2012-01-012024-05-17https://scholars.lib.ntu.edu.tw/handle/123456789/684559摘要：為更經濟及有效地探討人類疾病治療之動物模式，試驗動物的選擇扮演臨床應用參考之重要角色，學術界一般認定豬是除靈長類外，較適用於人類醫學研究之模式動物，因此試驗用豬隻之生產有其生物醫學產業化應用之潛力。賓朗豬之種原係由畜試花斑豬(Lanyu 100)於2010年近親成功選育之白膚及白毛色個體，因此產製表現綠色螢光蛋白質之賓朗豬做為試驗用豬隻，其體型小且種原具本土性及獨特性，有利於產業化過程之國際競爭力、學術性文章發表之可被接受性及後續提出專利申請之新穎性及進步性。 間葉幹細胞可分離自骨膜、骨小樑、脂肪組織、關節液、骨骼肌及脫落之牙齒等，其在特定培養環境下具高增殖能力及可分化為多種細胞系特性。由許多臨床前及臨床之研究試驗中已證實間葉幹細胞配合組織工程之生物可降解性材料，在再生醫學及具有其應用之潛力及治療功效。此外，攜報導基因的骨髓間葉幹細胞經移植後之表現穩定性將有助於確認其於體內之功能及分化潛能。分離自綠色螢光豬之骨髓間葉幹細胞具有相同報導基因之穩定表現型式，此特性有助於追蹤其移植及分化後之命運。因此，本年度計畫將延續第一年預期完成之試驗結果包括：綠色螢光賓朗豬及骨質疏鬆賓朗豬，進行表現綠色螢光蛋白質骨髓間葉幹細胞之骨質疏鬆症治療豬之模式研究，及成纖維母細胞初代分離培養，作為種原保存及體細胞核移植之供核細胞，研發將配合已建立之基因顯微注射技術，應用本土育成之小型、白毛色及白皮膚之賓朗豬進行綠色螢光基因之基因轉殖，以育成生醫用途之基因轉殖新品系小型綠色螢光豬；本第二年計畫將建立骨質疏鬆症之幹細胞治療豬模式、生殖細胞種原保存及賓朗豬體細胞核移置，另進行骨髓間葉幹細胞之擴增並寄存於食品工業研究所，同時將提出中華民國及美國專利申請。 <br> Abstract: Porcine shared an underlying genetic and physiologic similarities with human are ideal appropriate model animals for tissue engineering and regenerative medicine researches. In terms of cost and efficiency considerations, candidate animals for experiments will play key roles. Small size Binlang-pig, outer surface coating with white skin and hairs, was bred from domestic Lanyu-100 in 2010. Thus, generation of EGFP (enhanced green fluorescence protein) transgenic Binlang-pig for experiment purposes possess the native and industrialized potentiality of biomedical application, prospects for academic excellence, innovations in scientific research or technological development, and impact on research field and international competitiveness, advantageous to paper acceptable and patent apply subsequently. Mesenchymal stem cells (MSCs) have been isolated from bone marrow, periosteum, trabecular bone, adipose tissue, synovium, skeletal muscle and deciduous teeth, and so on. They also can be expanded with high efficiency, and induced differentiation into multiple lineages under defined culture conditions. These cells have generated a great deal of interest because of their potential use in regenerative medicine and tissue engineering, and there are some dramatic examples, derived from both pre-clinical and clinical studies, that illustrate their therapeutic value. In addition, consistent expression of reporter gene incorporating into genome of implanted MSCs will facilitate the progress for verifying their functions and differentiation potential in vivo. The bone marrow-derived MSCs isolated from transgenic animal expressing reporter gene possessed the characteristics of identical expression pattern, and this strategy is favorable to track their fate after transplantation and differentiation. Hence, extend the first year outcome to generate transgenic Binlang-pig carrying PCX-EGFP transgene which includes EGFP cDNA under the control of a chicken beta-actin promoter and cytomegalovirus enhancer by pronuclear microinjection. The second year will establish the pig model of stem cell therapy for osteoporosis, germ-plasma preservation, somatic cell nuclear transfer, and the marrow-derived mesenchymal stem cells will be isolated from EGFP transgenic Binlang pigs, ready to keep in Bioresource Collection and Research Center, Food Industry Research and Development Institute for patent apply of Taiwan, R. O. C. and USA.基因轉殖豬骨髓間葉幹細胞綠色螢光蛋白質賓朗豬體細胞核移置Transgenic pigMesenchymal stem cellsEnhanced Green fluorescence proteinBinlang pigSomatic cell nuclear transfer