臺灣博碩士論文加值系統

中文摘要
糖尿病是全世界日益普遍的慢性疾病，在臨床證實噻唑烷二酮類藥物可以改善患有胰島素阻抗性患者的胰島素敏感性，但被發現其具有肝毒性等副作用。ENERGI 為本實驗室所發現之新穎 AMPK 活化劑，為一水溶性之小分子代謝化合物，透過活化 AMPK 活性上升可以抑制脂肪細胞之轉錄因子進而有效的抑制脂肪細胞分化。本論文研究方向是探討 ENERGI 與噻唑烷二酮類藥物 Rosiglitazone 的協同效果。方法為培養 3T3-L1 小鼠前驅脂肪細胞分化 9 天加入 ENERGI 及Rosiglitazone 進行 western blot 分析，結果顯示 PPAR-γ 與 C/EBPα在 添 加 ENERGI 之 後 的 表 現 量 都 有 提 升 ， 而 Leptin 在 只 有Rosiglitazone 的時候似乎沒有明顯差異，但再加入 ENERGI 後會提升表現量，後續影響的機轉有待探討。另外將 Oil Red O 染色的果顯示出 ENERGI 能促進脂肪細胞中油滴之累積，在有添加濃度Rosiglitazone 及 ENERGI 的時候油滴累積的狀況都較對照組來的更好。顯示在 ENERG I 與 Rosiglitazone 協同作用下可以降低 Rosiglitazone使用量，推測有機會降低其副作用而達到減毒增效的效果。

英文摘要
Diabetes is an increasingly common chronic disease worldwide, and it has been confirmed in trampolines that thiazolidinediones can improve insulin sensitivity in patients with insulin resistance, but have been found to have side effects like hepatotoxicity. ENERGI is a novel AMPK activator discovered in our laboratory. It is a water-soluble small molecular compound. By increasing the activity of activated AMPK, it can inhibit the activity of transcription factors in fat cells and the differentiation of adipocytes. The aim of this research is to explore the synergistic effect of ENERGI and Rosiglitazone. After the cultured cells 3T3-L1 were differentiated for 9 days, ENERGI and Rosiglitazone were added and western blot analysis was conducted. The results showed that the expression of PPAR-γ and C/EBPα increased after ENERGI being added. There was no distinct difference when Leptin was with Rosiglitazone. The mechanism of ENERGI being added and promoted being increased awaits further discussions. In addition, the results of Oil Red O staining showed that ENERGI could promote the accumulation of oil droplets in fat cells, and the efficacy was better when Rosiglitazone and ENERGI were added. It shows that the Synergism of ENERGI and Rosiglitazone can reduce the use of Rosiglitazone. It is also assumed that the side effects of Rosiglitazone and toxicity can be reduced and effects cab be multiplied.

目錄
目錄 ................................................................................................... 1
圖目錄 ............................................................................................... 5
中文摘要............................................................................................ 6
英文摘要............................................................................................ 7
縮寫表 ............................................................................................... 8
第 1 章、 文獻回顧 ............................................................................ 9
1.1. 糖尿病 ( Diabetes mellitus )..........................................................9
1.2. 主要治療第二型糖尿病的方式 ..................................................10
1.2.1. AICAR......................................................................................12
1.2.2. 噻唑烷二酮藥物 ( Thiazolidinediones, TZD ) .......................15
1.2.3. Pioglitazone............................................................................18
1.3. 3T3-L1 脂肪細胞...........................................................................20
2
1.3.1. AMPK 在脂肪組織中的調節.................................................23
1.3.2. AMPK 於脂肪細胞分化之相關性.........................................24
1.3.3. Peroxisome proliferator-activated receptor ( PPAR )..............25
1.4. 小分子化合物 ENERGI................................................................26
1.5. 實驗動機與目的..........................................................................26
第 2 章、 材料與方法....................................................................... 27
2.1. 細胞培養 .....................................................................................27
2.1.1. 貼附型細胞及繼代...............................................................27
2.2. 細胞保存 .....................................................................................31
2.3. 細胞破裂法..................................................................................32
2.4. 蛋白質定量..................................................................................35
2.5. BCA method ..................................................................................36
2.6. 電泳檢定法..................................................................................37
3
2.7. 蛋白質轉印法..............................................................................41
2.8. 酵素免疫染色..............................................................................44
2.9. 脂肪細胞分化..............................................................................47
2.10. 油紅染色 (Oil Red O staining) ...................................................48
2.11. 細胞內 Triglyceride 測定..........................................................50
2.12. RNA 萃取 ...................................................................................52
2.13. 定量即時聚合酶連鎖反應 (qPCR)............................................54
2.14. 葡萄糖攝取測定法....................................................................56
第 3 章、 實驗結果 .......................................................................... 57
3.1. 3T3-L1 脂肪細胞分化處理 ENERGI 與 Rosiglitazone 之影響 ..57
3.2. 3T3-L1 脂肪細胞分化不同時期處理 ENERGI 與 Rosiglitazone
於細胞內三酸甘油脂含量之影響 ......................................................57
3.3. 3T3-L1 脂肪細胞分化處理 ENERGI 與 Rosiglitazone 於細胞內
三酸甘油脂含量之影響......................................................................58
3.4. 3T3-L1 脂肪細胞分化處理 ENERGI 與 Rosiglitazone 之脂肪細
胞相關 mRNA 表現量.........................................................................58
4
3.5. 3T3-L1 脂肪細胞分化處理 ENERGI 與 Rosiglitazone 之脂肪細
胞相關蛋白質表現..............................................................................59
第 4 章、 討論.................................................................................. 68
第 5 章、 文獻參考 .......................................................................... 70

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