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研究生:王姿琇
研究生(外文):Tzy-Shiow Wang
論文名稱:高甲硫胺酸飲食對大鼠熱中風之影響
論文名稱(外文):Effects of high methionine diet on heatstroke in rat
指導教授:楊彩秀楊彩秀引用關係
指導教授(外文):Tsai-Hsiu Yang
學位類別:碩士
校院名稱:嘉南藥理科技大學
系所名稱:營養與保健科技研究所
學門:醫藥衛生學門
學類:營養學類
論文種類:學術論文
論文出版年:2011
畢業學年度:99
語文別:中文
論文頁數:120
中文關鍵詞:同半胱胺酸S-腺核苷同半胱胺酸甲硫胺酸熱中風葉酸
外文關鍵詞:homocysteineS-adenosylhomocysteinefolic acidheatstrokemethionine
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根據我們先前研究證實,當動物體的血漿中具高濃度S-腺核苷同半胱胺酸或同半胱胺酸,可能是誘發熱中風生成的高危險因子之一。然而,目前卻未有利用飲食預防醫學延緩熱中風發生的研究。本研究目的,探討給予高量甲硫胺酸或高量葉酸飲食,對大白鼠血漿中的S-腺核苷同半胱胺酸或同半胱胺酸濃度之影響,以更加確定S-腺核苷同半胱胺酸或同半胱胺酸對於熱中風的影響,及是否可透過高量葉酸飲食作為預防熱中風損傷作用。將Sprague-dawley(S.D.)雄性大白鼠分為正常飲食控制組(Control)(正常飲食含有0.3% methionine和2.1 ppm folic acid)、高甲硫胺酸飲食組(High methionine diet; H.Met)(含有1.5% methionine和2.1 ppm folic acid)及高量葉酸飲食組(High folic acid diet; H.FA)(含有 0.3% methionine和1000 ppm folic acid)及高甲硫胺酸同時高葉酸飲食组(High methionine and folic acid diet; H.MF) (含有 1.5% methionine和1000 ppm folic acid),持續四週。飼養期間利用HPLC(High-Performance Liquid Chromatography)進行分析三組間血漿中同半胱胺酸(Homocystine; Hcy)、S-腺核苷同半胱胺酸(S-Adeosylhomo-
cysteine; SAH)及葉酸(Folic acid)的濃度變化。於四週後,將他們暴露於43℃室溫下誘發熱中風。而常溫對照組另外暴露於室溫下(24℃)。觀察他們的生理與病理變化,並且進行丙二醛(Malondialdehyde; MDA)、總抗氧化能力(Oxygen-radical absorbance capacity, ORAC)分析、去氧核糖核酸損傷(Deoxyribonucleic acid ladder; DNA Ladder)和病理組織切片。結果顯示,餵食高量甲硫胺酸飲食四週後的大白鼠,血漿中S-腺核苷同半胱胺酸或同半胱胺酸的濃度明顯高於其他組。高量甲硫胺酸飲食組在熱暴露下確實明顯縮短中風生成時間(64±0.7 min);其平均動脈壓(Mean arterial pressure; MAP)及心跳速率(Heartrate; HR)等生理參數也提早達高點後驟降。然而,餵食高量葉酸飲食四週後的大白鼠,血漿中S-腺核苷同半胱胺酸或同半胱胺酸的濃度明顯比其他組低。相反地,高葉酸飲食組在熱暴露下確實明顯延長中風生成時間(79±1.9 min);也明顯延緩其平均動脈壓(MAP)及腦血流量 (Cerebral blood flow; CBF)等生理參數下降的趨勢。在熱中風生成過程中,高量甲硫胺酸飲食的大白鼠,血漿及組織(腦部、心臟、肝臟和腎臟)中脂質過氧化和去氧核糖核酸損傷明顯比其他組提前出現且較為嚴重;反之,高量葉酸飲食組的大白鼠血漿中總抗氧化能力程度明顯增加,且血漿及組織(腦部、心臟、肝臟和腎臟)中脂質過氧化與去氧核糖核酸損傷程度明顯低於其他組。本研究證實,飲食中所攝入高量的甲硫胺酸確實會增加體內S-腺核苷同半胱胺酸或同半胱胺酸生成,確實與加速熱中風生成具有密切相關性;相反之,飲食中攝入高量葉酸的確明顯降低體內S-腺核苷同半胱胺酸及同半胱胺酸之濃度,不但也明顯延後熱中風之生成,同時也降低大白鼠血漿或組織中脂質過氧化與去氧核糖核酸損傷之程度。因此,可透過飲食中攝取或補充葉酸來改善體內S-腺核苷同半胱胺酸或同半胱胺酸濃度達到預防熱中風發生的效果。
According to our previous studies, the high plasma levels of S-adenosyl- homocysteine or homocysteine, one of the high-risk factors of heatstroke, may be easy to induce heatstroke induction. However, there is less attention on the use of dietary preventive medicine in treatment of heatstroke. Purpose of this study is firstly to observe the plasma levels of S-adenosylhomocysteine and homocysteine in rats after feeding the methionine or folic acid diet. Furthermore, it would be examined the effect of treatment with high folic acid diet on the physiological and pathological changes during the heatstroke formation. Male Sprague-dawley (S.D.) rats were used in this study and divided into normal diet control group (Control) (normal diet containing 0.3% methionine and 2.1 ppm folic acid), high methionine diet group (High methionine diet; H.Met) (containing 1.5% methionine and 2.1 ppm folic acid), high folic acid diet group (High folic acid diet; H. FA) (containing 0.3% methionine and 1000 ppm folic acid), and high methionine and folic acid diet group (H.MF) (containing 1.5% methionine and 1000 ppm folic acid) for four weeks. The plasma levels of homocysteine and S-adenosylhomocysteine change in each group rats were monitored by HPLC (High-Performance Liquid Chromatography) analysis. After dietary pretreatment for four weeks, rats were exposed to ambient temperature of 43℃ to induce heatstroke. In addition, rats of the room temperature control group exposed to room temperature (24 ℃). Rats were observed their physiological and biochemical changes, plasma level of the lipid peroxidation (MDA analysis) and oxygen-radical absorbance capacity (ORAC analysis), and deoxyribonucleic acid ladder and histopathology examination in the vital organs during heatstroke induction. After pretreatment with high methionine diet for four weeks, the results show that the levels of homocysteine and S-adenosylhomocysteine in plasma were significantly higher than other groups. Rat of high methionine diet group were significantly shrunk the duration time of heatstroke induction (64±0.7 min), and values of the mean arterial pressure (MAP), heart rate (HR) and other physiological parameters were early dropped from the normal range after heatstroke formation. However, after pretreatment with high folic acid diet for four weeks in rats, the levels of homocysteine and S-adenosylhomocysteine in plasma were significantly lower than other groups, and could reverse the influence of high methionine dietary feeding on abnormal plasma biochemical levels. Meanwhile, rats of high folic acid diet group were significantly prolonged the duration time of heatstroke induction (79±1.9 min), and significantly improved the heatstroke-induced arterial hypotension, cerebral ischemia and high oxidative situation. During heatstroke induction, it showed that lipid peroxidation and deoxyribonucleic acid ladder in plasma and vital organs (brain, heart, liver and kidney) in rats of high methionine dietary feeding group were significantly higher than other groups. In contrast, rats of pretreatment with high folate diet group displayed appreciate performance in the plasma level of oxygen-radical absorbance capacity, and ameliorated the conditions of high lipid peroxidation and deoxyribonucleic acid ladder fragmentation in plasma and vital organs. This study demonstrated, when high levels of methionine diet feeding, the body did increase the generation of homocysteine and S-adenosylhomocysteine, and accelerate the generation of heatstroke is closely associated. On the contrary, dietary of high levels folic acid intake did significantly decrease concentration of S-adenosylhomocysteine and homocysteine in body. Not only significantly decrease the generation of heatstroke, but also decrease the lipid peroxidation and deoxyribonucleic acid ladder of plasma or tissue in rats. Therefore, through diet intake or supplements of folic acid may be a good strategy to diminish the heatstroke-induced damage or keep away from the heatstroke induction .
目 錄
中文摘要 …………………………………………………………………….. Ⅰ
英文摘要 …………………………………………………………………….. Ⅳ
誌 謝 ………………………………………………………………………. VII
壹、 前言 ………………………………………………………………….. 1
一、 胺基酸(Amino acid; AA)與甲硫胺酸(Methionine; Met) …………... 1
二、 甲硫胺酸代謝循環(Methionine cycle) ……………………………… 2
三、 同半胱胺酸(Homocysteine; Hcy) …………………………………… 4
四、 葉酸(Folic acid) ……………………………………………………… 8
五、 S-腺核苷甲硫胺酸(S-adenosyl- methionine; SAM)及S-腺核苷同半胱胺酸(S-adenosylhomocysteine; SAH) …………………………... 11
六、 熱中風(Heatstroke) ………………………………………………… 16
貳、 研究動機及目的 …………………………………………………… 22
研究架構 …………………………………………………………… 24
參、 材料與方法 ………………………………………………………… 25
一、 實驗動物 …………………………………………………………. 25
二、 研究方法 …………………………………………………………. 25
三、 餵食期間大白鼠血漿中同半胱胺酸、半胱胺酸、SAH、SAM、葉酸濃度之測定 ……………………………………………………. 28
四、 基本動物手術及熱中風誘發 ……………………………………. 32
五、 血漿及組織中脂質過氧化測定 …………………………………. 35
六、 血漿總抗氧化能力(Oxygen-radical absorbance capacity, ORAC)測定 …………………………………………………………………. 37
七、 去氧核糖核酸損傷(Deoxyribonucleic acid ladder; DNA Ladder)分析 …………………………………………………………………. 39
八、 病理組織學的探討 ………………………………………………. 40
九、 統計分析 …………………………………………………………. 43
肆、 結果 ………………………………………………………………… 44
一、 動物生長及飼料攝取狀況 ………………………………………. 45
二、 觀察給予高量甲硫胺酸或高葉酸飲食期間,對大白鼠血漿中半胱胺酸濃度與同半胱胺酸濃度之影響 ……………………………. 48
三、 觀察給予高量甲硫胺酸或高葉酸飲食期間,對大白鼠血漿中SAM、SAH濃度與SAM/SAH比值的變化 ……………………... 49
四、 觀察給予高量甲硫胺酸或高葉酸飲食期間,對大白鼠血漿中葉酸濃度的變化 ………………………………………………………. 51
五、 給予高甲硫胺酸或高葉酸飲食對大白鼠熱中風生成時間和熱中風存活時間之影響 …………………………………………………. 53
六、 給予高甲硫胺酸或高葉酸飲食後,誘發熱中風生成過程中生理參數以及腦血流變化情形 …………………………………………. 54
七、 給予高甲硫胺酸或高葉酸飲食,對大白鼠熱中風後血漿中脂質過氧化程度之影響 …………………………………………………. 57
八、 給予高甲硫胺酸或高葉酸飲食,對大白鼠熱中風後各組織中脂質過氧化程度之影響 ………………………………………………. 59
九、 給予高甲硫胺酸或高葉酸飲食,對大白鼠熱中風後血漿中總抗氧化能力之影響 ……………………………………………………. 62
十、 給予高甲硫胺酸或高葉酸飲食,對大白鼠熱中風後DNA損傷之影響 ………………………………………………………………. 64
十一、 給予高甲硫胺酸或高葉酸飲食,對大白鼠熱中風後病理組織學之影響 ……………………………………………………………. 67
伍、 討論 ………………………………………………………………… 76
陸、 結論 ………………………………………………………………… 87
參考文獻 …………………………………………………………………….. 88
附錄 ………………………………………………………………………… 103
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