臺灣博碩士論文加值系統

中文摘要
Angiotensin II (Ang II)或其他的血管收縮素需要在氯離子的存在下，才能引起血管的收縮。Ang II引發血管平滑肌細胞內鈣離子的增加作用亦受到氯離子的調節，然而氯離子如何影響細胞內鈣離子的釋出其機制不是很清楚。我們以螢光染劑測定大鼠血管平滑肌細胞內鈣離子及活性氧的量，並進一步探討氯離子調節鈣離子釋出的作用是否經由細胞內活性氧的代謝而來。血管平滑肌細胞分別於不同氯離子濃度之生理鹽溶液中處理平衡後，移除細胞外之鈣離子，Ang II所引起的鈣離子釋出量，於氯離子120 mEq/L之生理鹽溶液約是20 mEq/L之1.5倍。此現象於Sprague Dawley (SD)大鼠及Spontaneously hypertensive rat (SHR)血管平滑肌細胞，皆可觀察測得。抗氧化劑vitamin E及N-acetylcysteine以及superoxide dismutase之抑制劑diethyldithiocarbamic acid可於高氯溶液中抑制Ang II引起的鈣離子釋出，顯示活性氧在Ang II引起的鈣離子訊息傳導過程中，可能具有相當的重要性。而Ang II的確可引發血管平滑肌細胞產生活性氧，其於高氯溶液中產生的量約為低氯溶液中之2.6倍。當外加H2O2於血管平滑肌細胞時，高氯溶液中以流式細胞儀所測得細胞內活性氧的量，在1至5分鐘時顯著的高於低氯溶液，但10分鐘後兩者的差異即消失。進一步實驗發現，在高氯溶液中，純化之catalase及血管平滑肌萃出物分解H2O2之速率僅為低氯溶液中之二分之一，顯示氯離子可能因藉由抑制catalase的活性，而增加細胞內活性氧的量。而SD大鼠其catalase的活性是SHR之3.5倍，因此氯離子對catalase之抑制作用可能於不同品種大鼠之血管平滑肌細胞所產生對氯離子的依賴有不同的重要性。我們推論，氯離子可能經由增加細胞內H2O2的穩定性而增強Ang II引發血管平滑肌鈣離子之釋出作用。

ABSTRACT
Cl- is essential for vasoconstriction induced by angiotensin II (Ang II) and other vasoconstrictors. Ang II-induced transient increase in intracellular [Ca2+] is also Cl--dependent. To further investigate whether Cl--dependent Ca2+ release from intracellular stores is mediated by altering metabolism of reactive oxygen species (ROS), intracellular Ca2+/ROS levels were measured using fluorescence dyes in cultured vascular smooth muscle cells (VSMC). After equilibration with solutions of variant [Cl-], VSMC was treated with Ang II in the absence of extracellular Ca2+ to induce Ca2+ release. The peak increase of released Ca2+ at 120 mEq/L of [Cl-] was 1.5 fold of that at 20 mEq/L of [Cl-] in VSMC from both Sprague Dawley (SD) and spontaneously hypertensive rat (SHR). Antioxidants, vitamin E and N-acetylcysteine, and a superoxide dismutase, diethyldithiocarbamic acid, significantly attenuated Ang II-induced Ca2+ release at high [Cl-] without altering those at low [Cl-]. In addition, Ang II-induced increase in intracellular ROS at high [Cl-] was 2.6 fold of that at low [Cl-]. Exogenous addition of H2O2 elicited a bigger increase in intracellular ROS with high vs. low [Cl-] one and five, but not ten min after addition of H2O2, suggesting that H2O2 degradation might be inhibited by Cl-. Further experiments demonstrated that the rate of H2O2 hydrolysis with high [Cl-] by both VSMC extract and purified catalase preparation, measured using spectrophotometer, was only half of that with low [Cl-], indicting that the activity of catalase was inhibited by Cl-. Catalase activity in VSMC from SD rat was 3 fold of that from SHR, suggesting that catalase may play a more important role in Cl--dependent Ca2+ release with SD rat than with SHR. In conclusion, Cl- may increase stability of H2O2 to enhance Ca2+ signaling in VSMC of rat.

目錄 (Contents)
指導教授推薦書
口試委員會審定書
授權書 iii
誌謝 iv
目錄 v
縮寫表 vi
中文摘要 viii
英文摘要 xi
緒論 1
材料與方法 10
結果 16
討論 23
圖表 31
參考文獻 46
附圖 57

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