氟化钠对中华大蟾蜍胚胎发育的影响
发布时间：2018-07-23 19:37
【摘要】：氟是存在于地球上最丰富的化学元素之一,由于氟化学性质活泼,在自然界中几乎不以氟单体的形式存在,而是与其它元素结合形成无机和有机氟化物。氟是动物及人体发育过程中必需的微量元素之一。适量摄入氟有益于龋齿的预防和骨骼发育,然而过量的氟摄取会对动物及人体产生毒性效应,造成氟斑牙、氟骨症外,还会引起机体神经系统、内分泌腺等非骨相组织器官的损伤。近年来,随着现代工业的发展,人类生产活动造成大量氟化物的产生,如金属冶炼、有机肥料的加工、含氟“三废”的排放,导致水环境中氟含量持续升高。中华大蟾蜍作为我国两栖类最具代表性的物种,其胚胎发育和幼体发育均在水域中进行,水域氟污染会直接影响其发育。在本研究中,首先研究了氟化钠对中华大蟾蜍胚胎的急性毒性效应,以及亚致死浓度下氟化钠对中华大蟾蜍胚胎生长发育的影响,其次对胚胎的体重和体长进行了测量和分期,和电镜观察,最后采用RT-PCR和Real-time PCR方法,对不同分期中华大蟾蜍胚胎样本中的Dio2, Dio3, TRα和TRβ mRNA水平进行定量研究。本文的主要研究结果如下：1.采用半静态生物毒性试验方法对中华大蟾蜍G11期-G22期(本研究依照Gosner (1960)对蛙类胚胎的分期方法来确定蛙胚的不同发育时期)的胚胎进行氟化钠的高浓度(200-500 mg/L)毒性试验,统计了氟化钠对中华大蟾蜍胚胎的致畸个数和致死个数,并计算了半数致畸浓度(EC50)和半数致死浓度(LC50)。结果显示中华大蟾蜍胚胎暴露在氟化钠中216小时的ECso和LC50分别为416.37mg/L和561.64 m/L,并且暴露在500 mg/L的畸形率和致死率分别是66.25%和47.50%。2.氟化钠可以造成中华大蟾蜍胚胎畸形,主要包括卵黄囊水肿、体轴弯曲和组织畸形增生。扫描电镜进一步了解氟化钠对胚胎的致畸效应,结果显示氟化钠会导致胚胎尾部表皮细胞畸形增生和变性。3.采用半静态生物毒性试验方法对中华大蟾蜍G7期-G22期的胚胎进行氟化钠的亚致死浓度(1-150 mg/L)毒性试验,通过测定体重和体长等相关形态学数据,推测氟化钠对中华大蟾蜍胚胎生长发育的影响。结果显示,氟暴露对体重和体长均呈现出了剂量相关性的影响。1 mg/L氟暴露对胚胎的体重和体长没有显著性影响(p0.05),10-150 mg/L氟暴露会造成胚胎的体重和体长的显著降低(p0.05)。4.在亚致死浓度毒性试验中,记录暴露在不同浓度氟化钠,中华大蟾蜍胚胎发育到G22期的累计发育时间；记录6d-9d,胚胎发育的平均时期。采用两种不同的统计方法,分析氟化钠对中华大蟾蜍胚胎生长发育的影响。累计发育时间结果的统计分析显示,在氟化钠处理组,胚胎的发育延迟显著(p0.05)；胚胎发育分期的结果统计分析表明,1 mg/L氟暴露对胚胎的发育进程无显著性影响(p0.05),10-150 mg/L氟暴露会显著延迟胚胎发育(p0.05)5.采用实时定量Real-time PCR方法,检测氟化钠在中华大蟾蜍胚胎发育过程中对脱碘酶基因Dio2,Dio3以及甲状腺激素受体基因TRa和TRβ表达水平的影响。结果显示：在G11期(原肠胚期),氟化钠处理组中Dio3和TRβ基因的表达量显著性增加p0.05),而Dio2和TRa基因的表达水平与对照组相比,无显著性差异(p0.05)；在G16期(神经胚期),经氟化钠暴露的实验组,其Dio2,Dio3,TRα和TRβ基因的表达水平与对照组相比均有显著性升高(p0.05)；在G18期(肌肉效应期),经100 mg/L氟化钠处理的实验组中,Dio2和TRa基因的表达量显著性降低(p0.05),而Dio3和TRβ基因的表达量在100和150 mg/L氟化钠暴露组中显著性增加(p0.05)；在G22期(尾鳍循环期),经氟化钠暴露的实验组,其Dio2,Dio3,TRα和TRβ基因的表达水平与对照组相比均有显著性降低(p0.05)。推测氟化钠可能通过干扰脱碘酶基因Dio2和Dio3,以及甲状腺激素受体基因TRa和TRβ的正常表达,从而影响中华大蟾蜍胚胎的发育。本研究首次从传统毒理学和现代分子生物学水平较系统的研究了氟化钠对中华大蟾蜍胚胎发育的影响,取得了较完整的研究数据,揭示了氟化钠对中华大蟾蜍胚胎的中毒机理,为两栖类毒理学研究及其栖息地的保护研究提供了参考。
[Abstract]:Fluorine is one of the most abundant chemical elements in the earth. Because of the active chemical properties of fluorine, it is almost not in the form of fluorine monomers in nature, but is combined with other elements to form inorganic and organic fluorides. Fluorine is one of the essential trace elements in the development of animals and human beings. Bone development, however, excessive fluoride intake can produce toxic effects on animals and human body, causing dental fluorosis, fluorosis, and the injury of non osteogenic organs such as the nervous system and endocrine gland. In recent years, with the development of modern industry, human production activities caused a large number of fluoride production, such as metal smelting, organic fertilizer. As the most representative species of amphibians in China, Bufo bufo gargarizans, as the most representative species of amphibians in China, are the most representative species in the water environment. The development of the fluorine pollution in the waters will directly affect its development. In this study, the first study of sodium fluoride to Bufo Bufo bufo gargarizans embryos The effects of acute toxicity and sublethal concentration of sodium fluoride on the growth and development of Bufo bufo gargarizans were observed. Secondly, the weight and body length of the embryos were measured and staging and observed by electron microscopy. Finally, the RT-PCR and Real-time PCR methods were used to improve the levels of Dio2, Dio3, TR a and TR beta mRNA in different stages of the embryo samples of Bufo bufo gargarizans. The main results of this study are as follows: 1. the high concentration (200-500 mg/L) toxicity test of sodium fluoride (200-500 mg/L) in the G11 phase -G22 phase of Bufo bufo gargarizans (this study was determined by Gosner (1960) stage method for frog embryo at different developmental stages) was carried out, and the fluorination was statistically analyzed. The number of teratogenicity and lethal number of sodium to Bufo gargarizans embryos were calculated, and half of the teratogenic concentration (EC50) and half lethal concentration (LC50) were calculated. The results showed that the 216 hours of ECso and LC50 were 416.37mg/L and 561.64 m/L respectively in the embryo exposure of Bufo bufo gargarizans, respectively, and the malformation rate and mortality rate of 500 mg/L were 66.25% and 47, respectively. .50%.2. sodium fluoride can cause fetal malformation of Bufo bufo gargarizans, mainly including ovoid sac edema, body axis bending and tissue malformation. The scanning electron microscope is used to further understand the teratogenic effect of sodium fluoride on embryos. The results show that sodium fluoride can lead to abnormal proliferation and denaturation of embryonic tail epidermal cells in.3. by semi static biotoxicity test method. The sublethal concentration (1-150 mg/L) of sodium fluoride (1-150 mg/L) toxicity test of sodium fluoride in the -G22 phase of Bufo bufo gargarizans was conducted. The effects of sodium fluoride on the growth and development of Bufo bufo gargarizans were estimated by measuring the morphological data of body weight and body length. The results showed that fluorine exposure had a dose-dependent effect on the.1 mg/L fluorine storm. Exposure had no significant influence on the weight and body length of the embryo (P0.05). 10-150 mg/L fluorine exposure could cause a significant decrease in the body weight and body length of the embryo (P0.05).4. in the sublethal concentration toxicity test, to record the cumulative time of exposure to different concentrations of sodium fluoride, the embryo development of Bufo bufo gargarizans to the G22 period, and record 6d-9d, the flat development of embryo. Two different statistical methods were used to analyze the effects of sodium fluoride on the growth and development of Bufo bufo gargarizans. The statistical analysis of the cumulative development time results showed that the development delay of embryos was significant (P0.05) in the sodium fluoride treatment group, and the results of embryo development staging showed that 1 mg/L fluorine exposure had no effect on the development of embryos. Significant effect (P0.05), 10-150 mg/L fluorine exposure significantly delayed embryo development (P0.05) 5. using real-time quantitative Real-time PCR method to detect the effect of sodium fluoride on the expression level of deiodonase gene Dio2, Dio3, and thyroid hormone receptor gene TRa and TR beta during the development of Bufo bufo gargarizans. The results showed that in the G11 phase (the primary gut embryo) The expression of Dio3 and TR beta gene in the sodium fluoride treatment group was significantly increased by P0.05), while the expression level of Dio2 and TRa genes had no significant difference compared with the control group (P0.05). In the G16 phase (neural embryo stage), the expression level of Dio2, Dio3, TR A and TR beta gene in the experimental group with sodium fluoride exposure was significantly higher than that of the control group (P). 0.05); in the G18 phase (muscle effect period), the expression of Dio2 and TRa genes decreased significantly in the experimental group treated with 100 mg/L sodium fluoride (P0.05), while the expression of Dio3 and TR beta genes increased significantly in the 100 and 150 mg/L sodium fluoride exposure group (P0.05); in the G22 phase (caudal fin cycle), the experimental group of sodium fluoride exposure was Dio2, Dio3, and alpha. The expression level of the TR beta gene was significantly lower than that in the control group (P0.05). It is speculated that the sodium fluoride may affect the normal expression of the deiodinase gene Dio2 and Dio3, as well as the normal expression of the thyroid hormone receptor gene TRa and TR beta, thus affecting the development of the embryo of Bufo bufo gargarizans. The effect of sodium fluoride on the embryo development of Bufo bufo gargarizans was studied in a relatively systematic way. The complete research data were obtained, and the poisoning mechanism of sodium fluoride on Bufo bufo gargarizans embryos was revealed, which provided a reference for the study of amphibian toxicology and the protection of its habitat.
【学位授予单位】：陕西师范大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：X171.5;Q954.4
[Abstract]:Fluorine is one of the most abundant chemical elements in the earth. Because of the active chemical properties of fluorine, it is almost not in the form of fluorine monomers in nature, but is combined with other elements to form inorganic and organic fluorides. Fluorine is one of the essential trace elements in the development of animals and human beings. Bone development, however, excessive fluoride intake can produce toxic effects on animals and human body, causing dental fluorosis, fluorosis, and the injury of non osteogenic organs such as the nervous system and endocrine gland. In recent years, with the development of modern industry, human production activities caused a large number of fluoride production, such as metal smelting, organic fertilizer. As the most representative species of amphibians in China, Bufo bufo gargarizans, as the most representative species of amphibians in China, are the most representative species in the water environment. The development of the fluorine pollution in the waters will directly affect its development. In this study, the first study of sodium fluoride to Bufo Bufo bufo gargarizans embryos The effects of acute toxicity and sublethal concentration of sodium fluoride on the growth and development of Bufo bufo gargarizans were observed. Secondly, the weight and body length of the embryos were measured and staging and observed by electron microscopy. Finally, the RT-PCR and Real-time PCR methods were used to improve the levels of Dio2, Dio3, TR a and TR beta mRNA in different stages of the embryo samples of Bufo bufo gargarizans. The main results of this study are as follows: 1. the high concentration (200-500 mg/L) toxicity test of sodium fluoride (200-500 mg/L) in the G11 phase -G22 phase of Bufo bufo gargarizans (this study was determined by Gosner (1960) stage method for frog embryo at different developmental stages) was carried out, and the fluorination was statistically analyzed. The number of teratogenicity and lethal number of sodium to Bufo gargarizans embryos were calculated, and half of the teratogenic concentration (EC50) and half lethal concentration (LC50) were calculated. The results showed that the 216 hours of ECso and LC50 were 416.37mg/L and 561.64 m/L respectively in the embryo exposure of Bufo bufo gargarizans, respectively, and the malformation rate and mortality rate of 500 mg/L were 66.25% and 47, respectively. .50%.2. sodium fluoride can cause fetal malformation of Bufo bufo gargarizans, mainly including ovoid sac edema, body axis bending and tissue malformation. The scanning electron microscope is used to further understand the teratogenic effect of sodium fluoride on embryos. The results show that sodium fluoride can lead to abnormal proliferation and denaturation of embryonic tail epidermal cells in.3. by semi static biotoxicity test method. The sublethal concentration (1-150 mg/L) of sodium fluoride (1-150 mg/L) toxicity test of sodium fluoride in the -G22 phase of Bufo bufo gargarizans was conducted. The effects of sodium fluoride on the growth and development of Bufo bufo gargarizans were estimated by measuring the morphological data of body weight and body length. The results showed that fluorine exposure had a dose-dependent effect on the.1 mg/L fluorine storm. Exposure had no significant influence on the weight and body length of the embryo (P0.05). 10-150 mg/L fluorine exposure could cause a significant decrease in the body weight and body length of the embryo (P0.05).4. in the sublethal concentration toxicity test, to record the cumulative time of exposure to different concentrations of sodium fluoride, the embryo development of Bufo bufo gargarizans to the G22 period, and record 6d-9d, the flat development of embryo. Two different statistical methods were used to analyze the effects of sodium fluoride on the growth and development of Bufo bufo gargarizans. The statistical analysis of the cumulative development time results showed that the development delay of embryos was significant (P0.05) in the sodium fluoride treatment group, and the results of embryo development staging showed that 1 mg/L fluorine exposure had no effect on the development of embryos. Significant effect (P0.05), 10-150 mg/L fluorine exposure significantly delayed embryo development (P0.05) 5. using real-time quantitative Real-time PCR method to detect the effect of sodium fluoride on the expression level of deiodonase gene Dio2, Dio3, and thyroid hormone receptor gene TRa and TR beta during the development of Bufo bufo gargarizans. The results showed that in the G11 phase (the primary gut embryo) The expression of Dio3 and TR beta gene in the sodium fluoride treatment group was significantly increased by P0.05), while the expression level of Dio2 and TRa genes had no significant difference compared with the control group (P0.05). In the G16 phase (neural embryo stage), the expression level of Dio2, Dio3, TR A and TR beta gene in the experimental group with sodium fluoride exposure was significantly higher than that of the control group (P). 0.05); in the G18 phase (muscle effect period), the expression of Dio2 and TRa genes decreased significantly in the experimental group treated with 100 mg/L sodium fluoride (P0.05), while the expression of Dio3 and TR beta genes increased significantly in the 100 and 150 mg/L sodium fluoride exposure group (P0.05); in the G22 phase (caudal fin cycle), the experimental group of sodium fluoride exposure was Dio2, Dio3, and alpha. The expression level of the TR beta gene was significantly lower than that in the control group (P0.05). It is speculated that the sodium fluoride may affect the normal expression of the deiodinase gene Dio2 and Dio3, as well as the normal expression of the thyroid hormone receptor gene TRa and TR beta, thus affecting the development of the embryo of Bufo bufo gargarizans. The effect of sodium fluoride on the embryo development of Bufo bufo gargarizans was studied in a relatively systematic way. The complete research data were obtained, and the poisoning mechanism of sodium fluoride on Bufo bufo gargarizans embryos was revealed, which provided a reference for the study of amphibian toxicology and the protection of its habitat.
【学位授予单位】：陕西师范大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：X171.5;Q954.4
【相似文献】
相关期刊论文 前10条
1 王春元;汪安琦;;金鱼胚胎发育不同时期辐射敏感性的研究[J];科学通报;1963年04期
2 吕柳新;余小玲;叶明志;肖振通;;荔枝胚胎发育机制的探讨[J];福建农学院学报;1989年02期
3 朱勇飞,张天宝;热休克蛋白与胚胎发育的关系[J];国外医学(卫生学分册);2004年05期
4 袁静;任育军;赵洁;;水稻胚胎发育过程相关基因的鉴定(简报)(英文)[J];分子细胞生物学报;2008年03期
5 何登菊;姚俊杰;赵云龙;姜海波;;瓯江彩鲤胚胎发育过程中脂、蛋白及碳水化合物水平的变化[J];动物学杂志;2011年02期
6 杨国辉;杨自忠;李孟然;王志强;宋宇;;红瘰疣螈胚胎发育的初步观察[J];生物学通报;2011年08期
7 曾弥白;胚胎发育中各胚层的相互作用[J];动物学杂志;1958年01期
8 钦俊德,郭郛,翟US慧,沙槎云;蝗卵的研究Ⅳ.浸水对于蝗卵胚胎发育和死亡的影响[J];昆虫学报;1959年04期
9 薛社普;;人的生殖及胚胎发育[J];生物学通报;1962年01期
10 秦鹏春;吕连升;杨山;刘凤仪;李齐华;;北京，
本文编号：2140437
本文链接：https://www.wllwen.com/kejilunwen/huanjinggongchenglunwen/2140437.html
最近更新
教材专著
