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研究生:鄭汝翔
研究生(外文):Cheng, Ju-Hsiang
論文名稱:黃金銀耳於糖尿病大鼠模式肝臟中降血糖機制之探討
論文名稱(外文):Studies on Mechanism of Anti-hyperglycemic Activity of Tremella mesenterica Retz.:Fr. in Liver of Diabetic RatsTremella mesenterica Retz.:Fr. in Liver of Diabetic RatsStudies on Mechanism of Anti-hyperglycemic Activity of Tremella mesenterica Retz
指導教授:盧錫祺盧錫祺引用關係
指導教授(外文):Lu, Hsi-Chi
學位類別:碩士
校院名稱:東海大學
系所名稱:食品科學系
學門:農業科學學門
學類:食品科學類
論文種類:學術論文
論文出版年:2010
畢業學年度:98
語文別:中文
論文頁數:116
中文關鍵詞:胰島素敏感性肝臟黃金銀耳酸性多醣體糖尿病降血糖
外文關鍵詞:Tremella mesenterica Retz.:Fr.hepatocytesacidic polysaccharidesinsulin sensitivityDiabetes
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糖尿病是由於胰島素分泌不足或其無法作用所形成的內分泌疾病,其形成原因複雜,因此目前無切確的成因。近年由於飲食精緻化,導致糖尿病患於全球人口有逐年攀升的趨勢,並且有年輕化的現象。有研究指出,糖尿病的形成可能是由於基因與生活形態所造成,如久坐少動的習慣及能量過剩的情況,在這些因子交互作用之下便造成胰島素阻抗現象,如無及時改善,最終將形成第二型糖尿病。糖尿病患於長期血糖異常下,容易衍生出併發症,如能有效控制血糖將可避免出現的機率。目前控制血糖主要以運動及飲食控制等非藥物及天然食材為發展主軸,因此,本實驗對於黃金銀耳進行研究。於先前的動物研究中證實,黃金銀耳具有降低糖尿病大鼠高血糖的症狀,但卻未改善其胰島素分泌不足的缺失,因此推測黃金銀耳可能經由更有效的訊息傳遞,來提高組織間胰島素敏感性,達到降血糖的目的。由於黃金銀耳之血糖調控機制至今尚未完全建立,因此本研究將針肝臟此一重要的血糖調控器官進行降血糖機制探討。爲釐清黃金銀耳的餵食是否影響胰島素傳訊與肝臟組織中葡萄糖的攝入以及脂聯素傳訊,本研究將對此三條路徑之傳訊分子進行探討。本研究以酵素免疫連結法分析血漿中脂聯素濃度,及利用反轉錄同步定量聚合酶連鎖反應分析基因層次之表現量,並以西方轉漬法分析蛋白質表現量。結果顯示,在胰島素傳訊方面,黃金銀耳可提升胰島素受體(IR)及胰島素受體基質(IRS-1、IRS-2)基因的轉錄及其蛋白質的表現,及提升Akt與其磷酸化程度,增強肝臟組織中胰島素傳訊。於葡萄糖的轉運方面,黃金銀耳則增加葡萄糖轉運蛋白(GLUT2)於膜上及質內的蛋白質密度及胞內葡萄糖激酶基因與蛋白質的表現。在脂聯素方面,黃金銀耳增加實驗大鼠脂聯素的分泌,並且於脂聯素傳訊中,提高AMPK的蛋白質表現及其磷酸化程度。另一方面,於黃金銀耳各型態物中,發現以純化之酸性多醣體最具顯著改善各傳訊路徑之功效,因此我們認為黃金銀耳可能藉由酸性多醣體達到降血糖效果。黃金銀耳在動物實驗降血糖能力,可能是透過增強肝臟組織中胰島素的傳訊及增加肝臟細胞對於葡萄糖的攝入,以及促進脂聯素的分泌,藉此提升脂聯素傳訊,達到降血糖的效果。
Diabetes, due to insufficient secretion or malfunction of insulin, is an endocrine disease. The pathogenesis of diabetes is complicated and still unclarified. Because of delicate diet, the global prevalence and young population increased dramatically in recent years. Studies indicate that the genetic deposits and lifestyles might influence the progression of diabetes, especially lack of exercises and energy excess which would ultimately result in insulin resistance and type II diabetes without proper intervention. Diabetic patients with long-term abnormal blood glucose might result in complications, which can be prevented by effective blood glucose control. Currently, the regimes of blood glucose control include exercise and diet, with intervention supplementation of non-pharmaceutical and natural ingredients. In previous animal studies, Tremella mesenterica Retz.:Fr. was proved to be effective in decreasing blood concentration in diabetic rats, without improving the deficiency status of insulin.Therefore, the lowering of blood glucose level might be attributed to an enhanced insulin sensitivity in tissues via efficient signal transduction in Tremella mesenterica Retz.:Fr. fed rats. In order to further elucidate the mechanism of utilized by Tremella mesenterica Retz.:Fr., liver was chosen for its importance in regulating glucose homeostasis. To clarify the effects brought with Tremella mesenterica Retz.:Fr. feeding, the insulin signal pathway, glucose uptake and adiponectin signal pathway were investigated. Oral feeding of Tremella mesenterica Retz.:Fr. increased the mRNA and protein expression levels of IR, IRS-1 and IRS-2. Futhermore, Akt production as well as phosphorylation of Akt were enhanced, indicating the enhancement of insulin signaling in liver. Elevated expression of membranous and cytosolic GLUT2 protein and cytosolic fraction of Akt were observed in hepatocytes. Serum level of adiponectin and as well as phosphorylation of AMPK were also elevated. Gavage with acidic polysaccharides purified from Tremella mesenterica Retz.:Fr. showed most prominent effect in activating two signaling pathways. Therefore acid polysaccharides are supposed to be the main component of Tremella mesenterica Retz.:Fr. that exerted anti-diabetic activities. Elevated hepatic insulin signaling and glucose uptake, enhanced adiponectin secretion and adiponectin signaling may all contribute to the hypoglycemic effects of Tremella mesenterica Retz.:Fr. in experimental rats.
摘要 1
英文摘要 3
一、前言 5
1-1黃金銀耳與其生理功效 5
1-2糖尿病 5
1-2-1糖尿病的分類 6
1-2-2糖尿病診斷 7
1-2-3糖尿病併發症 8
1-2-4糖尿病的治療 8
1-3肝臟組織與血糖恆定 8
1-4肝臟中胰島素傳訊 10
1-4-1胰島素傳訊分子 11
1-5肝臟中葡萄糖轉運與代謝途徑 14
1-5-1葡萄糖轉運蛋白 14
1-5-2六碳糖激酶 15
1-5-3葡萄糖的代謝與肝醣合成 16
1-5-4肝臟組織之糖質新生作用 16
1-6脂聯素傳訊 16
1-6-1脂聯素與其傳訊分子 17
1-7前驅動物實驗 20
1-7-1藥物誘發第二型糖尿病模式 20
1-7-2 Streptozotocin(STZ)藥物與Nicotinamide(NT)藥物 20
1-7-3前驅實驗結果 21
1-8研究目的 22
二、材料方法 23
2-1 動物實驗 23
2-2 脂聯素酵素免疫連結分析 24
2-3 肝臟組織抽取RNA 24
2-4 cDNA製備 25
2-5 同步定量聚合酶連鎖反應 25
2-6 肝臟組織之蛋白質萃取 26
2-7 蛋白質定量 27
2-8 西方轉漬法 27
2-9統計分析 28
三、結果 31
3-1 胰島素傳訊方面 31
3-1-1肝臟中胰島素受體(IR)及胰島素受體基質(IRS-1、IRS-2) mRNA相對表現量 31
3-1-2肝臟中胰島素受體(IR)、胰島素受體基質(IRS-1、IRS-2)及PKB/Akt蛋白質相對表現量與其磷酸化量 36
3-2葡萄糖轉運及代謝方面 44
3-2-1肝臟中葡萄糖轉運蛋白(GLUT2)及葡萄糖激酶(GK)之mRNA相對表現量 44
3-2-2肝臟中葡萄糖轉運蛋白(GLUT2)及葡萄糖激酶(GK)之蛋白質相對表現量與釐清葡萄糖轉運蛋白(GLUT2)是否具有轉位現象 47
3-2-3肝臟組織之糖質新生關鍵酵素(PEPCK) mRNA相對表現量 55
3-3脂聯素傳訊 57
3-3-1血清中脂聯素 57
3-3-2肝臟組織中脂聯素受體(AdipoR2)基因相對表現量 58
3-3-3肝臟組織中AMP-activated protein kinase (AMPK)蛋白質相對表現量 60
四、討論 65
4-1前驅動物實驗數據與本研究之探討 65
4-2胰島素傳訊方面 66
4-3肝臟組織中葡萄糖轉運及代謝方面 68
4-4 脂聯素傳訊方面 69
五、總結 72
六、未來展望 76
附圖一、動物試驗流程 78
附圖二、生化分析數據 78
附表一、前驅動物實驗生化分析數據表 81
附表二、實驗中使用的化合物 82
附圖三、同步定量聚合酶連鎖反應結果電泳圖 84
附圖四、西方轉漬法結果底片 93
參考文獻 110
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