醋酸铅对PC-12细胞Bax、Bcl-2和p53蛋白表达的影响

用蛋白免疫印迹法研究醋酸铅对PC-12细胞Bax、Bcl-2和p53蛋白表达的影响.结果显示,当醋酸铅浓度达到0.1μmol·L-1时,Bax和p53的表达水平显著上升,决定凋亡是否发生的重要因素Bax/Bcl-2比率也明显上升,并存在浓度依赖关系.当醋酸铅浓度达到1μmol·L-1时,Bcl-2的表达显著降低.结合相关文献报道,Bax、Bcl-2和p53可能在铅诱导的细胞凋亡中起了重要的调控作用.     

环境浓度下铜锰复合暴露对人角膜上皮细胞的毒性效应

人正常角膜上皮细胞(HCE)经室内环境浓度下的铜(10μmol·L-1)、锰(20、40 μmol·L-1)单独及复合暴露后,采用CCK-8法对细胞活力进行检测,使用Annexin V-FITC/PI试剂盒测定了细胞凋亡率,并通过实时荧光定量PCR法从分子水平比较了不同暴露组炎症(NLRP3,IL-1β)和凋亡调控关键...     

乙酸铅对PC12细胞形态的影响

为了从形态学角度研究铅诱导细胞凋亡的机制,用100 μmol·L-1乙酸铅对PC12细胞进行染毒24 h,检测细胞整体形态、细胞骨架和细胞超微结构的改变.实验结果显示:乙酸铅染毒24 h后.细胞整体形态和细胞骨架无明显改变.但电子显微镜下已出现细胞核固缩;线粒体肿胀、正常结构消失并伴有空泡化;胞内出现鼓泡等现象.结合前期研究结果,推测线粒体在铅诱导细胞凋亡中起了重要作用.而细胞骨架不参与此过程.     

营养盐对东海浮游植物生长影响的现场培养实验

2005年6月在东海赤潮高发区通过现场培养实验探讨了NO3-N和PO4-P对浮游植物生长的影响.结果表明,在培养期间,初始浓度NO3-N 11.98 μmol·L-1,PO4-P 0.16 μmol·L-1以上时,总细胞数可以达到赤潮数量.培养期间主要优势种为米氏凯伦藻、东海原甲藻和中肋骨条藻.NO3-N和PO4-P的初始浓度为23.82 μmol·L-1及 0.41 μmol·L-1以上时,东海原甲藻最大细胞数可以达到106·L-1,当NO3-N和PO4-P浓度分别增至50.34 μmol·L-1和2.32 μmol·L-1时,米氏凯伦藻和中肋骨条藻最大细胞数也可达到106·L-1.初步判定几种浮游植物对营养盐的需求不同,其中中肋骨条藻与米氏凯伦藻要求较高,东海原甲藻次之,双角多甲藻要求最低.     

长期铅暴露对小鼠胸腺细胞活化后诱导细胞凋亡的影响

为探讨铅暴露对小鼠胸腺T细胞活化后诱导细胞凋亡的影响,将24只健康初断乳21日龄清洁级雄性KM小鼠随机分为4组:对照(蒸馏水)组和低(200 mg·L-1)、中(400 mg·L-1)、高(800 mg·L-1)剂量乙酸铅染毒组,每组6只.采用自由饮水方式进行染毒,连续染毒12周.染毒结束后,分离胸腺细胞,采用脂质体介导法将NFκB、AP-1、NFAT的荧光素酶报告基因转染到胸腺T细胞,刺激48 h后,检测荧光素酶活性.刺激胸腺细胞后,采用流式细胞仪检测凋亡相关膜分子Fas、FasL活性及细胞凋亡率.结果表明,与健康对照组小鼠相比,中、高剂量铅暴露组小鼠胸腺T细胞膜表面Fas及FasL表达明显增强(p＜0.05),细胞凋亡率明显增加(p＜0.05);相对于刺激前,anti-CD3刺激后各组小鼠胸腺T细胞的NFκB、AP-1、NFAT转录活性明显增加(p＜0.05),Fas/FasL的表达明显增强(p＜0.05),细胞凋亡率明显增加(p＜0.05),但相对于刺激后健康对照组,各剂量铅暴露组小鼠胸腺T细胞3种核因子转录活性明显下降(p＜0.05),Fas及FasL表达明显减弱(p＜0.05),细胞凋亡率明显降低(p＜0.05),且随着染铅剂量增加,均呈现下降趋势.     

三丁基锡对正常人胚胎羊膜细胞凋亡相关蛋白表达的影响

首次采用蛋白质印迹法研究三丁基锡(tributyltin,TBT)对正常人胚胎羊膜细胞FL(human amniotic cells)的Bcl-2、Bax和p53表达的影响.结果显示,在0、1、2、3、4μmol·L-1三丁基锡下作用2 h后,FL细胞的Bax蛋白表达量随浓度增加逐渐升高,Bcl-2蛋白表达量仅在3、4μmol·L-1浓度组相比于对照组明显下降.作为凋亡是否发生的决定因素,Bax/Bcl-2比值在三丁基锡高浓度组(3、4μmol·L-1)相比于对照有显著升高;而p53蛋白表达量没有明显变化.研究表明,Bcl-2和Bax参与了TBT诱导的细胞凋亡.     

Cr(Ⅵ)染毒对小鼠肝脏细胞凋亡以及凋亡相关蛋白的影响

为研究六价铬(Cr(Ⅵ))对生物体的细胞凋亡以及凋亡相关蛋白的改变,用0、25、50和100 mg·kg-1的重铬酸钾(K2Cr2O7)对小鼠进行1d或持续5 d灌胃,检测肝脏细胞凋亡以及p53、Bcl-2、Bax蛋白表达水平和caspase-3酶的激活.实验结果显示,Cr(Ⅵ)染毒1 d或5 d后,小鼠肝细胞凋亡率随染毒剂量的增加而增加,并呈明显的剂量-反应关系;p53、Bax蛋白表达升高,Bcl-2蛋白表达下降,easpase-3酶激活增加,且p53、Bcl-2蛋白表达水平和caspase-3酶激活水平在100mg·kg-1剂量组的2个染毒时间段与对照组相比均具有显著性差异,Bax蛋白表达水平仅在100 mg·kg-1染毒5 d的实验组与对照组相比具有显著性差异.结果表明,Cr(Ⅵ)能诱导小鼠肝脏细胞凋亡,p53、Bcl-2、Bax和caspase-3在这个过程中可能起重要作用.     

离子液体氯化1-辛基-3-甲基咪唑对EMT6细胞的毒性及其机理研究

为评估咪唑类离子液体的生物毒性,研究了氯化1-辛基-3-甲基咪唑([C8mim] [C1])对EMT6细胞的毒性作用和可能的机制.不同浓度(0.06、0.25、1 mmol·L-)的[C8 mim][Cl]对EMT6细胞染毒12h后,采用MTT方法检测细胞活力,二乙酸荧光素(FDA)方法检测细胞膜通透性的变化,Rhodamine 123染色方法检测线粒体膜电位的变化,ELISA方法检测了Caspase-3的活性,并测定了细胞内活性氧(ROS)的含量.结果表明,经[Csmim] [Cl]染毒12h后,EMT6细胞活力下降,并呈剂量依赖关系.当[Csmim] [Cl]浓度高于0.25 mmol·L-1时,细胞活力与对照相比,差异显著.研究还发现,[Csmim][Cl]染毒增加了EMT6细胞膜通透性,降低了线粒体膜电位并诱导产生过量的活性氧,增强了Caspase-3活性.实验结果表明,[C8mim] [Cl]染毒造成了EMT6细胞膜通透性的改变、活性氧的过量产生和凋亡分子表达的增强,这可能是[Csmim] [Cl]导致细胞凋亡和活力下降的主要原因.     

亚硝酸钠增强SMMC-7721细胞拮抗镉细胞毒性的研究

为了研究亚硝酸钠对肝源性细胞的毒物兴奋效应和拮抗镉诱导的细胞凋亡作用,以人体肝癌细胞SMMC-7721为模型,用3.13～3200mg·L-1亚硝酸钠作用24h,并采用四甲基偶氮唑蓝(MTT)法检测细胞的增殖活性.结果表明,细胞增殖活性表现出毒物兴奋效应.在毒物兴奋效应浓度范围内,选取最小效应浓度的亚硝酸钠(17mg·L-1)预适应细胞24h,然后再用25μmol·L-1氯化镉处理12h,细胞增殖活性结果表明,预适应的细胞能够耐受镉损害,这种细胞保护作用可以被一氧化氮清除剂抑制.流式细胞术及Hoechst33258/PI荧光双染结果表明,亚硝酸钠预适应的细胞能够耐受镉诱导的细胞凋亡.亚硝酸钠预适应的细胞再用氯化镉处理,与单纯氯化镉处理组相比,细胞内丙二醛(MDA)含量下降,超氧化物歧化酶(SOD)和过氧化氢酶(CAT)活性升高.同时,免疫细胞化学结果也显示,金属硫蛋白(MT)表达升高.总体而言,亚硝酸钠在3.13～800mg·L-1浓度范围内可促进细胞增殖活性并能够耐受镉诱导的细胞凋亡.     

铜、镉及其交互作用对泡泡草细胞超微结构的影响

为了解铜、镉对泡泡草根、茎、叶细胞亚显微结构的毒害及作用位点 ,以期为进一步研究鸭跖草属植物的耐重金属机理提供理论基础 ,本文通过透射电子显微技术研究了 75 μmol·L-1铜和 5 0 μmol·L-1镉及其交互污染对泡泡草根、茎、叶细胞超微结构的影响和铜、镉在细胞中的分布 .结果表明 :75 μmol·L-1铜和 5 0 μmol·L-1镉均对泡泡草根细胞造成明显损伤 .铜使根细胞产生质壁分离、细胞质浓缩、部分细胞空泡化 ,使线粒体脊突消失、结构模糊、外膜破坏 ;镉使根细胞空泡化 ,并在部分空泡化的细胞里产生大小不等的颗粒状物 ;铜、镉交互污染使根细胞受害程度加深 ,并兼有两者的受害症状特征 :线粒体结构彻底破坏、空泡化细胞里的颗粒物更大电子密度更高、质壁分离现象更普遍、质膜上的颗粒物沉淀更大 .铜、镉及其交互污染使泡泡草茎细胞产生质壁分离 ,对叶细胞无明显伤害 .所以铜、镉及其交互作用在试验浓度下 ,对泡泡草各器官的损伤程度为根 >茎 >叶 .     

Toxic effects of acetochlor, methamidophos and their combination on nifH gene in soil

Toxic effects of two agrochemicals on nifH gene in agricultural black soil were investigated using denaturing gradient gel electrophoresis （DGGE） and sequencing approaches in a microcosm experiment. Changes of soil nifH gene diversity and composition were examined following the application of acetochlor, methamidophos and their combination. Acetochlor reduced the nifH gene diversity （both in gene richness and diversity index values） and caused changes in the nifH gene composition. The effects of acetochlor on nifH gene were strengthened as the concentration of acetochlor increased. Cluster analysis of DGGE banding patterns showed that nifH gene composition which had been affected by low concentration of acetochlor （50 mg/kg） recovered firstly. Methamidophos reduced nifH gene richness that except at 4 weeks. The medium concentration of methamidophos （150 mg/kg） caused the most apparent changes in nifH gene diversity at the first week while the high concentration of methamidophos （250 mg/kg） produced prominent effects on nifH gene diversity in the following weeks. Cluster analysis showed that minimal changes of nifH gene composition were found at 1 week and maximal changes at 4 weeks. Toxic effects of acetochlor and methamidophos combination on nifH gene were also apparent. Different nifH genes （bands） responded differently to the impact of agrochemicals： four individual bands were eliminated by the application of the agrochemicals, five bands became predominant by the stimulation of the agrochemicals, and four bands showed strong resistance to the influence of the agrochemicals. Fifteen prominent bands were partially sequenced, yielding 15 different nifH sequences, which were used for phylogenetic reconstructions. All sequences were affiliated with the alpha- and beta-proteobacteria, showing higher similarity to eight different diazotrophic genera.     

Arsenic uptake by arbuscular mycorrhizal maize （Zea mays L.） grown in an arsenic-contaminated soil with added phosphorus

The effects of arbuscular mycorrhizal (AM) fungus (Glomus mosseae) and phosphorus (P) addition (100 mg/kg soil) on arsenic (As) uptake by maize plants (Zea mays L.) from an As-contaminated soil were examined in a glasshouse experiment.Non-mycorrhizal and zero-P addition controls were included.Plant biomass and concentrations and uptake of As,P,and other nutrients,AM colonization,root lengths,and hyphal length densities were determined.The results indicated that addition of P significantly inhibited root colonization and development of extraradical mycelium.Root length and dry weight both increased markedly with mycorrhizal colonization under the zero-P treatments,but shoot and root biomass of AM plants was depressed by P application.AM fungal inoculation decreased shoot As concentrations when no P was added,and shoot and root As concentrations of AM plants increased 2.6 and 1.4 times with P addition,respectively.Shoot and root uptake of P,Mn,Cu,and Zn increased,but shoot Fe uptake decreased by 44.6%,with inoculation, when P was added.P addition reduced shoot P,Fe,Mn,Cu,and Zn uptake of AM plants,but increased root Fe and Mn uptake of the nonmycorrhizal ones.AM colonization therefore appeared to enhance plant tolerance to As in low P soil,and have some potential for the phytostabilization of As-contaminated soil,however,P application may introduce additional environmental risk by increasing soil As mobility.     

Degradation of metabolites accumulated of benzo[a]pyrene by coupling Penicillium Chrysogenum with KMnO4

Several main metabolites of benzo[a]pyrene （BaP） formed by Penicillium chrysogenum, Benzo[a]pyrene-1,6-quinone （BP 1,6- quinone）, trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene （BP 7,8-diol）, 3-hydroxybenzo[a]pyrene （3-OHBP）, were identified by high-performance liquid chromatography （HPLC）. The three metabolites were liable to be accumulated and were hardly further metabolized because of their toxicity to microorganisms. However, their further degradation was essential for the complete degradation of BaP. To enhance their degradation, two methods, degradation by coupling Penicillium chrysogenum with KMnO4 and degradation only by Penicillium chrysogenum, were compared; Meanwhile, the parameters of degradation in the superior method were optimized. The results showed that （1） the method of coupling Penicillium chrysogenum with KMnO4 was better and was the first method to be used in the degradation of BaP and its metabolites; （2） the metabolite, BP 1,6-quinone was the most liable to be accumulated in pure cultures; （3） the effect of degradation was the best when the concentration of KMnO4 in the cultures was 0.01% （w/v）, concentration of the three compounds was 5 mg/L and pH was 6.2. Based on the experimental results, a novel concept with regard to the bioremediation of BaP-contaminated environment was discussed, considering the influence on environmental toxicity of the accumulated metabolites.     

Impact of atrazine and nitrogen fertilizers on the sorption of chlorotoluron in soil and model sorbents

Sorption of chlorotoluron in ammonium sulfate, urea and atrazine multi-solutes system was investigated by batch experiments. The results showed application of nitrogen fertilizers to the soil could affect the behavior of chlorotoluron. At the same concentration of N, sorption of chlorotoluron decreased as the concentration of atrazine increased on the day 0 and 6 in soil, respectively. The sorption of chlorotoluron increased from 0 to 6 d when soils were preincubated with deionized water, ammonium sulfate and urea solution for 6 d. That indicated incubation time was one of the most important factors for the sorption of chlorotoluron in nitrogen fertilizers treatments. The individual sorption isotherms of chlorotoluron in rubbery polymer and silica were strictly linear in single solute system, but there were competition sorption between pesticides or between pesticides and nitrogen fertilizers. That indicated the sorption taken place by concurrent solid-phase dissolution mechanism and sorption on the interface of water-organic matter or water-mineral matter.     

Potential of plant polyphenol oxidases in the decolorization and removal of textile and non-textile dyes

In this study an effort has been made to use plant polyphenol oxidases; potato （Solanum tuberosum） and brinjal （Solanum melongena）, for the treatment of various important dyes used in textile and other industries. The ammonium sulphate fractionated enzyme preparations were used to treat a number of dyes under various experimental conditions. Majority of the treated dyes were maximally decolorized at pH 3.0. Some of the dyes were quickly decolorized whereas others were marginally decolorized. The initial first hour was sufficient for the maximum decolorization of dyes. The rate of decolorization was quite slow on long treatment of dyes. Enhancement in the dye decolorization was noticed on increasing the concentration of enzymes. The complex mixtures of dyes were treated with both preparations of polyphenol oxidases in the buffers of varying pH values. Potato polyphenol oxidase was significantly more effective in decolorizing the dyes to higher extent as compared to the enzyme obtained from brinjal polyphenol oxidase. Decolorization of dyes and their mixtures, followed by the formation of an insoluble precipitate, which could be easily removed simply by centrifugation.     

Effects of two sludge application on fractionation and phytotoxicity of zinc and copper in the soil

The potential harm of heavy metals is a primary concern in application of sludge to the agricultural land. A pot experiment was conducted to evaluate the effect of two sludges on fractionation of Zn and Cu in soil and their phytotoxicity to pakchoi. The loamy soil was mixed with 0%, 20%, 40%, 60% and 80% (by weight) of digested sewage sludge (SS) and composted sludge (SC). The additions of both sludges caused a significant raise in all fractions, resulting in that exchangeable (EXCH) and organic bound (OM) became predominance of Zn and organic bound Cu occupied the largest portion. There was more available amount of Zn and Cu in SS treatments than SC treatments. During the pot experiment, the concentration of Zn in EXCH, carbonate (CAR) and OM and Cu in EXCH and OM fractions decreased in all treatments, so their bioavailability reduced. Germination rate and plant biomass decreased when the addition rate was high and the best yield appeared in 20% mixtures at the harvest of pakchoi. The two sludges increased tissue contents of Zn and Cu especially in the SS treatments. Zn in pakchoi was not only in relationship to ΔEXCH and ΔCAR forms but also in ΔOM forms in the sludge-soil mixtures. Tissue content of Cu in pakchoi grown on SC-soils could not be predicted by ΔEXCH. These correlation rates between Zn and Cu accumulation in pakchoi and variation of different fractions increased with time, which might indicate that sludges represented stronger impacts on the plant in long-term land application.     

Intraspecific differences in effects of co-contamination of cadmium and

A hydroponic experiment was carried out to study intraspecific differences in the effects of different concentrations of cadmium (Cd)(0-10 mg/L) and arsenate (As(V)) (0-8 mg/L) on the growth parameters and accumulation of Cd and As in six wheat varieties Jing-9428, Duokang-1, Jingdong-11, Jing-411, Jingdong-8 and Zhongmai-8. The endpoints of wheat seedlings, including seed germination,biomass, root length and shoot height, decreased with increasing the Cd and As concentrations. Significant differences in seed germination, biomass, root length, shoot height and the accumulation of Cd and As were observed between the treatments and among the varieties (p ＜ 0.05). The lethal dosage 50% were about 20, 80, 60, 60, 80 and 20 mg As/L for Jing-9428, Duokang-1, Jingdong-11,Jing-411, Jingdong-8 and Zhongmai-8, respectively, and the corresponding values for Cd were about 30, 80, 20, 40, 60 and 10 mg Cd/L, respectively. Among the six varieties, Duokang-1 was found to be the most resistant to Cd and As toxicity, and Zhongmai-8 was the most sensitive to Cd and As co-contamination. The resistance of the six varieties was found dependant on the seedling uptake of Cd and As. Duokang-1 was the most suitable for cultivation in Cd and As co-contaminated soils.     

Removal of heavy metals from sewage sludge by low costing chemical method and recycling in agriculture

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Isolation and preliminary characterization of a 3-chlorobenzoate degrading bacteria

A study was conducted to compare the diversity of 2-, 3-, and 4-chlorobenzoate degraders in two pristine soils and one contaminated sewage sludge. These samples contained strikingly different populations of mono-chlorobenzoate degraders. Although fewer cultures were isolated in the uncontaminated soils than contaminated one, the ability of microbial populations to mineralize chlorobenzoate was widespread. The 3- and 4-chlorobenzoate degraders were more diverse than the 2-chlorobenzoate degraders. One of the strains isolated from the sewage sludge was obtained. Based on its phenotype, chemotaxonomic properties and 16S rRNA gene, the organism S-7 was classified as Rhodococcus erythropolis. The strain can grow at temperature from 4 to 37℃. It can utilize several （halo）aromatic compounds. Moreover, strain S-7 can grow and use 3-chlorobenzoate as sole carbon source in a temperatures range of 10-30℃ with stoichiometric release of chloride ions. The psychrotolerant ability was significant for bioremediation in low temperature regions. Catechol and chlorocatechol 1,2-dioxygenase activities were present in cell free extracts of the strain, but no （chloro）catechol 2,3- dioxygenase activities was detected. Spectral conversion assays with extracts from R. erythropolis S-7 showed accumulation of a compound with a similar UV spectrum as chloro-cis,cis-muconate from 3-chlorobenzoate. On the basis of these results, we proposed that S-7 degraded 3-chlorobenzoate through the modified ortho-cleave pathway.     

Single and joint effects of pesticides and mercury on soil urease

The influence of two pesticides including chlorimuron-ethyl and furadan and mercury （Hg） on urease activity in 4 soils （meadow burozem and phaeozem） was investigated. The soils were exposed to various concentrations of the two pesticides and Hg individually and simultaneously. Results showed that there was a close relationship between urease activity and organic matter content in soil. Chlorimuron-ethyl and furadan could both activate urease in the 4 soils. The maximum increment of urease activity by chlorimuronethyl was up to 14%-18%. There was almost an equal increase （up to 13%-21%） in the urease activity by furadan. On the contrary, Hg markedly inhibited soil urease activity. A logarithmic equation was used to describe the relationship （P〈0.05） between the concentration of Hg and the activity of soil urease in the 4 tested soils. Semi-effect dose （ED50） values by the stress of Hg based on the inhibition of soil urease in the 4 soils were 88, 5.5, 24 and 20 mg/kg, respectively, according to the calculation of the corresponding equations. The interactive effect of chlorimuron-ethyl or furadan with metal Hg on soil urease was mainly synergic at the highest tested concentrations.