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研究生:吳育亨
研究生(外文):YU-HENG WU
論文名稱:金屬鋅在斑馬魚幼體中對於金屬鎘造成的毒性之保護作用
論文名稱(外文):Protection Effect of Zinc in Cadmium-Induce Toxicity Using Zebrafish Embryo as a Model
指導教授:蔡淦仁
學位類別:碩士
校院名稱:中山醫學大學
系所名稱:醫學檢驗暨生物技術學系碩士班
學門:醫藥衛生學門
學類:醫學技術及檢驗學類
論文種類:學術論文
論文出版年:2017
畢業學年度:105
語文別:中文
論文頁數:42
中文關鍵詞:鎘離子鋅離子ZIP10鋅離子通道蛋白活性氧化物
外文關鍵詞:CadmiumzincZIP10ZnTROS
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鎘金屬是一種對生物有嚴重危害性的金屬,生活在受鎘污染水體
的魚類容易出現鰓功能衰結和神經失能的症狀。另一方面,金屬鋅是 一種生物必須的金屬元素,它參與許多生理功能,例如:發育、代謝、 信息傳遞、神經系統和免疫系統等等。目前有研究發現,鎘離子的毒 性可能與和鋅離子競爭有關,而額外的鋅離子可以減緩鎘的毒性。為 理解鎘離子在斑馬魚胚胎體內的影響,我們觀察不同濃度的金屬離子 對胚胎死亡率的影響,以及使用DCFH-DA染色觀察魚體內活性氧化 物,再以即時定量聚合酶連鎖反應觀察胚胎體內mRNA表現量。就結 果來看,鎘離子的毒性以及額外鋅離子的保護作用確實是可以被證實 的,但在胚胎對鋅離子耐受最高濃度以下,加入更多的鋅離子,在存 活率上沒有明顯效果;從活性氧化物的角度來看,鎘離子確實會造成 氧化壓力,但是這並不致死的原因;最後在mRNA的表現量變化看 來,並沒有明確的證據證實,鎘離子在胚胎時期能對ZnT1, ZIP10基因 有劇烈的影響,因此結論出:鎘離子確實能對斑馬魚胚胎造成致死的 影響,但其主要原因並不是在影響氧化壓力以及通道蛋白ZnT1, ZIP10 的mRNA表現量,另外,鋅離子也確實有緩和鎘毒性的作用。
Cadmium is an omnipresent metal ion with strong detrimental effects to all living creatures. Fish live in water with cadmium contamination will curse grill and neural disfunction. On the other hand, zinc is a trace element of great importance for all organisms, involved in many metabolic processes and well known as a cadmium antagonist. Among those previous studies, supplement of additional Zinc ion can induce cadmium-induce toxicity in adult female zebrafish. In this study, we determine the effect of zinc ion in cadmium-induced toxicity environment, by a serial of examinations using different concentrations of zinc to protect cadmium toxicity at different concentrations and different time intervals. We have demonstrated that zebrafish embryo co-treated with zinc, even at a lower concentration than cadmium, could efficiently protect the cadmium toxicity by measuring the death rate of the fish embryos. To further determine the mechanism for zinc protection in cadmium-induced toxicity, the embryo ROS levels are measured with DCFH-DA. Furthermore, real-time PCR examinations are performed to quantitate the RNA expression of SOD3, ZnT1 and Zip10, to determine the gene involvements for zinc induced protection to cadmium-induced toxicity in zebrafish embryo. These result shows that cadmium has no significant effect on intracellular ROS, and the mRNA expression of SOD3, ZnT1 and Zip10. In conclusion, there is no doubt with cadmium-induced toxicity can cause zebrafish death, but the mechanism of cadmium-induced toxicity in zebrafish embryo is still unknown. We may only sure about that oxidative stress is not the mean mechanism curse zebrafish embryo death.
目錄........................................................................... i
誌謝........................................................................... iii
中文摘要..................................................................... iv
英文摘要..................................................................... v
第一章、緒論............................................................... 1
第一節、斑馬魚 .......................................................... 1
第二節、鎘 (Cadmium) ............................................. .2
第三節、鋅(Zinc)...................................................... ..3
第四節、鋅運輸蛋白 ................................................... 4
第五節、超氧化物歧化酶 (Superoxid Dismutase, SODs) ...... 6
第六節、研究動機 .......................................................6
第二章實驗材料與方法 ............................................... 8
第一節、斑馬魚飼養 .................................................. 8
第二節、豐年蝦卵的養殖 ............................................ 8
第三節、魚卵收集 ................................................... 8
第四節、重金屬環境模擬 .......................................... 9
第五節、細胞內活性氧化物染色 ................................. 10
第六節、RNA純化萃取 ............................................. 10
第七節、反轉錄作用 ................................................ 11
第八節、聚合酶連鎖反應.......................................... 12
第九節、膠體電泳........................................................... 12
第十節、即時定量聚合酶鏈鎖反應 (Real-Time PCR)......... 13
第三章、實驗結果......................................................... 15
第四章、討論............................................................... 19
第五章、參考文獻......................................................... 24
第六章、圖表說明......................................................... 31
第七章、附錄............................................................... 41
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