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標題: | 吳郭魚仔魚鎘致毒的可能機制 Mechanisms Involved in Cadmium Toxification on Tilapia Larva ( Oreochromis mossambicus ) |
作者: | 餘明俊 |
出版年 : | 1995 |
學位: | 碩士 |
摘要: | 鎘是常見於礦業、冶鍊、電鍍、塑膠、電池及廢五金的污染性重金屬,離子形式的鎘對於水環境與生物有著嚴重的威脅,更直接或間接影響到人類的安全。本論文擬就鎘對生物的毒性(包括仔魚鎘累積量、死亡率、全長及鈣含量的變化)以及生物避免鎘中毒的保護機制(金屬硫蛋白的表現量)兩方面來探討鎘造成吳郭魚仔魚中毒的機制。
預備實驗中將剛孵化的吳郭魚仔魚(以下簡稱仔魚),浸泡在鎘濃度分別為 0 (控制組)、 50 或 150 μg / l 的水(鈣濃度0.lmM)中進行毒性實驗 ,發現 150 μg / l鎘處理 96 小時後即對仔魚有致死的作用。然而 96 小時仔魚體內的鎘累積量, 150 μg / l鎘處理組低於 50μg / l 鎘處理組,顯示鎘的累積量與仔魚的死亡之間沒有正相關,意味著仔魚有避免鎘中毒的保護機制存在。仔魚成長過程中,由於表面積或通透性的增加,造成仔魚體內的鈣與鎘離子都會不斷地增加,然而 150μg / l鎘處理 96 小時後,仔魚體內的鈣含量與鎘累積量都低於 50μg / l鎘處理組,意味著高濃度的鎘會造成仔魚成長的抑制,最後可能導致仔魚的死亡。 為了瞭解金屬硫蛋白在保護吳郭魚仔魚避免鎘中毒的功能,運用分子生物的技術從吳郭魚成魚的肝臟中,選殖出吳郭魚的金屬硫蛋白基因,再藉由北方雜合反應( northern hybridization )來偵測仔魚在不同鎘濃度處理後,其金屬硫蛋白基因表現的情形 。 其結果顯示,在 96 小時實驗期間,仔魚金屬硫蛋白 mRNA 的表現量,會隨鎘的浸泡濃度增加而增加,意味著金屬硫蛋白可能提供了,鎘處理後的仔魚避免鎘中毒的保護作用,至於金屬硫蛋白蛋白質表現量的部份仍待進一步的探討。目前已純化出一個吳郭魚的鎘的結合蛋白待定性後、製備抗體,以便瞭解金屬硫蛋白在保護吳郭魚仔魚避免鎘中毒的功能。 為了釐清仔魚的鎘累積量、鈣含量與成長的抑制三者,在鎘所造成的仔魚死亡中的角色,再以高鈣水(鈣濃度 2.0mM)與低鈣水(鈣濃度 0.1 mM ) 分別進行仔魚鎘的毒性實驗。與預備實驗結果相同,在低鈣、高鎘濃度的水中會造成仔魚成長的抑制與死亡。而在高鈣水中,除 300μg / l 處理組外,不論控制組50 、100 或 150μg / l 處理組都維持了相同的鈣含量與全長,而且死亡率也明顯降低,顯示定量的鈣吸收與仔魚的正常成長甚至存活有著明顯的正相關性。而且由於正常成長的結果,使得仔魚體內的鎘累積量隨鎘的處理濃度增加而增加,顯示高鈣的水不會阻礙鎘進入仔魚體內,也再次驗證鎘累積量的高與低不與仔魚的死亡成正相關。 總而言之,對於吳郭魚仔魚而言,正常成長的進行是很重要的,而成長中鈣的吸收是相當重要的生理需求。而高濃度的鎘造成吳郭魚仔魚鈣吸收量的不足,使其成長受到了抑制,可能導致其死亡。至於鎘與鈣之間究竟是如何的交互作用,以及金屬硫蛋白的在保護吳郭魚仔魚避免鎘中毒的功用如何則值得深入的探討。 The main objective of the present study is to elucidate the possible mechanisms involved in cadmium (Cd) toxification in tilapia larvae. The effects of environmental Cd and calcium (Ca) on the mRNA expression of metallothionein (MT), Cd accumulation, calcium content and growth in tilapia larvae were examined. In preliminary studies, high Cd concentration had lethal effect on tilapia larvae. The contents of Cd at 96 h was higher in 50 than 150μg/l Cd treatments, which suggests that failure in self- protective function may lead to death. Both the contents of Cd and Ca of tilapia larvae in each treatment increased as larval growth. The increasing ion contents of larvae may be due to the increasing surface area and permeability to ions when larvae grew up. However, the contents of both Ca and Cd of larvae in 100μg/l were less than those in 50μg/l Cd treatment. Thus, high cadmium concentration may inhibit the Ca uptake of larvae, which is necessary for the normal growth of larvae. To study the protective function of MT in preventing tilapia larvae from Cd toxicity, the MT cDNA sequence was cloned and the expression of the larval MT mRNA in various Cd concentration treatments were analyzed by northern blot hybridization. The expression of MT mRNA increased as the increasing Cd concentrations, which implied that the MT contributed to tilapia larvae the protective ability in preventing from Cd toxicity. A Cd-binding protein had been purified from adult tilapia liver, however the sequence and characteristics remained to be confirmed further. Because Ca uptake of larvae may be inhibited by Cd, the other experiments were conducted in high Ca environments ([Ca++]=2.0mM). Compared to the larvae in normal tap water ([Ca++1=0.lmM), the mortality of larvae in high—Ca water was much lower, even in 300μg/l Cd treatment. Moreover, in high—Ca water, the Ca uptake and the total length of larvae were not inhibited by high Cd concentrations. These data suggested that high Ca concentration supplied a normal Ca uptake and growth of tilapia larvae, which may prevented the larvae from death. Our results indicated that Ca uptake plays a important role in normal growth of tilapia larvae. The mechanism of Cd toxicification in larvae may be due to that Cd competes with Ca for the entrance of Ca, which may lead to inhibition of growth and death. The interaction between Ca and Cd and the function of MT remains to be studied further. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/76175 |
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顯示於系所單位: | 漁業科學研究所 |
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