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博碩士論文 etd-0514103-134312 詳細資訊
Title page for etd-0514103-134312
論文名稱
Title
汞對於腎小管細胞鈣傳訊及細胞毒殺之作用
Effect of mercury-induced Ca2+ increase and cytotoxicity in renal tubular cells
系所名稱
Department
畢業學年期
Year, semester
語文別
Language
學位類別
Degree
頁數
Number of pages
46
研究生
Author
指導教授
Advisor
召集委員
Convenor
口試委員
Advisory Committee
口試日期
Date of Exam
2003-05-02
繳交日期
Date of Submission
2003-05-14
關鍵字
Keywords
鈣離子、MDCK細胞、汞離子、腎小管細胞、fura-2、thapsigargin
mercury, Ca2+, MDCK cells, fura-2, renal, thapsigargin
統計
Statistics
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The thesis/dissertation has been browsed 5717 times, has been downloaded 21 times.
中文摘要
中文摘要

目前已知汞離子(Hg2+)對腎細胞會造成毒性,病變的腎臟常伴隨著鈣沈積,腎病常引發不正常之腎小管鈣傳訊。本研究是以Fura-2為鈣離子指示劑,探討氯化汞(HgCl2)對狗腎細胞(Madin Darby canine kidney cells)內鈣離子濃度變化之影響。當Hg2+濃度大於1 mM時,能增加細胞內鈣離子濃度,反應與Hg2+濃度呈正相關性,其有效半濃度為6 mM (EC50=6 mM)。而當細胞外鈣被移除後約可抑制67%的[Ca2+]i上升,此結果顯示Hg2+所引起之[Ca2+]i上升是來自於外鈣流入和鈣蓄池的內鈣釋出所致。在不含鈣溶液中,當加入內質網上鈣離子幫浦抑制劑(thapsigargin 1 mM)時,能引發[Ca2+]i上升效應,同時減弱Hg2+所誘導之[Ca2+]i上升效應,反之,若細胞先以Hg2+處理,會破壞thapsigargin所誘導產生[Ca2+]i上升之效應。而Hg2+所引發之[Ca2+]i上升並不受磷酯酶C (phospholipase C)的改變,而可能與蛋白質激酶C (protein kinase C) 的活化有關。另由分析細胞增生試驗的結果得知,發現1 mM的Hg2+不會改變細胞增生速率,但濃度10 mM即會殺死細胞。綜合此結果顯示Hg2+所誘發之內質網鈣蓄池鈣釋出,非經由磷酯酶C活性相關的作用機轉而是經由蛋白質激酶C的調控所致。且高濃度的Hg2+會對細胞有毒殺作用。

Abstract
Abstract
The effect of mercury (Hg2+), a known nephrotoxicant, on intracellular free Ca2+ levels ([Ca2+]i) in Madin Darby canine kidney (MDCK) cells was explored. [Ca2+]i was measured by using the Ca2+-sensitive dye fura-2. Hg2+ increased [Ca2+]i in a concentration-dependent manner with an EC50 of 6 mM. The Ca2+ signal comprised a gradual increase. Removal of extracellular Ca2+ decreased the Hg2+-induced [Ca2+]i increase by 67%, suggesting that the Ca2+ signal was due to both extracellular Ca2+ influx and store Ca2+ release. In Ca2+-free medium, the Hg2+-induced [Ca2+]i increase was nearly abolished by pretreatment with 1 mM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor), and conversely, pretreatment with Hg2+ abolished thapsigargin-induced Ca2+ increase. Hg2+-induced Ca2+ release was not altered by inhibition of phospholiase C but was potentiated by activation of protein kinase C. Overnight treatment with 1 mM Hg2+ did not alter cell proliferation rate, but 10 mM Hg2+ killed all cells. Collectively, this study shows that Hg2+ induced protein kinase C-regulated [Ca2+]i increases in renal tubular cells via releasing store Ca2+ from the endoplasmic reticulum in a manner independent of phospholipase C activity. Hg2+ also caused cytotoxicity at higher concentrations.

目次 Table of Contents
目  錄
誌 謝 1
中文摘要 2
英文摘要 3
目 錄 4
圖表目錄 6
專有名詞縮寫 7
第一章 緒論 8
第一節 研究動機與目的 8
第二節 汞的分類及生理毒性 10
一、元素汞 10
二、無機汞 10
三、有機汞 11
第三節 鈣與細胞生理 12
一、鈣:細胞中所伴演的角色 12
二、細胞鈣訊號的作用機轉 13
三、鈣與細胞的凋亡和毒殺 17
第二章 實驗的研究設計 21
第三章 材料與方法 22
第一節 儀器與材料 22
一、實驗儀器 22
二、實驗藥品 22
三、實驗用溶液 22
第二節 實驗方法 23
一、細胞內游離鈣離子濃度的測定 23
二、細胞活性分析 24
第三節 統計分析 24
第四章 結果 26
一、汞離子所誘發之鈣離子傳訊作用 26
二、外鈣移除時汞離子所誘發之鈣離子傳訊作用 26
三、汞離子對內質網所誘發之鈣離子傳訊作用 27
四、汞離子對磷脂酶C所誘發之鈣離子傳訊作用 27
五、汞離子對蛋白質激酶所誘發之鈣離子傳訊作用 28
六、汞離子對於腎小管細胞的毒殺作用 28

第五章 討論 37
第六章 結論 39
參考文獻 41
論文目錄 46

參考文獻 References
参考文獻
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