在本論文研究中嘗試利用兩種不同的方式來減輕糖尿病症狀或治療第2型糖尿病。首先，研究探討pepino polyphenolic extract（PPE）多酚提取物對糖尿病性神經病變的作用。PPE是一種多酚，其中含有香豆酰基和咖啡酰基衍生物。研究結果顯示PPE可以顯著改善糖尿病小鼠的血糖。經PPE治療後可以保留穀胱甘肽（GSH）水平、並減少硫代巴比妥酸反應物質（TBARS）水平、降低ROS、降低白細胞介素（IL）-6、腫瘤壞死因子（TNF）-α，以及果糖和糖化在糖尿病小鼠的坐骨神經中的晚期糖基化終末產物（AGE）的中間體和前體，例如甲基乙二醛（MG）和N-（羧甲基）賴氨酸（CML）。 PPE還具有改善糖尿病小鼠神經的作用-顯示促進許多小髓纖維束的分裂。 此外，PPE可以保持穀胱甘肽過氧化物酶（GPX）的蛋白和mRNA水平，並且減少坐骨神經中晚期糖基化終產物（RAGE）的醛糖還原酶（AR）和受體的表達。上述結果顯示PPE可以減輕糖尿病周圍神經中的氧化，炎症和糖原壓力。
由於間質幹細胞（MSCs）能夠分化成不同具功能性的細胞，因此，以此方式的細胞治療已成為組織修復與再生的應用潛力。本論文研究的第二部分即是探討結合第1型膠原蛋白奈米分子與菸鹼醯胺（NCT）和exendin-4（EX4）對MSCs分化成胰島素分泌細胞的影響，並評估此分化的胰島素分泌細胞對大鼠第2型糖尿病（T2DM）的治療效果。初步結果顯示，在結合第一型膠原蛋白奈米分與NCT，MSCs表達增加的Nkx6.1和早期PDX-1 mRNA表現，顯示MSCs能有效分化成具胰島素分泌的細胞。此外，添加第一型膠原蛋白奈米纖維可增加具胰島團簇之分化細胞數量。T2DM動物治療植入實驗中顯示：在植入NCT /第一型膠原蛋白奈米分子/ EX4所分化的細胞後，T2DM大鼠可恢復其正常的HbA1值，並顯示修復的胰島結構。簡言之，植入的細胞治療配方顯示可修復T2DM大鼠中被破壞的胰島，並能恢復葡萄糖的調節作用。

In this present study, we have utilized two different approaches tried to alleviate diabetes mellitus (DM) related stress or advanced to treat the DM. The effect of pepino polyphenolic extract (PPE) on diabetic neuropathy was examined first. PPE was demonstrated to contain coumaroyl and caffeoyl derivatives among polyphenols. The results demonstrated that PPE significantly improved glycaemic control in diabetic mice. These PPE treatments reserved the glutathione (GSH) level, and decreased the thiobarbituric acid reactive substances (TBARS) level, reactive oxygen species (ROS), interleukin (IL)-6, tumour necrosis factor (TNF)-alpha, fructose, and glycation intermediates and precursors of advanced glycation end products (AGEs), such as methylglyoxal (MG) and N-(carboxymethyl)lysine (CML), in the sciatic nerves of diabetic mice. PPE also had the effects in improving the nerves of diabetic mice, showing disorganization of the fascicle with numerous small myelinated fibers. The PPE can retain the protein and mRNA levels of glutathione peroxidase (GPX), and decrease the expressions of aldose reductase (AR) and the receptor for the advanced glycation end product (RAGE) in sciatic nerves. These findings support that pepino polyphenolic extract could attenuate oxidative, inflammatory and glycative stress in diabetic peripheral nerves.
Advances in mesenchymal stem cells (MSCs) and cell therapies have been promising approaches to treat (DM) since their potent capacities for differentiation into various functional cells match the demands of tissue repair and regeneration. The aim of this study is secondly to examine the effects of nano-sized type I collagen molecules in combination with nicotinamide (NCT) and exendin-4 (EX4) on MSCs differentiation into insulin-secreting cells in vitro and to evaluate their reparative effects against type 2 diabetes mellitus (T2DM) rat in vivo. The obtained results revealed that MSCs showed increased differentiation into insulin-secreting cells with higher mRNA expression for Nkx6.1 and early PDX-1 in the presence of NCT and nano-sized type I collagen molecules. Addition of nano-sized type I collagen fibrils increased morphologically islet-like clusters in differentiated cells. T2DM rats reverted to their normal HbA1 values and exhibited structurally repaired islets in the pancreas implanted with NCT/nano-sized collagen I molecule/EX4-incubated differentiated cells. In short, the combined recipe showed reparative actions on the destructive islet of Langerhans in the pancreas coupled with glucoregulatory effects in T2DM rats in vivo.

Republic of China i
致謝 i
摘要 ii
ABSTRACT iv
目錄 vii
表目錄 xi
圖目錄 xii
第一章概論 1
1-1 前言 1
1-2 治療糖尿病-再生醫學 2
1-3 糖尿病的概況及簡介 5
1-3-1 糖尿病定義 5
1-3-2糖尿病的分類 6
1-4糖尿病臨床診斷 8
1-4-1糖尿病診斷標準 8
1-4-2糖化血色素（HbA1c）指標與糖尿病關係 8
1-4-3第2型糖尿病的病因 9
1-5 糖尿病臨床症狀 13
1-5-1 三多症狀 13
1-5-2 高血糖 13
1-5-3 糖尿 13
1-5-4 酮尿 13
1-5-5 體重減輕 13
1-5-6 類似流行性感冒的症狀 13
1-5-7 視力模糊 14
1-5-8 創傷痊癒緩慢或經常受到感染 14
1-6 併發症 14
1-6-1 大血管疾病 14
1-6-2 小血管疾病 14
1-6-3 神經病變[52.53] 15
1-6-4 末梢動脈血管 15
1-6-5 微循環不良 15
1-6-6 性功能障礙 15
1-7 病理生理學 15
1-7-1 糖尿病的代謝作用[55.56] 16
1-7-2 葡萄糖的利用降低 16
1-7-3 脂肪活動增加 16
1-7-4. 蛋白質的利用增加 16
1-8 糖尿病與氧化壓力 17
1-9 糖尿病與多元醇路徑 19
1-10 第2型糖尿病與胰島素阻抗 20
1-11 香瓜茄 20
第二章研究目的 24
第三章 實驗藥品、材料與設備 25
3-1 實驗材料 25
3-1-1特殊飼料添加：1.澎湖地區種植之香瓜茄2.高脂飼料配方：市售包裝豬油。 25
3-1-2實驗動物：1.由國家實驗研究院國家動物中心購進4~5週齡雄性BALB/cByJNarl 小鼠2. 樂斯科生物科技股份有限公司，品系為Sprague-Dawley 雄性大白鼠 (250±20 g)。 25
3-1-3實驗誘發試藥：美國Sigma公司之Streptozocin(STZ) 25
3-1-4 實驗分析用藥：第一部分 (廠商/國家) 25
3-1-5 實驗分析用藥：第二部分 26
3-2 儀器設備 28
3-2-1儀器設備：第一部份(型號/廠商/國家) 28
3-2-1儀器設備：第二部份 29
第四章 研究材料與方法 30
4-1 香瓜茄萃取物製備 30
4-1-1流程步驟 30
4-1-2給予細胞之香瓜茄水萃取物製備 30
4-1-3香瓜茄水萃物之透析 30
4-2 香瓜茄萃取物高效液相色譜 - 電噴霧電離串聯質譜（HPLC / ESI-MS-MS）分析 31
4-3不同構形膠原蛋白奈米生物分子製備 31
4-4 間質幹細胞萃取/培養 31
4-5間質幹細胞誘導分化 31
4-6 免疫螢光染色 32
4-7 胰島素分泌測定 33
4-8逆轉錄聚合酶鏈式反應 (RT-PCR) 33
4-8-1 RNA抽取 33
4-8-2 cDNA合成 33
4-8-3 Real-time PCR 34
4-9 實驗動物來源及飼養 34
4-9-1 高脂飼料配方 35
4-9-2 第二型糖尿病鼠飼養 35
4-9-3 鏈脲佐菌素作用 35
4-10 STZ 誘發第二型糖尿鼠 36
4-11葡萄糖耐受度測試 36
4-12 胰島素-葡萄糖耐受度測試 36
4-13 誘導分化胰島素分泌細胞之第2型糖尿病病鼠動物植入治療實驗 36
4-14 H&E 染色分析 37
4-15 統計分析 37
第五章 實驗結果 38
5-1 以STZ誘發第2型糖尿病小鼠之動物模式特徵 38
5-1-1體重 38
5-1-2 三多症狀 39
5-1-3血糖值 39
5-1-4 insulin 胰島素 41
5-1-5 oral glucose tolerance test(OGTT) 葡萄糖耐量試驗 42
5-2 飲食中添加不同濃度生的香瓜茄，對以STZ誘發第2型糖尿病 小鼠血清氧化壓力的影響 42
5-3 PPE減少STZ誘導的糖尿病 43
5-4胰腺分離用於H＆E染色 45
5-5 PPE改善坐骨神經的組織學染色 46
5-6 PPE調節GPX，AR和RAGE的表達 47
5-7間質幹細胞的形態特徵觀察 50
5-8胰島素分泌 52
5-9即時聚合酶鏈式反應分析 53
5-10 SD 大鼠高脂飼料餵食模型 55
5-11 胰島素-葡萄糖耐受性測試 57
5-12 誘導分化胰島素分泌細胞之第2型糖尿病病鼠動物植入治療實驗 59
5-13 糖化血色素(HbA1c)含量測試 61
5-14 H&E 染色分析結果 62
第六章 討論 65
第七章 結論 67
第八章參考文獻 69

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