RTCA与CCK-8法用于检测柴油废气颗粒物致支气管上皮细胞毒性的比较研究

运用实时无标记细胞分析系统(RTCA)和Cell Counting Kit-8(CCK-8)法分别检测柴油废气颗粒物(DEP)致支气管上皮细胞(HBE)细胞毒性,从而对2种方法进行比较研究.分别以浓度为0、3.5、7、14、28和56 mg·L-1 2种柴油废气标准参考颗粒物(Standard Reference Material 1650b,SRM 1650b;Standard Reference Material 2975,SRM 2975)对 HBE 细胞进行暴露处理,分别暴露6、12、24和48 h后,检测不同DEP致HBE细胞毒性,比较各组之间细胞存活率或标准化细胞指数(normalized cell index,NCI)值的差异,考察2种方法的优缺点.并用细胞凋亡实验检测各差异组之间的凋亡率.在相同染毒浓度及暴露时间,与SRM 1650b相比,SRM 2975对HBE细胞的毒性更强.在RTCA检测DEP致HBE细胞毒性时,低浓度DEP组的NCI值已经表现出与对照组有统计学差异(P＜0.05),而相同时间条件下,CCK-8法在更高浓度的DEP组才检测出显著的细胞活性下降(P＜0.05).且由细胞凋亡实验证实,与对照组相比,低浓度DEP组的细胞凋亡率已经有统计学差异.相对于CCK-8法,RTCA更适用于检测DEP致贴壁HBE细胞毒性.CCK-8法更适用于检测DEP致悬浮细胞的细胞毒性或与气液暴露装置联用时的贴壁/悬浮细胞毒性.     

稀土矿城市不同季节大气可吸入颗粒物中稀土含量特征及颗粒物细胞毒性

为探讨典型稀土矿城市不同季节大气可吸入颗粒物(inhalable particulate matter,PM10)中稀土元素污染特征及其细胞毒性响应,将前期采集于包头市的PM10颗粒物进行提取,检测PM10中的稀土元素(rare earth elements,REEs)含量,并将人肺上皮细胞(A549)暴露于不同浓度水平(25,50,100μg·m L-1)的PM10样品和标准颗粒物1649b(standard reference material,SRM1649b)暴露液,用WST-1法测定暴露24 h后的细胞活性,用2’7’二氯荧光素二醋酸盐(2’7’-dichlorofluorescein diacetate,DCFH-DA)荧光探针法和彗星实验分别测定暴露3 h后的细胞内活性氧(reactive oxygen species,ROS)产生水平和DNA双链损伤程度。结果表明,包头春、夏季大气PM10和SRM1649b均引起A549细胞活性下降,并诱导细胞内ROS生成量增加,造成显著的细胞内DNA损伤,含REEs的大气颗粒物毒性显著高于标准颗粒物。与春季相比,包头夏季PM10对细胞活性的抑制程度更高,造成更多的DNA双链损伤,从而表现出更强的细胞毒性和遗传毒性。包头PM10呈现明显的轻稀土元素(light rare earth elements,LREEs)富集,铈(Ce)、钷(Pm)、镧(La)和钕(Nd)含量占稀土总量的50%以上。LREEs均与细胞活性和细胞内ROS产生水平呈负相关性,包头春季和夏季PM10中稀土元素含量的差异是导致包头PM10细胞毒性效应不同于标准颗粒物且具有季节性差异的原因之一。     

柴油尾气颗粒对秀丽隐杆线虫慢性毒性效应的初步研究

柴油机尾气颗粒(diesel exhaust particles,DEP)对PM_(2.5)有重要贡献,已有研究发现其与暴露人群的肺癌、慢性支气管疾病、心血管疾病等密切相关。以DEP为研究对象,以秀丽隐杆线虫(Caenorhabditis elegans)为模式生物,观察了柴油机尾气颗粒在线虫体内的分布情况,并以线虫头部摆动频率、体长体宽、活性氧自由基(ROS)水平、细胞凋亡水平、脂褐素水平为评价指标探究了DEP暴露对线虫的慢性毒性效应。结果表明,经过10 d的慢性暴露,1 mg·L~(-1)的DEP明显富集在线虫的咽部和肠道,中低浓度的(0.001、0.01、0.1 mg·L~(-1))DEP未出现明显的体内富集现象。随着浓度的增加,线虫的头部摆动频率、体长和体宽均呈先上升后降低的趋势,差异具有统计学意义(P0.01)。中浓度组(0.1 mg·L~(-1))线虫体内ROS水平和细胞凋亡水平较对照组均有所降低,而高浓度(1 mg·L~(-1))的DEP显著提高了线虫体内的ROS水平和细胞凋亡水平(P0.05)。中高浓度的DEP均能使线虫肠道脂褐素积累升高(P0.05)。研究揭示了DEP具有在生物组织体内富集的特点,长期积累会影响其运动行为,抑制其生长发育,导致其体内ROS水平和细胞凋亡升高并使肠道老化,具有潜在的神经和发育毒性效应。研究结果可为DEP的人群暴露的慢性毒性评价提供理论参考。     

镉诱导大鼠肾细胞凋亡及其机理的研究

为探讨镉(Cd)致大鼠肾细胞(NRK cell)凋亡的死亡受体途径,分别将终浓度为0、10、20、40、60μmol·L-1的CdCl2添加到培养液中,对NRK细胞暴露6h.应用流式细胞仪检测NRK细胞凋亡率;分光光度计检测Caspase-8蛋白活性;半定量RT-PCR检测Caspase-8、Fas的mRNA表达.结果表明,随着CdCl2暴露浓度的升高,NRK细胞凋亡率升高,Caspase-8蛋白活性增强,Caspase-8、Fas的mRNA表达增强,并呈现剂量-效应关系.说明镉可以通过死亡受体这条通路来引起NRK细胞凋亡.     

SO_2与CeO_2超细颗粒非均相反应产物的细胞毒性效应

大气二次细颗粒物(secondary fine particulate matters,SFPMs)是我国城市大气PM_(2.5)的主要组成部分。然而由于PM_(2.5)组成成份复杂,其毒性产生的来源并不明确。在本研究中,我们以二氧化铈(CeO_2)超细颗粒物(UFPs)为大气细矿物质颗粒模型,研究了SO_2气体在模拟大气环境中,如湿度(RH)、紫外光照(UV)和NO_2存在条件下,在CeO_2UFPs界面经多相反应生成的二次无机细颗粒物的性质及与细胞毒性的构效关系。实验通过实时高通量细胞分析系统,实时观察了CeO_2-SFPMs暴露对小鼠单核巨噬细胞(RAW264.7)增殖的影响;并进一步检测了CeO_2-SFPMs对细胞膜通透性和细胞凋亡的影响。结果表明,SO_2与CeO_2UFPs作用后可转化为硫酸盐,在有NO_2存在下转化更为明显。CeO_2-SFPMs对细胞毒性效应与其生成的环境条件相关,并具有时间效应性。RAW264.7细胞暴露于CeO_2-SFPMs 8 h,细胞增殖无明显变化;暴露8~25 h后,CeO_2-SFPMs对细胞增殖的抑制率随CeO_2@CeO_2+SO_2@CeO_2+SO_2+RH≈@CeO_2+SO_2+RH+UV@CeO_2+SO_2+RH+NO_2的顺序显著升高。CeO_2-SFPMs对Raw264.7细胞膜通透性和细胞凋亡的影响研究也证明CeO_2-SFPMs@CeO_2+SO_2+RH+NO_2产生的细胞毒性最明显。     

天然提取物抗PM2.5诱导A549细胞凋亡的作用

采集北京城区大气可吸人颗粒物中的细颗粒物(PM25),用其对人肺腺癌A549细胞染毒,探讨PM25对细胞增殖的毒性和诱导细胞凋亡的作用,并且考察了加入不同浓度的红豆越橘提取物和竹叶提取物对其的抗性作用.实验采用MTT法检测细胞增殖作用,采用Annexi V-FITC/PI双染法和流式细胞仪检测细胞凋亡.结果显示:PM2...     

重组人金属硫蛋白-Ⅲα短肽毒性效应的初步研究

研究重组人金属硫蛋白Ⅲα短肽(rh-MT-Ⅲα)致HaCaT细胞、L929细胞和秀丽隐杆线虫(Caenorhabditis elegans,C.ele-gans)毒性效应,为rh-MT-Ⅲα的安全应用提供实验基础.细胞暴露于不同浓度的rh-MT-Ⅲα溶液(0～400μg·mL-1),24h后检测细胞存活率和乳酸脱氢酶(LDH)活性,观察细胞形态并检测细胞凋亡情况.结果表明,各剂量组2种细胞的存活率,与对照组相比均无显著性差异,细胞形态也未观察到显著变化.在400μg·mL-1浓度下,LDH释放率明显增加,细胞凋亡率上升.线虫实验中,将线虫培养并同步化至L1期后进行染毒,剂量为5、50和500 μg·mL-1,暴露48 h后检测其运动行为(头部摆动和身体弯曲频率)、发育(体长)及摄食行为(咽泵频率),各指标均未有明显改变.研究表明,在一定的剂量水平下,rh-MT-Ⅲα对HaCaT细胞、L929细胞和秀丽隐杆线虫无明显毒性效应.     

微囊藻毒素-LR通过线粒体途径诱导人支气管上皮细胞凋亡

探讨线粒体Caspase依赖性途径是否参与微囊藻毒素-LR(microcystin-LR,MC-LR)诱导人支气管上皮细胞(human bronchial epithelial cells,16HBE)凋亡过程。将处于对数生长期的16HBE分别暴露于终浓度为0(对照组)、2.5、5、10μg·m L-1的微囊藻毒素-LR和10μg·m L-1MC-LR+50μmol·L-1Caspase广谱抑制剂Z-VAD-FMK,持续24 h和48 h。检测细胞凋亡率,线粒体跨膜电位(ΔΨm),Caspase-3和Caspase-9相对表达量。结果显示,与对照组相比,各浓度染毒组细胞凋亡率和Caspase-3、Caspase-9相对表达量均升高,10μg·m L-1MC-LR染毒组线粒体膜电位降低;与10μg·m L-1MC-LR组相比,10μg·m L-1MCLR+50μmol·L-1Z-VAD-FMK组细胞凋亡率明显降低,Caspase-3和Caspase-9相对表达量降低,差异均有统计学意义(P0.05)。且随着MC-LR染毒浓度的升高或染毒时间的延长,16HBE细胞凋亡率和Caspase-3、Caspase-9相对表达量呈升高趋势。研究表明,MC-LR可以通过线粒体Caspase依赖性途径诱导16HBE细胞凋亡。     

PCB77和Cd共同暴露诱导的线粒体损伤及其介导的联合毒性效应

本文以人肝癌细胞系SMMC-7721为生物模型,研究并比较了PCB77和Cd~(2+)对SMMC-7721细胞的单一与联合毒性.通过细胞活性检测发现,当Cd~(2+)暴露浓度为20μmol·L-1,PCB77暴露浓度为25μmol·L~(-1),暴露时间为48 h时,两种污染物产生明显的联合毒性.二者共同暴露虽然不影响细胞膜的完整性,但引起了线粒体膜电位下降、膜通透性显著增加,进而诱导细胞凋亡,同时胞内ROS水平升高.结果表明,PCB77和Cd~(2+)的共同暴露可引发联合肝细胞毒性,其毒性机理是线粒体膜电位下降和膜通透性增加,进而引发细胞凋亡.     

三种典型纳米颗粒物造成的人体肺细胞氧化应激和炎症效应(Oxidative Stress and Inflammatory Effect on A549 Cell Line Induced by Three Typical Nano-Particles )

采用体外细胞暴露实验研究了人肺腺癌细胞系(A549)单层细胞暴露于50和500μg·mL-1两种浓度纳米氧化钛、纳米氧化硅、碳纳米管和晶体石英砂等四种颗粒物后产生的氧化应激和炎症反应.用细胞活度、细胞内活性氧总量和细胞上清液中白细胞介素8(IL-8)表达量表征暴露效应.研究结果表明,纳米氧化钛、纳米氧化硅和碳纳米管在体外暴露实验过程中均发生不同程度的聚集;细胞暴露48h后,三种纳米颗粒物均使A549细胞活度下降,诱导细胞产生过量活性氧,同时刺激细胞IL-8表达量增高;三种纳米颗粒物中,纳米氧化钛和纳米氧化硅对细胞活度影响较大,碳纳米管诱发的炎症效应较另两种纳米材料强.     

Conservation by Design

Abstract: Conservation researchers are increasingly aware of the need to conduct interdisciplinary research and to engage nonscientists in practical applications of conservation biology. But so far, industrial designers have been left out of such collaboration and outreach efforts. Conservation of wildlife often depends on products such as nest boxes, feeders, barriers, and corridors, all of which have a designed component that is frequently overlooked. Furthermore, many products are adopted without testing on short or long time scales. We argue that the design of products for conservation, and hence their functionality, effectiveness, and value, can be improved through collaboration with industrial designers. We see four key benefits that can arise from interactions with industrial designers: improvement of product quality and value, innovation and improvement in functionality of products, harmonization of conservation products with local values, and development of a psychological biomimesis approach to design. The role of industrial designers in conservation projects would be to improve factors such as product durability, affordability, functionality, and aesthetic appeal to local people. Designers can also help to create multiple product options whose success can be tested in the field. We propose that collaborations with industrial designers can contribute to the development of improvements to existing products and innovations in the practice of animal conservation.     

Shipboard bilge water treatment by electrocoagulation powered by microbial fuel cells

Reveals the synergy between microbial fuel cells and electrocoagulation. Demonstrates MFC-ECC shipboard wastewater treatment is advantageous. MFC-ECC integration enables energy neutral bilge water treatment. Ships generate large amounts of wastewater including oily bilge water, blackwater and greywater. Traditionally they are treated separately with high energy consumption. In this study we demonstrate the feasibility that these waste streams can be treated using an integrated electrocoagulation cell (ECC) and microbial fuel cell (MFC) process, which not only synergized the contaminants removal but also accomplished energy neutrality by directly powering EC with MFC electricity. Results showed that MFC stack powered ECC removed 93% of oily organics, which is comparable to the performance of an external DC voltage powered ECC. In the meantime, more than 80% of COD was removed from MFCs when fed with either acetate or municipal wastewater. Moreover, the ECC electrode area and distance showed notable effects on current generation and contaminants removal, and further studies should focus on operation optimization to enhance treatment efficiency.     

Products of denitrification by a marine bacterium as revealed by gas chromatography

Gas chromatography was used to analyse the gases released by growing cultures of the denitrifying marine bacterium Pseudomonas perjectomarinus. When nitrate was provided as final oxidant, CO₂ was the first detectable gas released. Following the peak rate of release of CO₂ at approximately 16 h, N₂ liheration began and peaked at 30 h. When N₂O was provided as terminal oxidant, CO₂ and N₂ release began within a few hours and the rate of liberation of both increased for 45 to 50 h before leveling. Cell-free extracts of bacteria from denitrifying cultures of P. perfectomarinus were incubated anaerobically with nitrate, malic acid and electron transfer cofactors provided, and the gases in the atmosphere above the reaction mixtures were analysed. NO was found to be present transiently, and N₂O was discerned after prolonged incubation. The applicability of gas chromatography to analyses of marine ecosystems and the need for better means of sampling the atmosphere above enzymatic reaction mixtures were disoussed.     

Biodeposition by salt-marsh invertebrates

The importance of salt-marsh invertebrate biodeposits in biogeochemical cycles is discussed. Yearly biodeposition rates of nitrogen, phosphorus, potassium, magnesium, maganese, copper, zinc, and molybdenum are reported for 5 invertebrate species. The 5 species (Littorina irrorata, Arcatula demissa, Polymesoda caroliniana, Uca pugnax and U. pugilator) deposit 1709 g dry weight m² year^-1, of which 455 g m² year^-1 is organic material. Calculations suggest that the invertebrates are capable of processing 53% of the marsh's yearly production, but the actual amount utilized is probably substantially less than this.Contribution No. 716 from the Virginia Institute of Marine Science, and Contribution No. 295 from the University of Georgia Marine Institute.     

Echolocation by cave swiftlets

Summary The ability of cave dwelling swiftlets (Collocalia spodiopygius) to detect small cylindrical obstacles by echolocation was tested in a 3x9 m flight chamber. Although there was great individual variability, 6.3 mm obstacles were avoided much more often in total darkness than 1.5 wires (P<0.001). Flash photographs showed that the latter were avoided only by chance (about 50% misses). Obstacles 3.0 mm in diameter were avoided only slightly more often than 1.5 mm wires.Even in the light these swiftlets avoided the obstacles in only about 75% of the trials, probably because gentle touches with the primary feathers caused little discomfort.The 3.0 and 6.3 mm obstacles tended to be avoided more often on first encounters after they had been shifted horizontally by 10–30 cm, probably because the birds became more attentive.     

Human carcinogenesis by arsenic

Arsenic is one of the few human carcinogens for which there is not yet a reliable animal cancer model. As such, the classification of arsenic as a carcinogen is based upon data derived from human epidemiologic studies. Although the mechanisms of action of arsenic as a toxic agent have been known for many years, the inability to produce cancer with arsenic in laboratory animals has confounded the operational characterisation of arsenic as initiator, promoter, complete carcinogen, or cocarcinogen for humans. Arsenic is clearly a genotoxic agent that induces chromosomal aberrations, micronuclei and sister chromatid exchange in mammalian cells as well as neoplastically transforms Syrian hamster embryo cells; however, it is not a classical point mutagen. This paper reviews some of the scientifically based issues relating to arsenic and risk assessment.     

Direct measurement of dissolved organic carbon release by phytoplankton and incorporation by microheterotrophs

It is now possible to divide particulate primary production into algal and heterotrophic components without physical separation. This depends on two innovations, the introduction of isotope in the form of labelled dissolved product(s) of primary production and the employment of a data analysis specifically designed for tracer kinetic incorporation experiments. The ¹⁴C technique described by Steemann Nielsen (1952) is inapplicable in the analyses of certain classes of systems and kinetic tracer incorporation experiments must be employed instead. We show that measurement of PDOC production rate requires such kinetic tracer analyses. Measurements made in the laboratory on water taken from 2 m depth in South West Arm of the Port Hacking estuary showed that: (1) the steady-state rate of PDOC production was 0.10 to 0.13 mg C.m^-3.h^-1; (2) the rate of PDOC incorporation into microheterotroph particulate organic carbon was 0.10 to 0.12 mg C.m^-3.h^-1; (3) the rate at which PDOC was respired to CO₂ was 0.001 to 0.003 mg C.m^-3.h^-1. (4) the PDOC makes up only about 0.1% of the total dissolved organic carbon. The size class of particles associated with PDOC production differed from the size class responsible for uptake of PDOC. More than 50% of the PDOC production was associated with particles having a nominal diameter range of 20 to 63 m, while this fraction was responsible for <10% of the incorporation.     

Rapid degradation of carbon tetrachloride by commercial micro-scale zinc powder assisted by citric acid

We developed an effective method for degradation of carbon tetrachloride (CT) in contaminated water. Zinc metal as a reducing agent for CT in aqueous solutions has been previously studied in some detail, but the rapid corrosion of zinc surface usually reduces its efficiency in removing CT. We assumed that citric acid could enhance the degradation of CT by zinc powder due to the elimination of a passivation layer of Zn(II) (hydr)oxides on the surface of zinc powder through chelating of organic ligands with Zn(II) produced from the reaction and keeping the exposure of active sites to targets. Here the influence of citric acid on the decomposing of CT by commercial micro-scale zinc powder was investigated in a pH range of 3.5–7.5 at 25°C in batch experiments. Reaction mixtures were analysed by gas chromatography/headspace analysis, and Cl⁻ concentration was determined by turbidimetry. The results demonstrate that the degradation of CT by zinc metal alone is very weak, but the addition of citric acid can assist zinc powder to decompose CT more completely and rapidly at all pHs. Degradation of CT took place mainly in the first 10 min of reaction, coupled with 75–95% of CT removal. Maximum dechlorination percentage (82.4%) of CT was obtained at pH 5.5. In that case, chloroform and dichloromethane, as main intermediates, were found at low levels during the whole reaction, suggesting that CT may be sequentially and multiply degraded so quickly that methane is yielded before the intermediates can be desorbed and released to aqueous solution. When compared with the current methods of nano-scale zinc and bimetallic systems, the application of commercial micro-scale zinc particles assisted by organic ligands is of environmental significance since it allows decontamination of aqueous chlorinated organic compounds at low cost and with high efficiency.