RNA‑seq技术在水生生物生态毒理学中的应用进展

水域是地球环境的重要组成部分,也是最易受污染的生态系统之一。水生态系统中不同营养级别的水生生物可通过摄食、接触等多种途径摄入水体中的污染物。因此,监测水域污染物对水生生物和生态系统的影响,解析污染物对不同水生生物的毒性机制,筛选敏感、有效的生物标志物对生态毒理学研究和环境风险评价具有重要意义。RNA测序（RNA sequencing,RNA?seq）技术因所需样品量少,且不需参考序列,可在整体水平上鉴定基因差异表达,成为水生生物生态毒理学研究的最佳方法之一。基于此,介绍了RNA?seq技术的基本流程与数据分析过程,对该技术在不同生态位的水生生物（如鱼类、两栖类、贝类、甲壳类等）生态毒理学中的应用展开综述,并对RNA?seq技术面临的不足、挑战及发展趋势进行探讨,以期为该技术在水生生物生态毒理学研究中的应用,尤其是水生态环境中污染物胁迫水生生物机制的阐明及污染水域生态环境恢复提供参考。     

生态毒理基因组学和生态毒理蛋白质组学研究进展

将基因组学和蛋白质组学知识整合到生态毒理学中形成了生态毒理基因组学和生态毒理蛋白质组学.通过生态毒理基因组学和生态毒理蛋白质组学的研究能够在基因组和蛋白质组水平更深入理解毒物的作用机制,寻找更敏感、有效的生物标记物,形成潜在的强有力的生态风险评价工具.介绍了生态毒理基因组学和生态毒理蛋白质组学的研究进展,以及DNA芯片技术和2D-凝胶电泳技术在持久性有毒污染物的生态毒理学研究中的应用.     

环状RNA及其在生态毒理学研究中的应用进展

环状RNA（circular RNAs,circRNAs）是一类共价闭合环状非编码RNA,具有进化上保守、结构上稳定、组织特异性表达等特点。CircRNAs可作为miRNAs海绵影响其对基因的调控,还可与RNA结合蛋白（RNA binding proteins,RBPs）相互作用,也有研究表明某些circRNAs还具有被翻译成蛋白质的潜能。CircRNAs已被证实对某些疾病具有特异性、稳定性和调节功能,如癌症、糖尿病、心血管疾病、神经退行性疾病等,其可作为潜在的诊断、预后生物标志物和治疗靶点。最近,有研究发现circRNAs参与了环境化学污染物诱导的毒性效应发生及发展的过程。目前,生态毒理学研究中评价环境化学污染物和毒效应之间关系的毒性终点通常会受遗传多态性和表观遗传学影响,考虑到经环境化学污染物暴露后生物体内circRNAs差异性表达的现象,或许在生态毒理学研究中circRNAs也有作为生物标志物的可能性。基于此,对circRNAs的生物合成与降解、生物学功能、分析方法及其目前在生态毒理学研究中的应用展开综述,并对其作为分子生物标志物在环境污染物暴露早期诊断和生态风险评价中的应用进行了展望,以期为生态毒理学研究和环境风险评价提供参考。     

动物生物标志物在土壤污染生态学研究中的应用

应用陆栖无脊椎动物的生物标志物对土壤生态系统中污染物的暴露和效应进行评价日益受到重视,并取得了显著的研究进展,文中介绍了溶酶体、胁迫蛋白和金属硫蛋白(MTs)3种主要生物标志物,体腔细胞内溶酶体膜稳定性用中性红保持时间(NRR)进行检测;胁迫蛋白类多采其中的Hsp70和Hsp60;金属硫蛋白不同同分异构体的定量分析可用于反映不同的金属污染胁迫,对3种生物标志物机理、特性、检测实例以及在污染土壤生态毒理诊断中的应用前景进行了评述。     

重金属污染评估及其生物健康效应

重金属污染是目前全球面临的最为严峻的环境问题之一,不仅引发了粮食安全危机,还直接危害着动物和人类健康.土壤线虫作为陆地环境中重金属污染的重要指示生物,不仅会对重金属的毒害产生复杂的防御机制,其肠道微生物还被证明可以协助宿主抵御重金属胁迫.本文系统梳理了国内外重金属的污染现状及其评估方法,以线虫为模型概述了重金属对土壤动物行为、生理、生化三个方面的影响,总结了动物对重金属暴露的应激响应,主要包括细胞壁和细胞膜的屏障作用、活性物质改变、金属硫蛋白表达以及非编码RNA调节等四个层面,提出了肠道微生物的组成和多样性作为指示重金属毒性的新方法,归纳了长期重金属暴露导致的生物适应性进化,包括重金属超积累基因、抗性基因、解毒基因的出现和富集等,进而量化环境胁迫引起的动物遗传变异在基因组水平上的响应,比如基因的抗性突变和抗性基因的协同作用.最后,本文对重金属生态毒理学的研究进行了展望,指出未来研究需着眼于当下的生态环境和人类健康,探索土壤动物肠道微生物的生态作用,深入探究重金属毒性发生发展以及生物解毒的潜在机制.     

土壤污染的生态毒理诊断研究进展

土壤生态毒理学是污染土壤生态风险评价以及土壤污染控制的理论依据,土壤环境中有毒物质的生态毒理效应及其分子机制的研究是土壤生态毒理学的核心内容.土壤污染生态毒理诊断集合了土壤污染的全部信息,提供了土壤的整体毒性效应.因此,开展土壤污染生态毒理诊断研究有重要的理论意义和现实意义.本文在参考国内外土壤污染生态毒理诊断研究成果的基础上,介绍了常用土壤污染生态毒理效应诊断方法和指标,并探讨了不同方法的利弊.传统毒理诊断方法往往只能表征胁迫的程度,不能解释损伤和响应发生分子机理.污染土壤毒性评估的生物标记物和土壤污染分子诊断技术,可深入探讨外源胁迫下生物的解毒机制.本文对其进行了较为系统的介绍,并对未来的研究趋势进行展望.     

蚯蚓在环境安全研究中的应用

蚯蚓在环境污染的生态毒理学诊断方面具有重要生态学意义,这是蚯蚓在环境安全研究中应用的基础。本文概述了蚯蚓生态毒理诊断的一些试验方法,包括急性毒性试验法、田间生态毒性试验法、污染环境的生物检测法和微观生物指标检测法。在这基础上,就蚯蚓的环境安全生态指示研究进展进行了剖析与展望。还从蚯蚓处理生活垃圾与农业有机废弃物以及污水的蚯蚓过滤处理等应用方面,论述了蚯蚓在污染环境解毒过程中的重要生态作用及今后的发展方向。     

蛋白质组学在贝类研究中的应用

随着人类基因组计划的完成,功能基因组学的研究日益蓬勃发展。作为功能基因组学的重要支柱,蛋白质组学的研究旨在阐明基因组所表达的真正执行生命活动的全部蛋白质的表达规律和生物功能。伴随着研究的不断深入,蛋白质组学技术已涉及水产经济动物如鱼、虾、贝的研究。尽管在水产动物的研究中运用蛋白质组学技术较晚,但进展迅速。目前对于贝类相关蛋白质组学的研究尚未有一系统总结。因此,基于贝类蛋白质组学中双向电泳体系的构建和优化、生物矿化、发育与免疫和环境毒理学的相关研究做一综述。     

蛋白质组学在研究植物响应逆境机理上的应用

逆境条件下植物可以通过改变其基因表达和相关代谢活动来适应,探讨植物基因和蛋白表达谱的变化就成为植物逆境响应机制研究中的重要内容,蛋白质表达谱反映了植物细胞和组织的实际状态,是植物基因表达和最终代谢的关键环节。随着蛋白质分离技术、质谱鉴定技术和植物生物信息学的迅速发展,蛋白质组学在植物响应逆境方面的研究中的应用已经比较成功,加深了人们对植物响应逆境机制的认识,并为人们提供了新的线索和思维。本文主要对蛋白质组学在植物响应非生物逆境(干旱、盐胁迫、低温胁迫、高温胁迫等)和生物逆境(病虫害)的机制研究的应用上进行了综述。     

轮虫生态毒理学测试指标的敏感性研究进展

随着工业化的发展,人类活动对环境造成了严重污染.轮虫是常用于污染物毒性检测的模式生物之一.针对不同类型的污染物,轮虫各生态毒理学指标的敏感性不尽相同.因此,本文归纳了现有的轮虫生态毒理学测试指标,综述了各生态毒理学检测指标对不同污染物的敏感性:轮虫的生命表统计学参数和生理生化指标对重金属污染通常更敏感;在检测持久性有机...     

Environmental proteomics and metallomics

Monitoring environmental pollution using biomarkers requires detailed knowledge about the markers, and many only allow a partial assessment of pollution. New proteomic methods (environmental proteomics) can identify proteins that, after validation, might be useful as alternative biomarkers, although this approach also has its limitations, derived mainly from their application to non-model organisms. Initial studies using environmental proteomics were carried out in animals exposed to model pollutants, and led to the concept of protein expression signatures. Experiments have been carried out in model organisms (yeast, Arabidopsis, rat cells, or mice) exposed to model contaminants. Over the last few years, proteomics has been applied to organisms from ecosystems with different pollution levels, forming the basis of an environmental branch in proteomics. Another focus is connected with the presence of metals bound to biomolecules, which adds an additional dimension to metal-biomolecule and metalloprotein characterization - the field of metallomics. The metallomic approach considers the metallome: a whole individual metal or metalloid species within a cell or tissue. A metallomic analytical approach (MAA) is proposed as a new tool to study and identify metalloproteins.     

Selection of reference genes for quantitative RT-PCR studies in striped dolphin (Stenella coeruleoalba) skin biopsies

Background  

Odontocete cetaceans occupy the top position of the marine food-web and are particularly sensitive to the bioaccumulation of lipophilic contaminants. The effects of environmental pollution on these species are highly debated and various ecotoxicological studies have addressed the impact of xenobiotic compounds on marine mammals, raising conservational concerns. Despite its sensitivity, quantitative real-time PCR (qRT-PCR) has never been used to quantify gene induction caused by exposure of cetaceans to contaminants. A limitation for the application of qRT-PCR is the need for appropriate reference genes which allow the correct quantification of gene expression. A systematic evaluation of potential reference genes in cetacean skin biopsies is presented, in order to validate future qRT-PCR studies aiming at using the expression of selected genes as non-lethal biomarkers.     

Proteomic applications in ecotoxicology

Within the growing body of proteomics studies, issues addressing problems of ecotoxicology are on the rise. Generally speaking, ecotoxicology uses quantitative expression changes of distinct proteins known to be involved in toxicological responses as biomarkers. Unlike these directed approaches, proteomics examines how multiple expression changes are associated with a contamination that is suspected to be detrimental. Consequently, proteins involved in toxicological responses that have not been described previously may be revealed. Following identification of key proteins indicating exposure or effect, proteomics can potentially be employed in environmental risk assessment. To this end, bioinformatics may unveil protein patterns specific to an environmental stress that would constitute a classifier able to distinguish an exposure from a control state. The combined use of sets of marker proteins associated with a given pollution impact may prove to be more reliable, as they are based not only on a few unique markers which are measured independently, but reflect the complexity of a toxicological response. Such a proteomic pattern might also integrate some of the already established biomarkers of environmental toxicity. Proteomics applications in ecotoxicology may also comprise functional examination of known classes of proteins, such as glutathione transferases or metallothioneins, to elucidate their toxicological responses.     

A web-based tool for in silico biomarker discovery based on tissue-specific protein profiles in normal and cancer tissues

Here we report the development of a publicly available Web-based analysis tool for exploring proteins expressed in a tissue- or cancer-specific manner. The search queries are based on the human tissue profiles in normal and cancer cells in the Human Protein Atlas portal and rely on the individual annotation performed by pathologists of images representing immunohistochemically stained tissue sections. Approximately 1.8 million images representing more than 3000 antibodies directed toward human proteins were used in the study. The search tool allows for the systematic exploration of the protein atlas to discover potential protein biomarkers. Such biomarkers include tissue-specific markers, cell type-specific markers, tumor type-specific markers, markers of malignancy, and prognostic or predictive markers of cancers. Here we show examples of database queries to generate sets of candidate biomarker proteins for several of these different categories. Expression profiles of candidate proteins can then subsequently be validated by examination of the underlying high resolution images. The present study shows examples of search strategies revealing several potential protein biomarkers, including proteins specifically expressed in normal cells and in cancer cells from specified tumor types. The lists of candidate proteins can be used as a starting point for further validation in larger patient cohorts using both immunological approaches and technologies utilizing more classical proteomics tools.     

蚯蚓生物标记物在土壤生态风险评价中的应用

蚯蚓在土壤中行使了很多重要的生态功能,蚯蚓生物标记物常用作土壤污染风险评价研究。这篇综述的目的是探讨当前蚯蚓生物标记物研究是否可以应用到实际的土壤污染风险评价。1)讨论了蚯蚓生物标记物在土壤污染风险评价体系中的重要性,认为它是化学分析方法的有益补充,可以提供更为全面和客观的土壤污染信息;2)综述了相关研究中所使用的蚯蚓类型,土壤类型和生物标记物类型,及其它试验设计要素和最后结果的变异,认为目前蚯蚓生物标记物研究以实验室基础研究为主,筛选出了大量的生物标记物,一定程度上揭示了生物标记物的对各类典型污染物及其组合的应答机制;同时也认为,未来的蚯蚓生物标记物研究应该重点探讨将其应用到实际的土壤污染风险评价中的可行性及如何应用;3)目前不同研究之间从试验设计到结果都具有很大的变异,难以通过综合比较获得完全可靠的具有实践意义的结论和成果,因此,有必要通过建立标准化的蚯蚓生物标记物研究方法,推动生物标记物的研究工作;4)提出了蚯蚓生物标记物研究方法标准化的具体建议,推荐了蚯蚓生物标记物走向实际应用所需要解决的问题。     

Utility of proteomics to assess pollutant response of clams from the Doñana bank of Guadalquivir Estuary (SW Spain)

The utility of proteomics to assess pollutant response of Scrobicularia plana clams from three sites of Guadalquivir Estuary at the southern end of the National Park of Do?ana (SW Spain) has been studied. Protein expression profiles were analyzed by 2-DE in soluble fractions of S. plana gills. Nearly 2000 well-resolved spots were detected in silver-stained gels, with focused areas in the 4-6.5 pH range. Different protein expression signatures were found at each site, with the highest number of more intense spots in animals with the highest metal content. Nineteen more intense protein spots were analyzed out by nanospray-ion trap tandem mass spectrometry, de novo sequencing and a bioinformatics search for their possible identification. While sequence tags of 16 more intense protein spots were obtained, including several proteins induced by pollutant exposure of model organisms, only 2 proteins were unambiguously identified: hypoxanthine-guanine phosphoribosyltransferase (HPRT) and glyceraldehyde-3-phosphate dehydrogenase (G3PDH). Both enzymes were significantly higher in animals with the highest metal contents. Thus, we propose these two proteins, HPRT and G3PDH, as novel pollution biomarkers.     

Proteomics in clinical trials and practice: present uses and future promise

The study of clinical proteomics is a promising new field that has the potential to have many applications, including the identification of biomarkers and monitoring of disease, especially in the field of oncology. Expression proteomics evaluates the cellular production of proteins encoded by a particular gene and exploits the differential expression and post-translational modifications of proteins between healthy and diseased states. These biomarkers may be applied towards early diagnosis, prognosis, and prediction of response to therapy. Functional proteomics seeks to decipher protein-protein interactions and biochemical pathways involved in disease biology and targeted by newer molecular therapeutics. Advanced spectrometry technologies and new protein array formats have improved these analyses and are now being applied prospectively in clinical trials. Further advancement of proteomics technology could usher in an era of personalized molecular medicine, where diseases are diagnosed at earlier stages and where therapies are more effective because they are tailored to the protein expression of a patient's malignancy.     

Proteomic analysis of methyl parathion-responsive proteins in Sparus latus liver

The contamination of marine ecosystems by organophosphate pesticides is of great concern. The use of protein expression profiles may provide a good method to help us understand the methyl parathion (MP) toxicity to aquatic organisms. In this study, Sparus latus, was selected as the target organism. The toxicological effects of MP were investigated after 24 h exposure using proteomics to analyze their liver tissues. Certain enzyme activity parameters of the liver extracts were also examined, including CAT. After analyzing the proteomic profile of the liver using 2D gel electrophoresis, we found that the protein expression levels of 25 spots increased or decreased significantly in the exposed groups. Sixteen of the 25 protein spots were successfully identified using MALDI-TOF MS/MS. These proteins were roughly categorized into diverse functional classes such as cell redox homeostasis, metabolic processes and cytoskeleton system. These data demonstrated that proteomics was a powerful tool to provide valuable insights into the possible mechanisms of toxicity of MP contaminants in aquatic species. Additionally, these data may provide novel biomarkers for evaluation of MP contamination in the environment.     

Label-free quantification in clinical proteomics

Nowadays, proteomic studies no longer focus only on identifying as many proteins as possible in a given sample, but aiming for an accurate quantification of them. Especially in clinical proteomics, the investigation of variable protein expression profiles can yield useful information on pathological pathways or biomarkers and drug targets related to a particular disease. Over the time, many quantitative proteomic approaches have been established allowing researchers in the field of proteomics to refer to a comprehensive toolbox of different methodologies. In this review we will give an overview of different methods of quantitative proteomics with focus on label-free proteomics and its use in clinical proteomics.     

Clinical cancer proteomics: promises and pitfalls

Proteome analysis promises to be valuable for the identification of tissue and serum biomarkers associated with human malignancies. In addition, proteome technologies offer the opportunity to analyze protein expression profiles and to analyze the activity of signaling pathways. Many published proteomic studies of human tumor tissue are associated with weaknesses in tumor representativity, sample contamination by nontumor cells and serum proteins. Studies often include a moderate number of tumors which may not be representative of clinical materials. It is therefore very important that biomarkers identified by proteomics are validated in representative tumor materials by other techniques, such as immunohistochemistry. Proteome technologies can be used to identify disease markers in human serum. Tumor derived proteins are present at nanomolar to picomolar concentrations in cancer patient sera, 10(6)-10(9)-fold lower than albumin, and will give rise to correspondingly smaller spots/peaks in protein separations. This leads to the need to prefractionate serum samples before analysis. Despite various pitfalls, proteomic analysis is a promising approach to the identification of biomarkers, and for generation of protein expression profiles that can be analyzed by artificial learning methods for improved diagnosis of human malignancy. Recent advances in the field of proteomic analysis of human tumors are summarized in the present review.