內(nèi)分泌干擾物雙酚A對(duì)秀麗隱桿線蟲(chóng)的生態(tài)毒性效應(yīng)及其作用機(jī)制研究
發(fā)布時(shí)間：2018-11-08 15:57
【摘要】：雙酚A(BPA)是世界公認(rèn)的環(huán)境內(nèi)分泌干擾物,一直是毒理學(xué)研究中的明星化合物。BPA的世界年需求量超過(guò)400萬(wàn)噸/年,環(huán)境中的污染也日趨嚴(yán)重,在土壤、水、大氣和沉積物中都發(fā)現(xiàn)了BPA的蹤跡,甚至在人體組織和體液中也檢測(cè)到了BPA的存在。體內(nèi)和體外實(shí)驗(yàn)已經(jīng)證明BPA暴露能夠引起內(nèi)分泌干擾效應(yīng)、神經(jīng)毒性、生殖毒性、免疫毒性和致癌致畸性等。人群流行病學(xué)研究也發(fā)現(xiàn),糖尿病、肥胖、神經(jīng)行為障礙、肝功能異常、復(fù)發(fā)性流產(chǎn)、發(fā)育性疾病都與BPA暴露有關(guān)。目前關(guān)于BPA暴露對(duì)無(wú)脊椎動(dòng)物的毒性報(bào)道較少。作為一種從土壤中分離出來(lái)的代表性的線蟲(chóng)種類,秀麗隱桿線蟲(chóng)是環(huán)境暴露評(píng)價(jià)以及生態(tài)毒理學(xué)研究中最常用的模式生物之一。本研究選取秀麗隱桿線蟲(chóng)作為受試生物,主要從生理、生化、分子水平探究環(huán)境相關(guān)濃度下BPA急性與亞急性暴露、慢性暴露和多代連續(xù)暴露對(duì)秀麗隱桿線蟲(chóng)的生態(tài)毒性效應(yīng)；此外,本研究還考察了BPA的替代品雙酚S(BPS)對(duì)秀麗隱桿線蟲(chóng)的毒性影響,并與BPA的毒性進(jìn)行對(duì)比,分析BPA替代品的生態(tài)風(fēng)險(xiǎn)性。本研究主要的研究結(jié)果如下：(1)BPA急性與亞急性暴露能夠顯著抑制秀麗隱桿線蟲(chóng)的體長(zhǎng)、運(yùn)動(dòng)行為水平以及子代數(shù)目,顯著提高細(xì)胞凋亡水平以及脅迫相關(guān)基因的表達(dá)量。急性暴露后,秀麗隱桿線蟲(chóng)的頭部擺動(dòng)頻率是最敏感的生理指標(biāo),暴露濃度在0.01μM即可對(duì)秀麗隱桿線蟲(chóng)產(chǎn)生顯著的負(fù)面效應(yīng),而亞急性暴露后,體長(zhǎng)和頭部擺動(dòng)頻率相比其他生理指標(biāo)更加敏感,同樣在暴露濃度為0.01μM時(shí)能夠?qū)π沱愲[桿線蟲(chóng)產(chǎn)生顯著的抑制作用。BPA急性和亞急性暴露對(duì)線蟲(chóng)體內(nèi)脂褐素累積和ROS水平?jīng)]有產(chǎn)生顯著影響,但當(dāng)暴露濃度達(dá)到0.1μM時(shí)能夠顯著提高線蟲(chóng)體內(nèi)細(xì)胞凋亡水平,表明實(shí)驗(yàn)濃度下BPA暴露對(duì)秀麗隱桿線蟲(chóng)產(chǎn)生的毒性效應(yīng)并不是通過(guò)氧化損傷途徑,而很可能與細(xì)胞凋亡水平升高有關(guān)。BPA亞急性暴露能夠?qū)е滦沱愲[桿線蟲(chóng)體內(nèi)絕大部分脅迫相關(guān)基因表達(dá)量的升高,即使在最低暴露濃度0.001 μM, BPA暴露也能夠引起絕大多數(shù)脅迫相關(guān)基因表達(dá)量的顯著提升。(2)BPA慢性暴露能夠顯著抑制秀麗隱桿線蟲(chóng)的體長(zhǎng)、運(yùn)動(dòng)行為水平、壽命以及種群數(shù)目,顯著提高線蟲(chóng)體內(nèi)細(xì)胞凋亡水平、脅迫相關(guān)基因表達(dá)以及轉(zhuǎn)基因品系TJ375的熒光蛋白表達(dá)量。慢性暴露后,頭部擺動(dòng)頻率是最敏感的生理指標(biāo),暴露濃度在0.001μM即可對(duì)秀麗隱桿線蟲(chóng)產(chǎn)生顯著的負(fù)面效應(yīng)。BPA慢性暴露對(duì)線蟲(chóng)體內(nèi)脂褐素累積同樣沒(méi)有顯著影響,但能夠在暴露濃度達(dá)到0.01μM以上時(shí),顯著地提高秀麗隱桿線蟲(chóng)體內(nèi)的細(xì)胞凋亡水平。在實(shí)驗(yàn)濃度范圍內(nèi),BPA慢性暴露能夠明顯地提高線蟲(chóng)體內(nèi)絕大部分脅迫相關(guān)基因的表達(dá)量。與亞急性暴露的結(jié)果相比,同一暴毒濃度下,BPA慢性暴露能夠引起脅迫相關(guān)基因的mRNA水平更為明顯的提高。當(dāng)BPA暴露濃度達(dá)到0.1μM時(shí),秀麗隱桿線蟲(chóng)的種群數(shù)目顯著減小。轉(zhuǎn)基因品系TJ375只能夠檢測(cè)環(huán)境濃度達(dá)到1μM時(shí)的BPA毒性,與其它測(cè)試指標(biāo)相比不夠靈敏。(3)BPA多代連續(xù)暴露能夠在暴露濃度達(dá)到0.01μM以上時(shí)對(duì)秀麗隱桿線蟲(chóng)的生理指標(biāo)產(chǎn)生顯著的毒性影響。在較低暴露濃度時(shí),子代線蟲(chóng)會(huì)出現(xiàn)毒性逐代修復(fù)或者惡化的情況,造成秀麗隱桿線蟲(chóng)生理水平上的世代毒性差異,而在最高暴露濃度10μM時(shí),秀麗隱桿線蟲(chóng)G1代和G4代在毒性效應(yīng)上沒(méi)有顯著性的差異。在秀麗隱桿線蟲(chóng)G1代和G4代中,與高濃度暴露組相比,低濃度(≤0.01 μM)暴露組能夠引起體內(nèi)脅迫相關(guān)基因表達(dá)量更為明顯的提高；與G4代相比,秀麗隱桿線蟲(chóng)的G1代體內(nèi)脅迫相關(guān)基因的表達(dá)量增加更為顯著。(4)BPS亞急性暴露后,秀麗隱桿線蟲(chóng)的頭部擺動(dòng)頻率是最敏感的生理指標(biāo),暴露濃度在1 μM以上才能夠?qū)π沱愲[桿線蟲(chóng)產(chǎn)生顯著的負(fù)面效應(yīng),而其他生理指標(biāo)只有在最高濃度暴露組才會(huì)受到顯著的負(fù)面影響。亞急性暴露后,秀麗隱桿線蟲(chóng)體內(nèi)脂褐素的累積沒(méi)有受到顯著影響,細(xì)胞凋亡水平也只有在暴露濃度為1μM時(shí)顯著提高。BPS亞急性暴露能夠引起秀麗隱桿線蟲(chóng)體內(nèi)絕大部分基因表達(dá)量的小幅度上升。綜上所述,低濃度的BPA暴露能夠?qū)π沱愲[桿線蟲(chóng)產(chǎn)生發(fā)育毒性、神經(jīng)行為毒性以及生殖毒性。BPA暴露對(duì)秀麗隱桿線蟲(chóng)生理水平的毒性影響很可能與體內(nèi)細(xì)胞凋亡水平提高有關(guān)；BPA暴露能夠顯著提高秀麗隱桿線蟲(chóng)體內(nèi)脅迫相關(guān)基因的表達(dá),破壞機(jī)體的穩(wěn)態(tài)；轉(zhuǎn)基因品系TJ375對(duì)于BPA暴露不敏感,不能有效監(jiān)測(cè)BPA的毒性；與BPA毒性相比,BPS的毒性明顯較弱。
[Abstract]:Bisphenol A (BPA) is a world-recognized environmental endocrine disruptor and has been a star compound in toxicology studies. The world's annual demand for BPA is more than 4 million tons/ year, and the pollution in the environment is becoming more and more serious. The trace of BPA is found in soil, water, atmosphere and sediment, and BPA is also detected in human tissue and body fluids. In vivo and in vitro experiments have shown that the exposure of BPA can cause endocrine disruption, neurotoxicity, reproductive toxicity, immunotoxicity, and carcinogenicity. The epidemiological study of the population also found that diabetes, obesity, neurobehavioral disorders, liver function abnormalities, recurrent spontaneous abortion, and developmental disorders were associated with exposure to BPA. At present, the toxicity of BPA exposure to invertebrates is less. As a representative nematode species isolated from the soil, the Caenorhabditis elegans is one of the most commonly used modes in environmental exposure assessment and ecological toxicology studies. The study was conducted to study the ecological toxicity of Caenorhabditis elegans as the tested organism, mainly from the physiological, biochemical and molecular level to the acute and subacute exposure of BPA, the chronic exposure and the continuous exposure of multiple generations to the enorhabditis elegans; in addition, The toxicity of biphenol S (BPS) to the caenorhabditis elegans was also investigated in this study, and the toxicity of BPA was compared with the toxicity of BPA, and the ecological risk of the substitute of BPA was analyzed. The main results of this study are as follows: (1) The acute and subacute exposure of BPA can significantly inhibit the body length, the level of activity and the number of offspring of the caenorhabditis elegans, and significantly increase the level of cell apoptosis and the expression of stress-related genes. after acute exposure, the head swing frequency of the caenorhabditis elegans is the most sensitive physiological index, and the exposure concentration is 0. 01 & mu; m, which can produce a significant negative effect on the caenorhabditis elegans, and after the subacute exposure, the body length and the head swing frequency are more sensitive than other physiological indexes, It is also possible to have a significant inhibitory effect on the caenorhabditis elegans at a concentration of 0.01. m u.M. The acute and subacute exposure of BPA did not have a significant effect on the accumulation of lipofuscin and the level of ROS in the nematode, but when the exposure concentration reached 0.1. m The results show that the toxic effect of BPA exposure to the caenorhabditis elegans is not the way of oxidative damage, and it is likely to be related to the increase of the level of apoptosis. The subacute exposure of BPA can lead to a significant increase in the expression of most stress-related genes in the Caenorhabditis elegans, even when the minimum exposure concentration is 0. 001. m u.M, and the exposure of BPA can also cause a significant increase in the expression of most stress-related genes. (2) The chronic exposure of BPA can significantly inhibit the body length, the activity level, the service life and the population number of the caenorhabditis elegans, and obviously improve the cell apoptosis level, the stress-related gene expression and the fluorescent protein expression amount of the transgenic line TJ375. After chronic exposure, the swing frequency of the head is the most sensitive physiological index, and the exposure concentration is 0. 001. m The chronic exposure of BPA did not have a significant effect on the accumulation of lipofuscin in Nematode, but it was able to significantly increase the level of apoptosis in the Caenorhabditis elegans when the exposure concentration was above 0.01. m In the experimental concentration range, the chronic exposure of BPA can significantly improve the expression of most stress-related genes in the nematode. Compared with the results of subacute exposure, the chronic exposure of BPA could lead to a more significant increase in the mRNA level of stress-related genes. When the exposure concentration of BPA reached 0.1. m u.M, the number of Caenorhabditis elegans was significantly reduced. The transgenic strain TJ375 can only detect the toxicity of BPA when the environmental concentration reaches 1. m u.M, and is not sensitive to other test indicators. (3) The continuous exposure of BPA can have a significant effect on the physiological index of the Caenorhabditis elegans when the exposure concentration is above 0.01. m In the case of lower exposure, the generation toxicity of Nematode in the offspring was different from generation to generation, while at the highest exposure concentration of 10. m u.M, there was no significant difference in the toxicity effect of the G1 and G4 of the Caenorhabditis elegans. in that G1 and G4 generation of the Caenorhabditis elegans, the exposure group of low concentration (0. 01. mu.M) can induce a more significant increase in the expression of the related gene in the in vivo compared with the high-concentration exposure group; compared with the G4 generation, The expression of the stress-related genes in the G1-generation of the Caenorhabditis elegans was more significant. (4) After the subacute exposure of BPS, the head swing frequency of the caenorhabditis elegans is the most sensitive physiological index, and the exposure concentration is more than 1. m while other physiological indicators will be significantly negatively affected only in the highest concentration exposure group. After sub-acute exposure, the accumulation of lipofuscin in the caenorhabditis elegans was not significantly affected, and the level of apoptosis was significantly increased only when the exposure concentration was 1. m The sub-acute exposure of BPS can cause a small increase in the expression of most of the gene in the Caenorhabditis elegans. In conclusion, exposure of low concentrations of BPA can produce developmental toxicity, neurobehavioral toxicity, and reproductive toxicity to the Caenorhabditis elegans. The effect of BPA exposure on the physiological level of Caenorhabditis elegans is likely to be related to the improvement of the level of cell apoptosis in vivo; the exposure of BPA can significantly improve the expression of stress-related genes in the Caenorhabditis elegans and destroy the steady state of the body; the transgenic line TJ375 is not sensitive to the exposure of BPA, The toxicity of BPA could not be effectively monitored; the toxicity of BPS was significantly less than the toxicity of BPA.
【學(xué)位授予單位】：華東理工大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：X171.5
本文編號(hào)：2318956
[Abstract]:Bisphenol A (BPA) is a world-recognized environmental endocrine disruptor and has been a star compound in toxicology studies. The world's annual demand for BPA is more than 4 million tons/ year, and the pollution in the environment is becoming more and more serious. The trace of BPA is found in soil, water, atmosphere and sediment, and BPA is also detected in human tissue and body fluids. In vivo and in vitro experiments have shown that the exposure of BPA can cause endocrine disruption, neurotoxicity, reproductive toxicity, immunotoxicity, and carcinogenicity. The epidemiological study of the population also found that diabetes, obesity, neurobehavioral disorders, liver function abnormalities, recurrent spontaneous abortion, and developmental disorders were associated with exposure to BPA. At present, the toxicity of BPA exposure to invertebrates is less. As a representative nematode species isolated from the soil, the Caenorhabditis elegans is one of the most commonly used modes in environmental exposure assessment and ecological toxicology studies. The study was conducted to study the ecological toxicity of Caenorhabditis elegans as the tested organism, mainly from the physiological, biochemical and molecular level to the acute and subacute exposure of BPA, the chronic exposure and the continuous exposure of multiple generations to the enorhabditis elegans; in addition, The toxicity of biphenol S (BPS) to the caenorhabditis elegans was also investigated in this study, and the toxicity of BPA was compared with the toxicity of BPA, and the ecological risk of the substitute of BPA was analyzed. The main results of this study are as follows: (1) The acute and subacute exposure of BPA can significantly inhibit the body length, the level of activity and the number of offspring of the caenorhabditis elegans, and significantly increase the level of cell apoptosis and the expression of stress-related genes. after acute exposure, the head swing frequency of the caenorhabditis elegans is the most sensitive physiological index, and the exposure concentration is 0. 01 & mu; m, which can produce a significant negative effect on the caenorhabditis elegans, and after the subacute exposure, the body length and the head swing frequency are more sensitive than other physiological indexes, It is also possible to have a significant inhibitory effect on the caenorhabditis elegans at a concentration of 0.01. m u.M. The acute and subacute exposure of BPA did not have a significant effect on the accumulation of lipofuscin and the level of ROS in the nematode, but when the exposure concentration reached 0.1. m The results show that the toxic effect of BPA exposure to the caenorhabditis elegans is not the way of oxidative damage, and it is likely to be related to the increase of the level of apoptosis. The subacute exposure of BPA can lead to a significant increase in the expression of most stress-related genes in the Caenorhabditis elegans, even when the minimum exposure concentration is 0. 001. m u.M, and the exposure of BPA can also cause a significant increase in the expression of most stress-related genes. (2) The chronic exposure of BPA can significantly inhibit the body length, the activity level, the service life and the population number of the caenorhabditis elegans, and obviously improve the cell apoptosis level, the stress-related gene expression and the fluorescent protein expression amount of the transgenic line TJ375. After chronic exposure, the swing frequency of the head is the most sensitive physiological index, and the exposure concentration is 0. 001. m The chronic exposure of BPA did not have a significant effect on the accumulation of lipofuscin in Nematode, but it was able to significantly increase the level of apoptosis in the Caenorhabditis elegans when the exposure concentration was above 0.01. m In the experimental concentration range, the chronic exposure of BPA can significantly improve the expression of most stress-related genes in the nematode. Compared with the results of subacute exposure, the chronic exposure of BPA could lead to a more significant increase in the mRNA level of stress-related genes. When the exposure concentration of BPA reached 0.1. m u.M, the number of Caenorhabditis elegans was significantly reduced. The transgenic strain TJ375 can only detect the toxicity of BPA when the environmental concentration reaches 1. m u.M, and is not sensitive to other test indicators. (3) The continuous exposure of BPA can have a significant effect on the physiological index of the Caenorhabditis elegans when the exposure concentration is above 0.01. m In the case of lower exposure, the generation toxicity of Nematode in the offspring was different from generation to generation, while at the highest exposure concentration of 10. m u.M, there was no significant difference in the toxicity effect of the G1 and G4 of the Caenorhabditis elegans. in that G1 and G4 generation of the Caenorhabditis elegans, the exposure group of low concentration (0. 01. mu.M) can induce a more significant increase in the expression of the related gene in the in vivo compared with the high-concentration exposure group; compared with the G4 generation, The expression of the stress-related genes in the G1-generation of the Caenorhabditis elegans was more significant. (4) After the subacute exposure of BPS, the head swing frequency of the caenorhabditis elegans is the most sensitive physiological index, and the exposure concentration is more than 1. m while other physiological indicators will be significantly negatively affected only in the highest concentration exposure group. After sub-acute exposure, the accumulation of lipofuscin in the caenorhabditis elegans was not significantly affected, and the level of apoptosis was significantly increased only when the exposure concentration was 1. m The sub-acute exposure of BPS can cause a small increase in the expression of most of the gene in the Caenorhabditis elegans. In conclusion, exposure of low concentrations of BPA can produce developmental toxicity, neurobehavioral toxicity, and reproductive toxicity to the Caenorhabditis elegans. The effect of BPA exposure on the physiological level of Caenorhabditis elegans is likely to be related to the improvement of the level of cell apoptosis in vivo; the exposure of BPA can significantly improve the expression of stress-related genes in the Caenorhabditis elegans and destroy the steady state of the body; the transgenic line TJ375 is not sensitive to the exposure of BPA, The toxicity of BPA could not be effectively monitored; the toxicity of BPS was significantly less than the toxicity of BPA.
【學(xué)位授予單位】：華東理工大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：X171.5
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