A unified modeling framework for metabonomic profile development and covariate selection for acute trauma subjects

Acute trauma is often associated with progressive deterioration of multiple organ systems in humans and is the leading cause of death in trauma care units. Identification of specific organ failure in a non-invasive manner and the contribution of different demographic factors on the casual progression of acute trauma are of supreme interests for successful diagnosis, prognosis or monitoring of trauma status. Recently, electrospray ionization and matrix-assisted laser desorption time-of-flight mass spectrometry have been used to identify biomarkers in both proteomics and metabonomics studies. Data sets generated from mass spectrometers in such studies are generally very large in size and thus require the use of sophisticated statistical techniques to glean useful information. In a recent development, Ghosh et al. (BMC Bioinformatics 2008; 9:38) suggested a unified semiparametric approach to distinguish urinary metabolic profiles in a group of traumatic subjects from those of a control group consisting of normal individuals. In this study we have extended their approach by combining available covariate information in the development of metabonomic profile of acute trauma. We have shown that age is a statistically significant covariate across trauma and control group, thus pointing out the fact that prognosis of trauma may be acutely linked with subjects' age.     

质谱技术在蛋白质组研究中的应用

电喷雾电离 (ESI)和基质辅助激光解吸电离 (MALDI)是最近发展起来的生物质谱技术 ,本文概述生物质谱技术在蛋白质组研究中分子质量的测定、蛋白质全谱分析、肽指纹谱测定、肽序列测定等方面的应用。方法灵敏、准确、快速 ,而且可实现高通量和自动化。它们已成为蛋白质组研究的关键性支撑技术 ,对蛋白质组的研究和分析具有重要意义     

血清蛋白质图谱分析在卵巢癌早期诊断中的应用

低分子量血清蛋白的概况可以反映器官潜在的病理变化 ,有助于肿瘤的早期诊断。在质谱技术的基础上发展起来的表面加强激光解析及电离飞行时间质谱技术联合人工智能技术 ,可以对卵巢癌患者的血清进行分析从而得到蛋白质图谱 ,对血清蛋白质图谱分析可用于卵巢癌的筛查 ,有望成为卵巢癌新的诊断手段。     

Nutriproteomics: technologies and applications for identification and quantification of biomarkers and ingredients

Nutrition refers to the process by which a living organism ingests and digests food and uses the nutrients therein for growth, tissue maintenance and all other functions essential to life. Food components interact with our body at molecular, cellular, organ and system level. Nutrients come in complex mixtures, in which the presence and concentration of single compounds as well as their interactions with other compounds and the food matrix influence their bioavailability and bioefficacy. Traditionally, nutrition research mainly concentrated on supplying nutrients of quality to nourish populations and on preventing specific nutrient deficiencies. More recently, it investigates health-related aspects of individual ingredients or of complete diets, in view of health promotion, performance optimisation, disease prevention and risk assessment. This review focuses on proteins and peptides, their role as nutrients and biomarkers and on the technologies developed for their analysis. In the first part of this review, we provide insights into the way proteins are currently characterised and analysed using classical and emerging proteomic approaches. The scope of the second part is to review major applications of proteomics to nutrition, from characterisation of food proteins and peptides, via investigation of health-related food benefits to understanding disease-related mechanisms.     

结直肠癌有无肝转移血清差异蛋白质表达的研究

目的：利用蛋白质组学技术分离、鉴定结直肠癌有无肝转移患者血清中差异蛋白质,筛选诊断肝转移的血清蛋白标志物。方法：根据入组条件,收集结直肠癌无肝转移病例、有肝转移病例,在两组中随机抽取12例血清样本,同组血清等量混合进行双向凝胶电泳,建立两组血清蛋白质双向电泳图谱,用Image—MasterV5．0软件寻找两组差异蛋白质点,MALDI—TOF—MS对差异蛋白质进行鉴定,查询生物信息数据库对差异蛋白质进行初步分析。结果：两组比较差异在2倍以上蛋白质有8种,其中5个蛋白质表达上调,3个蛋白质表达下调;两组每个差异蛋白灰度体平均值比较差异均有统计学意义（P〈0．05）。通过数据库搜索鉴定出7种蛋白质,上调的5个蛋白质分别是Transferrin、Complement component C9、RN Adirected DNA polymerase（RNA指导的DNA合成酶）、Conserved hypothetical—protein（假定蛋白质）、SEC14L17 kDa protein;下调的2个蛋白质分别是Haptoglobin和Isoform 1 of Serumalbumin。结论：结直肠癌有无肝转移患者血清中的蛋白质表达谱有一定的差异性,这些差异蛋白质可能成为诊断肝转移的血清标志物。     

Celiac disease, inflammation and oxidative damage: a nutrigenetic approach

Celiac disease (CD), a common heritable chronic inflammatory condition of the small intestine caused by permanent intolerance to gluten/gliadin (prolamin), is characterized by a complex interplay between genetic and environmental factors. Developments in proteomics have provided an important contribution to the understanding of the biochemical and immunological aspects of the disease and the mechanisms involved in toxicity of prolamins. It has been demonstrated that some gliadin peptides resistant to complete proteolytic digestion may directly affect intestinal cell structure and functions by modulating gene expression and oxidative stress. In recent years, the creation of the two research fields Nutrigenomics and Nutrigenetics, has enabled the elucidation of some interactions between diet, nutrients and genes. Various dietary components including long chain ω-3 fatty acids, plant flavonoids, and carotenoids have been demonstrated to modulate oxidative stress, gene expression and production of inflammatory mediators. Therefore their adoption could preserve intestinal barrier integrity, play a protective role against toxicity of gliadin peptides and have a role in nutritional therapy of celiac disease.     

多种芯片检测和多元分析法在肝纤维化患者血浆蛋白表达谱分析中的初步应用

[目的]运用多种表面增强激光解吸-离子化飞行时间质谱（surface-enhanced laser desorption/ionization time of flight mass spectrometry,SELDI-TOF-MS）蛋白质芯片及多元分析方法寻找由乙型肝炎病毒（hepatitis B virus,HBV）感染引起的肝纤维化病程相关的血浆生物标志物。[方法]选用多种SELDI化学表面芯片,比较分析肝纤维化病人和正常血浆样本,筛选和确定3种最佳芯片类型。用这3种芯片分析无肝纤维化、轻度肝纤维化、重度肝纤维化和肝硬化4组共110例患者的血浆样本。运用主成分分析（PCA）、偏最小二乘回归（PLSR）、软独立建模分类法（SIMCA）等数据分析技术寻找差异蛋白。用聚类分析法研究差异蛋白的表达相似性。[结果]3种最适芯片类型分别是弱阳离子交换芯片（WCX2）、强阴离子交换芯片（SAX2）、固定化镍金属螯合亲和层析芯片（IMAC-Ni）。这3种芯片吸附的蛋白质种类互不相同,所发现的差异蛋白质峰也不同。经t检验分析,3种芯片共发现了20个差异蛋白峰。运用PCA、PLSR、SIMCA等数据分析技术,分别发现了105、98、62个差异峰,并对差异峰的重要性的可信度进行衡量。运用聚类分析技术,将差异蛋白的表达模式分组。[结论]联用多种SELDI芯片检测,结合多元分析方法,使SELDI技术成为筛选疾病相关的生物学标志物的有力工具。     

Genetic differences in the serum proteome of horses, donkeys and mules are detectable by protein profiling

Although horses and donkeys belong to the same genus, their genetic characteristics probably result in specific proteomes and post-translational modifications (PTM) of proteins. Since PTM can alter protein properties, specific PTM may contribute to species-specific characteristics. Therefore, the aim of the present study was to analyse differences in serum protein profiles of horses and donkeys as well as mules, which combine the genetic backgrounds of both species. Additionally, changes in PTM of the protein transthyretin (TTR) were analysed. Serum protein profiles of each species (five animals per species) were determined using strong anion exchanger ProteinChips? (Bio-Rad, Munich, Germany) in combination with surface-enhanced laser desorption ionisation-time of flight MS. The PTM of TTR were analysed subsequently by immunoprecipitation in combination with matrix-assisted laser desorption ionisation-time of flight MS. Protein profiling revealed species-specific differences in the proteome, with some protein peaks present in all three species as well as protein peaks that were unique for donkeys and mules, horses and mules or for horses alone. The molecular weight of TTR of horses and donkeys differed by 30 Da, and both species revealed several modified forms of TTR besides the native form. The mass spectra of mules represented a merging of TTR spectra of horses and donkeys. In summary, the present study indicated that there are substantial differences in the proteome of horses and donkeys. Additionally, the results probably indicate that the proteome of mules reveal a higher similarity to donkeys than to horses.     

Proteomics as a tool for the modelling of biological processes and biomarker development in nutrition research

Nutrition research has slowly started to adopt the proteomics techniques to measure changes in the protein complement of a biological system. This enables modelling of biological processes in response to dietary interventions, as well as the elucidation of novel biomarkers for health or disease that are sensitive to such interventions. There are limited studies on the effect of micronutrients on the proteome, so this review concentrates rather more on dietary intervention studies that have used proteomics (mainly classical 2D gel electrophoresis combined with mass spectrometry) to elucidate changes in pathways that relate to glucose and fatty acid metabolism, oxidative stress, anti-oxidant defence mechanisms and redox status. The ability to measure regulation of more low abundant proteins, such as those involved in inflammatory pathways, as well as the evaluation and validation of newly discovered candidate biomarkers in human biofluids, may depend on the introduction of more quantitative and sensitive methods like multiple reaction monitoring (MRM) and multiplexed immunoassays in nutrition research.     

Murine retrovirus infection and the effect of chronic alcohol consumption: proteomic analysis of cardiac protein expression

AIMS: The cardiovascular complications of acquired immunodeficiency syndrome (AIDS) are serious, including the occurrence of pathological heart conditions such as cardiomyopathy. Chronic alcohol consumption accentuates the severity of AIDS and may contribute to the development of cardiomyopathy. The aim of this work was to use a proteomics approach to investigate global alterations in protein expression in a mouse model of AIDS in the presence or absence of chronic alcohol consumption. METHODS: Cardiac proteins were separated by two-dimensional polyacrylamide gel electrophoresis and quantitative computer analysis was used to evaluate the resulting two-dimensional protein profiles. Proteins that were differentially expressed in the hearts of mice from the different experimental groups were identified by peptide mass finger-printing by matrix-assisted laser desorption/ionization mass spectrometry. RESULTS: A number of specific proteins were observed to be differentially expressed in the mouse heart due to the effect of ethanol feeding alone. Differentially expressed proteins were also observed that were due to viral infection alone. Ethanol feeding and viral infection appeared to have similar effects on the expression of a number of proteins. A total of 24 proteins were altered by infection alone. Of these 24 proteins, eight were affected by alcohol, with six alterations being ameliorated and two being exacerbated by alcohol. Two of these proteins have been identified as the 27 kDa heat-shock protein and mitochondrial long-chain acyl-CoA thioesterase 1. CONCLUSIONS: These results suggest that chronic alcohol consumption may exacerbate the effects of viral infection on the heart by lowering the stress response leading to de-protection and further cytotoxic effects.     

'Proteomics': the mapping of all human proteins

The genomes of many organisms, including humans, are now largely known. In the wake of this there is a need to identify and measure all proteins that are encoded by the genome (proteomics). This need leads to turbulent developments in the area of analytical techniques, such as two-dimensional electrophoresis, mass spectrometry, and protein chips. The rapidity of advancements justifies the expectation that in the next 5-10 years it will indeed become possible to determine the proteome of an organism or its components such as plasma, serum, or tissues. In combination with information on initiation and progress of disease, proteomics will contribute to improving health and to better primary and secondary prevention.     

Introduction of proteomic approach to environmental medicine

Recent progress in life science technology and the availability of much information on genes obtained by genome analysis has enabled us to analyze the changes of proteins on a large scale. Sets of proteins are called proteomes, and proteomics is the scientific field of proteome analysis including differential, post translational modification and interaction analyses. Various proteomic techniques, particularly two-dimensional gel electrophoresis (2-DE), mass spectrometry, protein chip methods, and surface plasmon resonance (SPR), are very useful for acquiring proteomes in cells, tissues and body fluid, and for analyzing interactions between a protein and other biofactors including proteins. A proteomic approach is also useful for determining biomarkers of diseases and key proteins involved in various stages of metabolism such as differentiation, cell cycle and apoptosis. Environmental pollutants including endocrine disruptors inhibit activities of various organs in wild animals and humans. Proteomic approaches could be very useful tools for elucidating the mechanisms of damage caused by environmental pollutants. In this review, we describe the application of a proteomic approach to the field of environmental medicine.     

Prioritization of biomarker targets in human umbilical cord blood: identification of proteins in infant blood serving as validated biomarkers in adults

Background: Early diagnosis represents one of the best lines of defense in the fight against a wide array of human diseases. Umbilical cord blood (UCB) is one of the first easily available diagnostic biofluids and can inform about the health status of newborns. However, compared with adult blood, its diagnostic potential remains largely untapped.Objectives: Our goal was to accelerate biomarker research on UCB by exploring its detectable protein content and providing a priority list of potential biomarkers based on known proteins involved in disease pathways.Methods: We explored cord blood serum proteins by profiling a UCB pool of 12 neonates with different backgrounds using a combination of isoelectric focusing and liquid chromatography coupled with matrix-assisted laser desorption/ionization tandem mass spectrometry (MALDI-MS/MS) and by comparing results with information contained in metabolic and disease databases available for adult blood.Results: A total of 1,210 UCB proteins were identified with a protein-level false discovery rate of ~ 5% as estimated by naïve target-decoy and MAYU approaches, signifying a 6-fold increase in the number of UCB proteins described to date. Identified proteins correspond to 138 different metabolic and disease pathways and provide a platform of mechanistically linked biomarker candidates for tracking disruptions in cellular processes. Moreover, among the identified proteins, 38 were found to be approved biomarkers for adult blood.Conclusions: The results of this study advance current knowledge of the human cord blood serum proteome. They showcase the potential of UCB as a diagnostic medium for assessing infant health by detection and identification of candidate biomarkers for known disease pathways using a global, nontargeted approach. These biomarkers may inform about mechanisms of exposure–disease relationships. Furthermore, biomarkers approved by the U.S. Food and Drug Administration for screening in adult blood were detected in UCB and represent high-priority targets for immediate validation.     

Proteomic analysis reveals changes in the liver protein pattern of rats exposed to dietary folate deficiency

Epidemiologic and experimental studies showed that folate deficiency is associated with increased risk of degenerative diseases by enhancing abnormal one-carbon metabolism. We studied the changes in the proteome of liver, the main tissue of folate storage and metabolism, in a rat model of dietary folate depletion. Four-month-old rats were fed for 4 wk an amino acid-defined diet without folate and compared with pair-fed rats given the same diet adequately supplemented with folic acid. Folate deprivation decreased plasma and hepatic folate concentrations dramatically, while increasing homocysteinemia significantly. Using 2-dimensional electrophoresis and matrix-assisted laser desorption/ionization time-of-flight MS, we identified 9 spots corresponding to differentially expressed proteins in the liver of folate-deficient rats compared with controls. Among those spots, 4 had a significantly increased volume, whereas the volume of the 5 other spots was decreased. Upregulated proteins included glutathione peroxidase (GPx) 1 and peroxiredoxin 6, 2 enzymes involved in the response to oxidative stress, and MAWD binding protein (MAWDBP), which has been associated with cancer. MAWDBP was simultaneously identified as a second spot with a lower isoelectric point (pI) that vanished almost completely after folate deficiency. Decreased abundance was also observed for cofilin 1, a protein linked to tumorigenesis, and for the GRP 75 precursor and preproalbumin, both of which are responsive to oxidative stress and/or inflammation. Moreover, an enzyme activity assay and/or Western blot analysis of GPx-1 and MAWDBP confirmed the proteomic findings. Our results show that folate deficiency modifies the abundance of several liver proteins consistently with adaptive tissue responses to oxidative and degenerative processes.     

Dietary proteins as determinants of metabolic and physiologic functions of the gastrointestinal tract

Dietary proteins elicit a wide range of nutritional and biological functions. Beyond their nutritional role as the source of amino acids for protein synthesis, they are instrumental in the regulation of food intake, glucose and lipid metabolism, blood pressure, bone metabolism and immune function. The interaction of dietary proteins and their products of digestion with the regulatory functions of the gastrointestinal (GI) tract plays a dominant role in determining the physiological properties of proteins. The site of interaction is widespread, from the oral cavity to the colon. The characteristics of proteins that influence their interaction with the GI tract in a source-dependent manner include their physico-chemical properties, their amino acid composition and sequence, their bioactive peptides, their digestion kinetics and also the non-protein bioactive components conjugated with them. Within the GI tract, these products affect several regulatory functions by interacting with receptors releasing hormones, affecting stomach emptying and GI transport and absorption, transmitting neural signals to the brain, and modifying the microflora. This review discusses the interaction of dietary proteins during digestion and absorption with the physiological and metabolic functions of the GI tract, and illustrates the importance of this interaction in the regulation of amino acid, glucose, lipid metabolism, and food intake.     

Matrix-assisted laser desorption ionization

Since its invention, matrix-assisted laser desorption ionization (MALDI) has found wide application in mass spectrometry of high molecular weight compounds such as synthetic polymers and biopolymers. Despite widespread application of MALDI, the fundamental processes of ion formation and desorption are still poorly understood. The chemistry of the MALDI process, occurring both during sample preparation and during ionization is reflected in the mass spectrum. As the MALDI technique now stands a low concentration of analyte molecules, which usually exhibit only moderate absorption per molecules, is embedded in matrix crystals consisting of a small, highly absorbing species. In this manner the efficient and controllable energy transfer is retained while the analyte molecules are separated from excessive energy that would lead to their decomposition. The matrix is believed to serve two major functions: adsorption of energy from the laser light and the isolation of analyte molecules from each other. There are 3 major methods for the preparation of samples for analysis which are quite quick and simple: dried droplet, surface and sandwich preparation. Experiments with pH indicator dyes serve as proof that analyte's charge state in the matrix crystals is the same as in solution. Upon laser desorption a sudden and explosive phase transition occurs and a dense plume of desorbed material is formed. The initial velocity of analyte ions in the plume depends only on the matrix used. Initial species formed as a result of laser desorption are tiny clusters. They consist of a matrix, analyte and other ionic species embedded in the matrix crystals all held together by hydrogen bonds and coulombic interactions. The first essential charging and thus ionization process is the statistical occurrence of clusters with a deficit/excess of anions of cations. Very small initial clusters are likely to be highly charged. Highly charged initial clusters cannot survive in the matrix plume and their charge drops. Clusters shrink by evaporation of neutral molecules. This paper presents only those cases leading to analyte ions and compares MALDI and the electrospray ionization technique.     

四氯化碳诱发大鼠肝脏蛋白质表达变化的分析

目的分析四氯化碳(CCl4)诱发肝纤维化大鼠肝脏细胞蛋白表达的变化,为寻找CCl4致肝纤维化早期生物效应的相关蛋白提供实验基础。方法采用皮下注射CCl4诱发实验性大鼠肝损伤模型,测定大鼠血清生化指标和观察肝组织病理学的变化,提取大鼠肝细胞总蛋白并采用二维凝胶电泳(2-DE)进行分离,Image Master 2D Platinum软件分析电泳图谱,选择表达有显著差异的蛋白点进行基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)测定。结果大鼠血清生化指标和病理学结果提示肝纤维化造模成功,CCl4损伤组肝细胞蛋白质2-DE图谱表达发生改变,差异点经MALDI-TOF-MS分析和数据库搜索,鉴定出4个已知序列蛋白。结论应用蛋白质组学技术筛选和鉴定出的蛋白可能与肝纤维化的发生发展有关,为进一步研究CCl4诱发肝纤维化机制提供依据。     

Translational regulation in nutrigenomics

The emergence of genome-wide analysis to interrogate cellular DNA, RNA, and protein content has revolutionized the study of the control network that mediates cellular homeostasis. Nutrigenomics addresses the effect of nutrients on gene expression, which provides a basis for understanding the biological activity of dietary components. Translation of mRNAs represents the last step of genetic flow and primarily defines the proteome. Translational regulation is thus critical for gene expression, in particular, under nutrient excess or deficiency. Until recently, it was unclear how the global effects of translational control are influenced by nutrient signaling. An emerging concept of translational reprogramming addresses how to maintain the expression of specific proteins during pathophysiological conditions by translation of selective mRNAs. Here we describe recent advances in our understanding of translational control, nutrient signaling, and their dysregulation in aging and cancer. The mechanistic understanding of translational regulation in response to different nutrient conditions may help identify potential dietary and therapeutic targets to improve human health.     

应用基质辅助激光解吸附电离飞行时间质谱分析宫颈鳞癌早期浸润蛋白质

目的探讨基质辅助激光解吸附电离飞行时间质谱(matrix assisted laser desorption ionization ti me of flight mass spectrometry,MALDI-TOF-MS)对宫颈鳞癌(squamous carcinoma of thecervix,SCC)组织切片直接分析,获取宫颈鳞癌早期浸润的分子标志物的有效性。方法利用超微量点样枪将化学基质直接点样到宫颈鳞癌组织切片上,然后利用基质辅助激光解吸附电离飞行时间质谱直接扫描宫颈鳞癌组织切片,分析宫颈鳞癌组织蛋白质分子。结果获得的25种蛋白质质谱峰强度比较,差异有显著意义(P<0.05)。其中7种差异蛋白质有分类意义,其质/核(M/Z)分别为:m/z3433,m/z4786,m/z10092,m/z10835,m/z15114,m/z15854和m/z16345。差异蛋白质m/z3433,m/z15114,m/z15854在宫颈鳞癌鳞状上皮组织高度表达,可作为宫颈鳞癌早期诊断的标志蛋白;差异蛋白质m/z16345在宫颈鳞癌的间质组织高度表达,可作为宫颈鳞癌早期间质浸润和扩散诊断的标志蛋白。结论利用基质辅助激光解吸附电离飞行时间质谱技术成功获取一组差异蛋白质。该差异蛋白组可望作为宫颈鳞癌早期诊断的标志蛋白。