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研究生:曾千育
研究生(外文):Chien-Yu Tseng
論文名稱:GABA茶對高血糖大白鼠抗心肌凋亡分子機轉之探討
論文名稱(外文):Molecular Mechanisms of Anti-apoptotic Effects of GABA Tea on Diabetic Rat Hearts
指導教授:王雪芳王雪芳引用關係黃志揚黃志揚引用關係
指導教授(外文):Hsueh-Fang WangChih-Yang Huang
學位類別:碩士
校院名稱:弘光科技大學
系所名稱:生物產業研究所
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2009
畢業學年度:97
語文別:中文
論文頁數:111
中文關鍵詞:Wistar 鼠GABA茶高血糖西方點墨法細胞凋亡
外文關鍵詞:WistarGABA TeaHyperglycemiaWestern BlottingApoptosis
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GABA茶是一種含豐富γ-胺基丁酸 (γ-aminobutyric acid, GABA) 的純天然茶葉製品。為了瞭解GABA茶對於高血糖改善效果之影響,本研究以Stereptozotocin (STZ) 誘發高血糖反應的Wistar大白鼠,經攝取不同劑量之GABA茶水萃取液後,針對血清生化分析以及心臟組織對於抗細胞凋亡之分子機轉探討。將八週齡大之雄性Wistar大白鼠分成六組,前兩組為未誘發高血糖組,分別餵食水 (Normal control) 和高劑量GABA茶(3010 μg GABA Tea/day/rat, normal-HG)。另外四組先以55 mg/kg/rat STZ腹腔注射誘導Wistar大白鼠產生高血糖反應後,分別餵食水 (DM-control) 及低、中、高劑量 (3.01, 30.1, 3010 μg/day/rat; DM-LG, DM-MG, DM-HG) GABA茶。所有試驗動物自由攝食飼料與分別管餵實驗樣品、安慰劑 (水) 6週後,犧牲,採血並取其心臟組織,分析血清GABA、脂質、賀爾蒙含量,並進一步探討餵食GABA茶水萃取物對高血糖鼠之心肌細胞凋亡及存活等相關路徑。結果:GABA茶水萃取物可降低餵食GABA茶水萃取物之高血糖鼠血糖及血脂,以DM-MG組的效果最佳外,並具有降低normal-HG血糖及血清三酸甘油脂,且有降低空腹血清胰島素(Insulin)及增加incretins (glucagon-like peptite-1 (GLP-1) 及glucose-dependent insulinotropic polypeptide (GIP)) 含量具有調節血糖與改善胰島素阻抗 (insulin resistance, IR) 之效果,且同樣以DM-MG組的效果最佳。由組織染色分析 (Histological analysis) 心臟切片中發現,GABA茶水萃取物皆有減緩高血糖鼠心肌細胞傷害,在TUNEL assay亦觀察到具有抑制高血糖鼠心肌細胞凋亡的效果,西方點墨法 (Western Blot) 的分析結果則證明了,GABA茶因可抑制death receptor dependent及mitochondria dependent此兩條凋亡路徑,而抑制了高血糖鼠之心肌細胞凋亡,且同時活化了IGF-I及PI3K/Akt心肌存活路徑來保護高血糖鼠的心肌細胞的效果以餵食低或中劑量GABA茶水萃取物為佳 (DM-LG, DM-MG group)。因此,餵食中劑量之GABA茶水萃取物降低血脂作用最佳,且更具有抗心肌肥大的效果,但低劑量已具有其療效。以上結果得知,適量的GABA 茶水萃取物具有改善高血糖鼠血糖、血脂、胰島素阻抗的效果,以及保護高血糖鼠心肌細胞。
GABA tea has the high content of g-aminobutyric acid (GABA). To find how GABA tea affects the hyperglycemia, the Stereptozotocin (STZ) administered Wistar diabetic rats were used in this study. For finding the apoptotic mechanisms, the male eight-week-old Wistar rats were separated into six groups. At the seventh day, the first two groups (not-induced hyperglycemia) were fed with water (Normal control) and GABA tea with high dose GABA 3010 μg /day/rat (Normal-HG), respectively. The other four groups with STZ induced hyperglycemia were fed with water (DM-control) and low, medium, and high dose GABA (3.01, 30.1, 3010 μg /day/rat; DM-LG, DM-MG and DM-HG). After six weeks, the cardiac tissues of all test animals were taken for analyzing, and the contents of GABA, lipids and hormones in blood were also examined.
The results of hematologic analysis showed that DM-MG demistrated the best effect to reduce the blood sugar and lipids. Besides, Normal-HG which was fed with the high dose of GABA in tea could also reduce the blood sugar and the triglyceride. The results of blood serum hormone measurement showed that reduced the fasting serum insulin and increased incretins (glucagon-like peptite-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP)) had the effect to adjust blood sugar and impoved insulin resistance, and DM-MG showed the best effect. The results of Histological analysis showed that the cardiac damage of rats with hyperglycemia could be reduced by fedding different dose of GABA tea. The TUNEL assay showed that extracts of GABA tea had the effect to inhibit the cardiomyocyte apoptosis. The Western Blot analysis showed that death of cardiomyocytes in the hyperglycemia rats was mediated through the receptor and mitochondria-dependent pathways, and GABA tea could inhibit these two pathway activity. Furthermore, GABA tea could activate IGF-I and PI3K/Akt signaling to protect the cardiomyocytes in the hyperglycemia rats. The study showed that DM-MG had also the best effect to resist the hypertrophy of cardiomyocytes. Besides, the results of GLP-1 and GIP measurement revealed that the extract of GABA tea had function to adjust the blood sugar, and groups of low and medium dose showed the best effect to resist the death of myocardial cells.
The overall results showed that taking the proper amount of GABA tea extract could improve the abnormal blood sugar, blood lipids and insulin resistance in the hyperglycemia rats, and protect cardiomyocytes in rats with hyperglycemia.
目 錄
頁次
中文摘要……………………………………………………………….. Ⅰ
英文摘要……………………………………………………………….. Ⅲ
目錄…………………………………………………………………….. Ⅴ
表目錄………………………………………………………………….. Ⅷ
圖目錄………………………………………………………………….. Ⅸ
第一章 前言…………………………………………………………... 1
第二章 文獻回顧…………………………………………………….. 3
一、糖尿病現況……………………………………………….. 3
二、認識糖尿病……………………………………………….. 3
(一)定義………………………………………………….. 3
(二)糖尿病分類………………………………………….. 4
(三)糖尿病症狀………………………………………….. 6
(四)糖尿病併發症……………………………………….. 7
(五)胰島素阻抗性……………………………………….. 9
三、茶與糖尿病……………………………………………….. 15
(一)茶的簡介……………………………………………. 15
(二)茶與糖尿病…………………………………………. 18
(三)茶與心血管疾病……………………………………. 19
四、茶與心肌細胞…………………………………………….. 21
(一)茶與細胞凋亡………………………………………. 21
(二)細胞程序性凋亡和壞疳……………………………. 23
五、認識GABA茶…………………………………………….. 28
六、GABA茶的生理功效…………………………………… 30
七、GABA茶與心血管疾病………………………………… 36
八、研究目的………………………………………………….. 37
第三章 材料與方法…………………………………………………… 38
一、試驗設計…………………………………………………. 38
二、實驗材料…………………………………………………. 39
三、實驗儀器…………………………………………………. 45
四、試驗方法…………………………………………………. 47
五、GABA茶水萃取物之成分測定………………………….. 48
六、實驗動物檢體收集及分析……………………………… 53
七、組織分析…………………………………………………. 55
八、統計分析…………………………………………………. 58
第四章 結果…………………………………………………………… 59
一、GABA茶水萃取物之GABA、總多元酚及礦物質含量... 59
二、GABA茶水萃取物對試驗鼠之血清脂質含量的影響….. 59
三、GABA茶水萃取物對試驗鼠之血清GABA及賀爾蒙
含量的影響……………………………………………... 60
四、GABA茶水萃取物對試驗鼠心臟重量與體重之影響…... 60
五、由心臟切片組織觀察GABA茶水萃取物對試驗鼠心臟型態的影響………………………………………... 61
六、由顯微鏡觀察餵食GABA茶水萃取物之試驗鼠心臟細胞的變化………………………………………………… 62
七、以TUNEL assay觀察試驗鼠心肌細胞凋亡…………….. 62
八、以Western blotting觀察心臟組織中蛋白質的變化……. 63
(一)高血糖與心肌細胞之Death receptor dependent pathway……………………………………………... 63
(二)高血糖與心肌細胞粒腺體的損傷………………… 64
(三) GABA茶水萃取物活化PI3K/Akt pathway保護心肌細胞……………………………………………… 66
第五章 討論…………………………………………………………… 67
第六章 結論…………………………………………………………… 73
第七章 圖表結果……………………………………………………… 76
第八章 參考文獻……………………………………………………… 97
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