多溴联苯醚和全氟烷基酸的分子毒理机制研究

多溴联苯醚(PBDEs)和全氟烷基酸(PFAAs)是两类使用量大、环境污染广泛、人体暴露严重的新型有机污染物,2009年已纳入《斯德哥尔摩公约》持久性有机污染物(POPs)名单,但其毒性效应及作用机制并不明确.本文综述了本课题组近几年针对多溴联苯醚PBDEs和全氟烷基酸PFAAs的分子毒理机制研究工作,主要集中在这两类污染物对甲状腺系统、雌激素系统和肝脏脂肪酸代谢系统干扰效应的分子机制研究.本文分别从分子、细胞和活体三个层面,研究了污染物与核受体的直接结合作用、结合后受体的构象变化、细胞内受体的转录活性、以及活体暴露后受体调控基因的表达变化,由此阐明了污染物通过与受体直接作用导致细胞和活体生物功能改变的分子机制.同时结合计算模拟,探讨了污染物生物效应与其化学结构之间的关系,发现污染物的受体活性取决于它们与受体结合的空间构型,而其活性强度基本与二者的结合能力一致,主要受疏水作用和氢键的影响.此外,还通过研究污染物与天然配体转运蛋白的相互作用,明确了各个污染物与转运蛋白的结合能力,探讨了其构效关系,并评估了污染物对天然配体在体内转运过程的潜在干扰效应.通过上述研究工作,提出了多层面、多靶点研究环境污染物分子毒理机制的新思路,建立和引进了研究污染物与生物靶分子相互作用的新方法,发现了PBDEs、PFAAs与TR、ER、PPARγ核受体结合的新模式,为深入了解这些污染物的分子毒理机制提供了有用的信息和有效的研究手段.     

吸烟烟气的细胞毒性作用和茶多酚保护作用的研究

以体外培养的中国仓鼠肺成纤维细胞V79为模型,用分光光度法和ESR自旋标记方法研究了吸烟烟气对细胞的损伤以及抗氧化剂茶多酚（GTP）的保护作用。结果表明,吸烟烟气能引发V79细胞膜的脂质过氧化,导致细胞死亡,使膜浅层的流动性增加,但对膜深层的动态性质没有明显改变。加入GTP后能抑制烟气引起的膜脂过氧化和膜流动性的改变以及细胞死亡,且这种抑制作用与GTP的浓度呈量效关系,而GTP本身对细胞膜浅层的流动性无明显作用,但对深层的流动性则有一定的影响。此外,还用马来酰亚胺自旋探针研究吸烟烟气对细胞膜膜蛋白巯基结合位点局部构象的影响,结果发现,随着烟气作用增强,ESR波谱的强弱固定化比值（S/W）增大,表明膜蛋白上巯基结合位点处结构变得紧密,这说明吸烟烟气改变了膜蛋白的构象,GTP本身在高浓度（＞0.1mg/mL）时也能使S/W增大,当低浓度（＜0.05mg/mL）则没有影响。当GTP浓度为0.01mg/mL时就可以抑制烟气引起的S/W的增大。     

应用应激蛋白HSP70作为生物标志物研究锌、铜及其联合毒性对鲫鱼肝脏的影响

将应激蛋白 (HSP70 )作为生物标志物 ,以银鲫 (Carassiusauratusgibelio)作为实验生物 ,经过 4 0d不同浓度铜 (Cu2 )、锌(Zn2 )及其混合物暴露后 ,运用SDS PAGE和WesternBlotting方法研究了鲫鱼肝脏组织内应激蛋白HSP70的诱导表达情况 .结果表明 :在实验浓度下 ,Cu2 ,Zn2 及其混合物对鱼肝脏内HSP70有显著的诱导 (P <0 0 5 ) ,而将Cu2 和Zn2 实验组与其混合物实验组相比 ,Cu2 ,Zn2 表现为一定的协同作用 .实验还发现 ,在国家渔业水质标准Zn2 浓度下 ,鱼肝脏组织内HSP70已经有明显的表达 ,说明分子生物学指标要比传统的环境检测指标敏感 ,对污染物具有早期预警的作用 ,将其作为生物标志物来监测环境中的重金属污染具有重要意义     

生活污水中蛋白质测试方法研究

分别用Folin-酚改进法、考马斯亮兰法和紫外分光光度法等3种测试方法测试了上海市某小区生活污水中的蛋白质含量,对这3种测试方法进行了比较,并采用氨基酸仪分析了生活污水中蛋白质水解后的氨基酸化学组成.试验结果表明,Folin-酚改进法测试蛋白质浓度的准确性与精确性优于考马斯亮兰法和紫外分光光度法,可用于测定生活污水中蛋白质的浓度;氨基酸仪法测得该生活污水中含有17种氨基酸,其中最多的为谷氨酸(3.6 mg/L),含量最低的是脯氨酸(0.8 mg/L);所测生活污水中蛋白质COD占总COD的18.9%.     

First and second line mechanisms of cadmium detoxification in the lichen photobiont Trebouxia impressa (Chlorophyta)

“First line” defence mechanisms, such as phytochelatin biosynthesis, and “second line” mechanisms, such as stress protein induction, were investigated in cadmium-exposed cells of Trebouxia impressa Ahmadjian, a green microalgal species that is a common photobiont of the lichen Physcia adscendens (Fr.) H. Olivier. When T. impressa cells were exposed to 0, 9 and 18 μM Cd for 6, 18 and 48 h, glutathione and phytochelatins efficiently protected the cells against Cd damage. By contrast, the highest Cd concentration (36 μM) at the longest exposure-time (48 h) caused marked drops in glutathione and phytochelatin content, several types of ultrastructural damage, and decreases in cell density and total chlorophyll concentration. In this case, induction of stress proteins was observed, but only long after the induction of phytochelatins. Thus, stress proteins could represent a “second line” mechanism to counteract Cd stress, activated when there is a decline in the “first line” mechanism of Cd detoxification given by phytochelatins.     

产朊假丝酵母CANDIDA UTILIS细胞壁对铜离子吸附位点的研究

目前认为所有生物大分子都对重金属离子有较强吸附性 ,但某些分子可特异地吸附重金属 ,并介导酵母细胞对重金属的抗性[1] .其中细胞表面的高分子聚合物倍受关注 ,酵母细胞壁约含 4 0种蛋白分子 ,但有关这些蛋白质在对重金属离子吸附中的作用目前还未见报道 .产朊假丝酵母Ｃａｎｄｉｄａｕｔｉｌｉｓ是重要单细胞蛋白生产菌株之一 ,因其营养条件要求不高 ,对工业废水有较强耐受和转化能力而广泛用于生产和研究 .我们观察了产朊假丝酵母细胞与分离纯化的细胞壁对铜离子的吸附能力差异 ,并对细胞壁上重金属离子吸附位点进行了研究 .1　材料与方…     

粘虫受中红侧沟茧蜂寄生后的生理变化

粘虫（Ｐｓｅｕｄａｌｅｔｉａ ｓｅｐａｒａｔａ）被中红侧沟茧蜂（Ｍｉｃｒｏｐｌｉｔｉｓ ｍｅｄｉａｔｏｒ）寄生以后生理生化上发生了一系列变化。通过被寄生粘虫的生长（体重）、发育（龄期）、血淋巴总蛋白含量和酚氧化酶活性等生理生化指标动态的研究,试图探讨中红侧沟茧蜂对寄主粘虫的生理调节方式和途径,进一步探讨寄生蜂同寄主相互协调、相互影响的关系。图６参２０     

Effect of a subchronic exposure to Simazine on energetic metabolism of common carp (Cyprinus carpio)

The energetic parameters, such as glycogen, glucose, proteins, lactate and adenosine triphosphate (ATP) content and lactate deshydrogenase (LDH) activity in tissues and blood of carps from simazine (2-chloro-4,6-bis(ethylamino)-s-triazine) contaminated waters were investigated. In order to confirm the field results, a laboratory experiment was designed in which carps were exposed to simazine at the concentration level of 45 μ g. L^{? 1} (10-fold of the amount found in natural waters) for 90 days. Fish from a contaminated reservoir showed low glycogen concentrations in hepatopancreas and muscle, while fish in another contaminated reservoir showed high LDH activity together with an increase in lactate content in muscle. Laboratory findings did not confirm field results, and fish exposed to simazine did not show alterations in the parameters studied. The results suggest that carps were not stressed by the presence of the simazine at the concentration levels found in both studies and the mechanisms of defense covered the energetic demand.     

Effect of in vitro administered 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on DNA-binding activities of nuclear transcription factors in NIH-3T3 mouse fibroblasts

Abstract

Most modern pesticides are expensive. Application of excessive dosage rates is likely to cause undesirable biological side‐effects and is economically wasteful. Non‐uniform distribution of the spray cloud, or application at the wrong time, may result in failure to control the pest. It is the responsibility of the field operator to acquire sufficient knowledge and skill to ensure proper use of the control agents, to increase efficiency of their usage and to reduce unwanted side‐effects. To achieve this goal, he must take into consideration the various physical factors that govern field performance of pesticides.

A simple relationship exists between the spray volume and emission rate used, and droplet size produced. The use of extremely low spray volumes (i.e., those less than 2.0 litre per ha) for forest insect control in Canada, as opposed to higher volumes used in agriculture, necessitates the release of fine droplets (ranging from 20 to 70 μm in diameter) to obtain adequate coverage of the target area. These droplets take a long time to sediment downwards, evaporate in‐flight, become smaller in size and/or form powdery residues, thus contributing to off‐target drift and impaired droplet adhesion to target surfaces. Physical factors such as rain washing, degradation by sunlight and erosion by wind also influence the longevity of pesticide deposits on foliage which is crucial during the critical period of pest control.

Factors affecting the mode of entry into insects are related to the type of ingredients used in formulation. If a pesticide acts via crawling contact, formulations which would provide surface deposits would be more beneficial than emulsions or oil‐based mixes which tend to undergo penetration into foliar cuticle. Physical factors that affect field performance of a pesticide tank mix are related to phase separation and ‘breakdown of emulsions’ in the application equipment; ‘agglomeration and caking’ of wettable powder dispersions at the bottom of the tank; impaired flow behaviour of highly viscous formulations; and coarse atomization of high‐viscosity tank mixes leading to poor target cover.     

2,3,7,8-四氯苯并二噁英（TCDD）短期染毒可造成着床前胚胎丢失并伴随雌性生殖器官中毒物相关蛋白的诱导表达（Short Time Exposure of TCDD Causing Loss of Implantation Failure of Embryos and Its Associated with Induced Expression of Relevant Proteins）

为探明妊娠早期胚胎的丢失是否与卵巢、输卵管、子宫组织受到2,3,7,8-四氯苯并二噁英(TCDD)直接毒害有关,检测了NIH小鼠胚胎着床前期和后期TCDD暴露对胚胎毒性影响的敏感性,并利用免疫组化方法分析了模型动物肝脏、子宫、输卵管和卵巢组织中TCDD所引起的AhR、ARNT以及Cyp1a2分子标记物的变化.检测发现:妊娠第9d,100ng·kg-1·d-1剂量TCDD经口染毒,造成胚胎着床数量减少,且着床前期暴露的影响大于着床后期;子宫蜕膜反应受到明显抑制;胚胎迁移率没有明显变化,但胚胎数量减少.免疫组织化学分析发现正常组小鼠的肝脏、子宫、输卵管和卵巢组织中有AhR和Cyp1a2弱阳性信号表达,ARNT有细胞核的强阳性信号表达;妊娠第1～8d、第1～3d和第4～8d处理组小鼠肝脏、子宫、输卵管和卵巢组织中的AhR、Cyp1a2的阳性面积和光密度值均高于正常组;随处理时间和组织蓄积量的增加,ARNT在组织中的变化由胞核(妊娠第1～3d组)表达到胞浆(妊娠第4～8d组)表达,然后完全无表达(妊娠第1～8d组).以上研究结果表明:TCDD对早期妊娠小鼠子宫、输卵管和卵巢组织中的AhR、ARNT和Cyp1a2的激活和代谢方式与肝脏相同,说明雌性生殖系统中的组织有TCDD蓄积和代谢活性,这可能是导致早期胚胎迁移、着床等过程改变,造成胚胎丢失的重要原因.