點。
The key issues involved in tissue engineering are how to culture specific cells on a suitable scaffold and to provide satisfactory growth factors to regulate the differentiation and proliferation of the cultured cells. Scaffolds function as the base for cell adhesion and migration, the place for the exchange of nutrients, and to deliver and retain cells and biochemical factors. Embryonic stem cell possesses the characteristics of cellular differentiation and self-renewing, being able to differentiate to various tissue. A specifically designed scaffold can enhance the division, proliferation, and differentiation of embryonic stem cell.
In this study, first four different mixing solutions of biocompatible material PLGA and acetone with weight percentage (w/w) ratio of 1:3, 1:4, 1:5, 1:6 were prepared. The prepared solutions were than combined with salt particles of two different particle sizes (13.6�2.8 m (Porous I) and 404.4�20.6 m (Porous II)), respectively, to form eight different types of porous scaffold with various moisture contents. The porous scaffolds were then filled with vascular endothelial cell conditioned medium (ECCM) as control release scaffolds to induce the differentiation of mouse embryonic stem cell to endothelial cell. It was measured that the (1:3)/(Porous I) and (1:6)/(Porous II) scaffolds are the minimum and the maximum moisture containing scaffolds, respectively. The moisture content difference between these two types of scaffold was measured to be 213%. The control release rate of the VEGF contained in the ECCM that was embedded in the (1:3)/(Porous I) scaffolds was measured to be twice of that of the (1:6)/(Porous II) scaffolds. The starting differentiation time points of the mouse embryonic stem cell to endothelial cell in these two types of scaffold observed through Western blot and ELISA analysis are consistent with the VEGF releasing results. The VEGF concentration in the proposed control release scaffolds was measured to be from 0.06�0.01 to 0.14�0.03 ng/mL. Our control release scaffolds consumed much less amount of VEGF when compared to the reported studied that used 20 to 50 ng/mL VEFG daily to induce the differentiation of mouse embryonic stem cell. Furthermore, the starting differentiation time point of mouse embryonic stem cell in the proposed control release scaffolds can be reduced to 1/2 of that of the conventional approaches.
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