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題名 | Canine Islet Isolation, Cryopreservation, and Transplantation to Nude Mice=狗胰島細胞之分離、冷凍儲存及移植至祼鼠 |
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作者 | 黃瑞助; 盧文聰; 許瑞旭; 郭健葒; 傅馨慧; 陳漢明; 姚南光; 莊峻鍠; Huang, Jui-chu; Lu, Wen-tsoung; Hsu, Brend Ray-sea; Kuo, Chien-hung; Fu, Shin-huei; Chen, Han-ming; Yao, Nan-kuang; Juang, Jyuhn-huarng; |
期刊 | 長庚醫學 |
出版日期 | 20031000 |
卷期 | 26:10 2003.10[民92.10] |
頁次 | 頁722-728 |
分類號 | 416.248 |
語文 | eng |
關鍵詞 | 狗胰島細胞; 胰島分離; 冷凍儲存; 胰島移植; 糖尿病; Diabetes; Canine islet; Islet isolation; Islet cryopreservation; Islet transplantation; |
中文摘要 | 背景:人體胰島細胞移植已可使第一型糖尿病患者不需依賴胰島素。狗與賽體胰島細胞的特性與分離方法類似,因此狗是人體試驗前最佳的研究模型。在此我們報告狗胰島細胞分離、冷凍儲存及移植的經驗。 方法:我們摘取27隻體重介於9~13公斤雜種狗的胰臟,以collagenase消化後,再用比重分層來純化胰島細胞。我們計算胰島細胞數目及純度,細胞活性在離體使用葡萄糖靜態培養及周流來評估其胰島素分泌,在體內則將其移植至糖尿病及非糖尿病裸鼠作研究。此外,我們將剛分離的狗胰島細胞冷凍儲存一週後進行離體研究。 結果:我們得到的胰島細胞數目及純度分別是每個胰臟121,000±26,000IEQ及81.4±1.2%。其在300 mg/dl比100 mg/dl葡萄糖所釋放腸島素量之刺激指數為6.6±1.9(N=7)。此外,這些細胞在周流研究顯現第一相和第二相的胰島素分泌。解凍後胰島細胞數目由1,000降至540(N=1),胰島素含量由50.95減為39.23μg/150個胰島細胞(N=1),但其在300比100mg/dl葡萄糖靜態培養明顯表現胰島素反應(6.97比3.98μg/30胰島給胞/2小時,p<0.05)。移植四週後,移本體可見豐富的β細胞,且含胰島素。植入2,000個胰島細胞至糖尿病裸鼠,則23隻中有14隻的血糖降至正常。 結論:我們實驗室分離的狗胰島細胞具活性,且在離體及動物體內保有其生理功能。 |
英文摘要 | Backgorund: Successful human islet transplantation has led to insulin independence in type 1 diabetes. Dogs constitute an animal model for preclinical studies. We present our recent experience in canine islet isolation, cryopreservation and transplantation. Methods: Twenty-seven pancreases from mongrel dogs, weighting 9-13 kg, were removed. Each pancreas was digested with collagenase, and then purified by density gradients. Islet number and purity were counted, and the viability of isolated islets was assessed in vitro by static incubation, perfusion study and in vivo transplantation into nondiabetic or diabetic nude mice. Additionally, freshly isolated islets were cryopreserved for 1 week, and then studied in vitro. Results: The islet yield and purity were 121,000±135,000 IEQ per pancrease and 81.4±1.2%, respectively. The stimulation index (insulin release in 300 mg/dl flucose/insulin release in 100 mg/dl glucose) of the isolated islets was 6.6±1.9(N=7), and first and second phases of insulin secretion were demonstrated during perifusion study. After 1-week cryoperservation, the islet number decreased from 1,000 to 540 (N=1) and insulin content decreased from 50.95 to 39.23 μg/150 islets (N=1). These islets maintained their insulin response to high glucose. Four weeks after transplantation, the grafts showed abundant β-cells and significant insulin content. Normoglycemia was achieved in 14 of 23 diabetic recipients after transplantation with 2,000 freshly isolated islets. Conclusion: Canine islets isolated at our laboratory were viable and maintained their physiological function both in vitro and in vivo. |
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