臺灣博碩士論文加值系統

中文摘要

研究目的：糖尿病在目前臨床醫學上仍然無法根治，幹細胞能夠治療許多疾病並能分化成胰島素製造細胞。本實驗期望以人類臍帶間質幹細胞分化成類胰島團塊，移植至胰臟全切除手術動物藉由觀察其生理變化以及存活率以了解胰島分化之幹細胞對於糖尿病動物的影響。
研究設計：人類臍帶間質幹細胞以分化培養基誘導後形成類胰島細胞團塊，以反轉錄聚合酶鏈式反應偵測類胰島細胞團塊中胰島相關基因和免疫螢光染色及酵素連結免疫吸附反應分析胰島荷爾蒙蛋白之表現情形。將類胰島細胞團塊移植至胰臟全切除手術的動物中觀測空腹血糖的變化以及移植後存活率。
研究結果：從反轉錄聚合酶鏈式反應、免疫螢光染色及酵素連結免疫吸附反應分析的結果顯示類誘導成胰島團塊具有胰島相關基因的表現同時也有胰島荷爾蒙蛋白的表現及分泌。移植類胰島團塊至胰臟全切除豬可以降低術後初期的空腹血糖且存活率有明顯差異。
研究結論：從我們的研究可以觀察到經胰島分化之間質幹細胞具有類似胰臟β細胞的特徵，移植類胰島團塊至胰臟全切除手術之豬可有效改善術後產生之高血糖症所造成的傷害並且增加存活率。

關鍵字： 人類臍帶間質幹細胞、胰臟全切除、糖尿病、類胰島細胞團塊

Summary

Objective: Diabetes mellitus is still a problem to yield an effective radical cure in current clinical medicine field. It has been shown that stem cells were not only clinically effective in therapeutic improvement on various diseases but also had the potential to be induced to insulin producing cells. This study was to investigate the functional influence of post-transplanted differentiated islet-like cell aggregates induced from umbilical cord stromal mesenchymal stem cells on the physiological changes and survival rates of total pancreatectomized pigs.
Design: The preparation of islet-like cell aggregates derived from Human umbilical cord stromal mesenchymal stem cells was performed by an induced protocol through several differentiation mediums. Methods involved in the quantitative analyses were the reverse transcription PCR for the assessment of the pancreatic lineage gene expression, ELISA and immunofluorescence staining for hormone protein expression. Fasting blood glucose level and survival rate were the measures to evaluate the effectiveness of the post-transplanted islet-like cell aggregates on the total pancreatectomized pigs.
Result: Significant levels of pancreatic lineage gene expressions as well as the amount of pancreatic hormone protein secretion were observed in the Islet-like cell aggregates detected by reverse transcription PCR, ELISA and immunofluorescence staining. The result data showed the effectiveness of the islet-like cell aggregates yielding significant decrease in the fasting blood level and increase in the survival rate of total pancreatectomized pigs with post-transplantation of these cell aggregates.
Conclusion: Islet-like cell aggregates bear the insulin secretion potential as the pancreatic beta cells. Post-transplantation of islet-like cell aggregates to the total pancreatectomized pigs solved out the lesion recovery problem reducing previous hyperglycemia condition so that the survival rates elevated.

Keywords: Human mesenchymal stem cells (MSCs), Total pancreatectomy, Diabetes, Islet-like cell aggregates

目 錄

論文電子檔著作權授權書............................................ i
論文審定同意書.....................................................ii
誌謝 ............................................................ iii
中文摘要.......................................................... iv
英文摘要......................................................... . v
目錄.............................................................. vi
第一章 緒論.........................................................1
1.1 糖尿病.......................................................1
1.2 幹細胞研究及應用.............................................3
1.3 幹細胞胰島分化..............................................4
1.4研究目的.....................................................6
第二章 材料與方法..................................................7
2.1實驗藥劑與材料..............................................7
2.2幹細胞初代培養..............................................11
2.3幹細胞繼代培養及放大........................................11
2.4間質質幹細胞胰島分化培養....................................12
2.5 RNA分離、純化和反轉錄聚合酶鏈式反應(RT-PCR) ...............12
2.6細胞免疫螢光染色............................................13
2.7酵素連結免疫吸附反應分析....................................14
2.8糖尿病動物模式之建立及動物生理變化測量......................14
2.9數據統計分析................................................15
第三章 實驗結果....................................................17
3.1偵測分化之間質幹細胞胰島相關基因之表現量....................17
3.2偵測分化之間質幹細胞胰島荷爾蒙蛋白之表現量..................17
3.3移植分化間質幹細胞對於糖尿病模式動物之生理變化以及存活率....18
第四章 討論........................................................19
第五章 參考文獻....................................................22
第六章 圖表與圖表說明..............................................25
第七章 附錄........................................................30

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