2-D DIGE and MALDI-TOF-MS analysis of the serum proteome in human osteosarcoma

We performed 2-D DIGE on proteins prepared from serum obtained from patients with osteosarcoma (OS) and controls, to identify differentially expressed proteins that might serve as serum biomarkers for OS prognosis. Proteins found to be differentially expressed were identified by MALDI-TOF mass spectrometric analysis, coupled with database interrogation. We compared serum samples from four individuals with OS to four age- and sex-matched healthy controls. We identified 24 protein spot-features that were significantly increased, and 34 that were significantly decreased in serum from patients with OS relative to the controls. The MS analysis revealed 18 unique proteins that were increased, and 25 unique proteins that were decreased in OS serum samples. Western blot and ELISA analysis confirmed increased levels of amyloid-related serum protein (SAA) in the OS serum samples. The increased expression levels of SAA were decreased after using MTX and cisplatin combination chemotherapy, and were further decreased after operation. Moreover, increased expression levels of sera SAA were seen in the relapsed patients. Our results suggested that the determination of serum SAA in OS patients might be utilized as a marker for relapse and in evaluation of the efficacy of therapy.     

Protein-induced changes during the maturation process of human dendritic cells: A 2-D DIGE approach

Dendritic cells (DCs) are unique antigen presenting cells, which upon maturation change from a specialized antigen‐capturing cell towards a professional antigen presenting cells. In this study, a 2‐D DIGE analysis of immature and mature DCs was performed, to identify proteins changing in expression upon maturation. The protein expression profile of immature and mature DCs, derived from CD14⁺ peripheral blood monocytes was investigated using two pH ranges (pH 4–7 and 6–9) (n = 4). Ninety one differentially expressed spots (p<0.01) were detected, from which we identified 74 spots (81.32%) corresponding to 41 different proteins. The proteins identified play a role in diverse processes, such as antigen processing/presentation, vesicle transport and cytoskeleton remodeling. In addition, a protein interaction network contained 29 (out of 41) proteins, suggesting that, although they functionally originate from distinct classes, these proteins are acting as a protein‐interactome. In conclusion, the proteins shown here to be altered in expression upon maturation are in line with the morphological and functional changes observed during the maturation process, providing a better understanding of the processes involved. This will open new avenues for investigating treatment regimens for immune‐associated disorders.     

State of the art of 2D DIGE

Difference gel electrophoresis enables the accurate quantification of changes in the proteome including combinations of PTMs and protein isoform expression. Here, we review recent advances in study design, image acquisition, and statistical analysis. We also compare DIGE to established and emerging mass spectrometric analysis technologies. Despite these recent advances in MS and the still unsolved limitations of 2DE to map hydrophobic, high molecular weight proteins with extreme pIs, DIGE remains the most comprehensive top-down method to study changes in abundance of intact proteins.     

An optimum method designed for 2-D DIGE analysis of human arterial intima and media layers isolated by laser microdissection

The formation and progression of atherosclerotic lesions involve complex mechanisms which are still not fully understood. A variety of cell types from the distinct arterial layers are implicated in the whole process from lipid accumulation within the vascular wall to plaque development and final rupture. In the present work, we employ the combination of laser microdissection and pressure catapulting and 2-D DIGE saturation labeling to investigate the human intima and media sub-proteomes isolated from atherosclerotic (coronary and aorta) or non-atherosclerotic vessels (preatherosclerotic coronary arteries). Laser microdissection and pressure catapulting allows the specific isolation of regions of interest. In turn, DIGE saturation labeling overcomes the limitation of extensive microdissection times to recover the protein amount required to perform comparative 2-DE, particularly when dealing with tissue regions rich in myofilament proteins, which result in low protein recovery. The compatibility and optimum performance of both techniques were investigated in detail, paying special attention to tissue staining and protein solubilization. Since scarce amount of protein obtained from microdissected tissue made it impossible to directly perform protein identification from 2-DE spots by MS, we performed in-solution digestion followed by LC-MS/MS analysis of total protein extracts from intima and media in order to get an overall picture of protein composition. Proteins so identified confirm the nature of the isolated regions. Finally, similar spot resolution on 2-D DIGE gels was obtained for the different human artery types (coronary, aorta) and studied layers (intima, media), setting the basis for future clinical comparative studies.     

Effects of endurance exercise on the urinary proteome analyzed by 2-D PAGE and Orbitrap MS

Purpose : Exercise‐induced proteinuria is a well‐known phenomenon and the influence of parameters such as intensity and duration was studied extensively. Usually, total protein or albumin was measured for diagnosis of a proteinuria, and the present study was performed to search for qualitative differences in the urinary proteome before and after endurance exercise. Experimental design : Urine samples were concentrated and proteins separated by means of 2‐D PAGE. Proteins differing in the investigated groups were identified by nano‐UPLC‐Orbitrap MS after trypsin digestion. Results : The study yielded several proteins such as hemopexin, albumin, orosomucoid 1, transferrin or carbonic anhydrase 1 that were elevated after a marathon run in comparison to a control group. These are linked to physiological changes resulting from endurance exercise such as destruction of erythrocytes or increased fat metabolism. On the contrary, 2‐D PAGE profiles of athletes at rest did not differ from those of control samples. Conclusions and clinical relevance: The study is a starting point to build up individual 2‐D PAGE protein maps of athletes. Further studies will investigate intra‐individual differences and further exercise parameters, which potentially lead to a physiological monitoring system for athletes in training and competition and may also complement the blood passport in doping control.     

基于平面图像实现三维仿真场景的技术方法研究

利用Windows操作系统平台正的DirectX多媒体接口技术,本文提出了利用平面图像实现三维仿真场景的相关实用技术方法。     

2-D Roesser模型的静态干扰解耦

本文讨论了2-D Roesser模型[1](RM)的静态干扰解耦问题[2](简称为2-D DDP),即寻求2-D状态反馈使相应的闭环系统具有抗干扰的能力,得到了问题有解的充分条件和计算相应反馈阵的算法.     

Considerations for powering a clinical proteomics study: Normal variability in the human plasma proteome

Proteomics is increasingly being applied to the human plasma proteome to identify biomarkers of disease for use in non‐invasive assays. 2‐D DIGE, simultaneously analysing thousands of protein spots quantitatively and maintaining protein isoform information, is one technique adopted. Sufficient numbers of samples must be analysed to achieve statistical power; however, few reported studies have analysed inherent variability in the plasma proteome by 2‐D DIGE to allow power calculations. This study analysed plasma from 60 healthy volunteers by 2‐D DIGE. Two samples were taken, 7 days apart, allowing estimation of sensitivity of detection of differences in spot intensity between two groups using either a longitudinal (paired) or non‐paired design. Parameters for differences were: two‐fold normalised volume change, α of 0.05 and power of 0.8. Using groups of 20 samples, alterations in 1742 spots could be detected with longitudinal sampling, and in 1206 between non‐paired groups. Interbatch gel variability was small relative to the detection parameters, indicating robustness and reproducibility of 2‐D DIGE for analysing large sample sets. In summary, 20 samples can allow detection of a large number of proteomic alterations by 2‐D DIGE in human plasma, the sensitivity of detecting differences was greatly improved by longitudinal sampling and the technology was robust across batches.     

Impact of incomplete DNase I treatment on human macrophage proteome analysis

The aim of our study was to analyze the proteomic pattern of human macrophages obtained over a 4 year period from blood donors. The purpose was to simulate a long-term clinical study to assess the application of 2-D DIGE technique for differential proteomic analysis of these scarce samples. Bioinformatic analysis of 2-D DIGE gels of 19 different cultures of macrophages assessed whether they did or did not contain at least specific five spots identified by MS as being or containing bovine deoxyribonuclease I (DNase I). Bovine DNase I was used during sample treatment to remove nucleic acids from protein extracts. Macrophages were classified in two groups, which appeared to be differentiated by the completeness of DNase I treatment. Further detailed analysis revealed a different proteomic pattern of macrophage protein samples according to the completeness of this treatment. The major group of proteins affected, accounting for one third of the differentially expressed proteins, included proteins involved in cell motion and actin cytoskeleton reorganization. The use of DNase I for the removal of nucleic acids from protein samples must be avoided in proteomic studies since it can generate bias in the analysis of protein expression patterns.     

卧式二维智能测头的设计

设计了一种在水平坐标面内测量的高精度二维智能测头;论述了采用硬件和软件结合方法设计智能测头的总体方案。测头采用两组平行片簧组合作为微位移机构,精密差动传感器作为检测器件,根据测头的动态数学模型对系统状态估值和实际测值比较判断测头与工件接触状态,实现了测头工作的自适应控制,保证了精度与快速性要求。静态和动态试验结果证明:该测头技术指标达到了设计要求。     

Multi-edge detection by isotropical 2-D ISEF cascade

Most research in edge detection has been based on the classical mono-edge model. In the present paper, we propose an optimal edge detector for complicated noisy multi-edge images and the isotropical 2-D realization of the optimal filter. We introduce at first the complicated multi-edge model and show that the optimal filter as a preparation for multi edge detection, is a cascade of Infinite size Symmetrical Exponential Filters (ISEF). We then propose a fast realization of the isotropical 2-D optimal filter. Experimental results are also presented.     

二维片上网络局部均匀随机通信性能分析

作为对全局均匀随机通信二维片上网络性能分析的延续和深入,首先描述了全局均匀随机通信模式和局部均匀随机通信模式的数学模型,分析了二者的关系;然后用链路数表示通信成本,基于作者独立设计的片上网络路由与通信协议,分析了不同结构和规模各结构网络性能随局部通信概率变化而变化的规律,并依据几种结构的性能相互关系及结构特点对它们进行了简单分类.结果表明,全局均匀随机通信模式其实是局部均匀随机通信模式的特例,随着局部通信概率的增大,各种结构的网络性能逐步提高;相比较而言,四边形、三角形网眼Mesh网络及其变形结构更适合于在本地通信概率较小或者通信密集型的应用,而当本地通信概率较大或者通信强度较低的情况下应用六边形网眼Mesh及其变形结构、多环相切及其回绕结构可能会取得更好的综合性能.     

基于Legendre多项式展开的双基SAR二维频域成像算法

针对任意构型双基SAR基于传统Taylor级数展开斜距方法的边缘点散焦问题,提出了一种基于Legendre正交多项式逼近的双基SAR二维频域成像算法。该算法基于对斜距函数的Legendre多项式展开推导了点目标二维频谱,解除了回波信号距离—方位的耦合,使成像处理更高效。在二维频域对场景区域的点目标采用二维频域成像算法进行成像。该算法改善了场景边缘点的聚焦效果,增加了场景边缘点的聚焦深度。理论推导和仿真结果验证了该算法的有效性和可行性。     

2-D DIGE identification of differentially expressed heterogeneous nuclear ribonucleoproteins and transcription factors during neural differentiation of human embryonic stem cells

Neural stem cells (NSC) are progenitors that can give rise to all neural lineages. They are found in specific niches of fetal and adult brains and grow in vitro as non-adherent colonies, the neurospheres. These cells express the intermediate filament nestin, commonly considered an NSC marker. NSC can be derived as neurospheres from human embryonic stem cells (hESC). The mechanisms of cellular programming that hESC undergo during differentiation remain obscure. To investigate the commitment process of hESC during directed neural differentiation, we compared the nuclear proteomes of hESC and hESC-derived neurospheres. We used 2-D DIGE to conduct a quantitative comparison of hESC and NSC nuclear proteins and detected 1521 protein spots matched across three gels. Statistical analysis (ANOVA n = 3 with false discovery correction) revealed that only 2.1% of the densitometric signal was significantly changed. The ranges of average ratios varied from 1.2- to 11-fold at a statistically significant p-value <0.05. MS/MS identified 15 regulated proteins previously shown to be involved in chromatin remodeling, mRNA processing and gene expression regulation. Notably, three members of the heterogeneous nuclear ribonucleoprotein family (AUF-1, and FBP-1 and FBP-2) register a 54, 70 and 99% increased expression, highlighting them as potential markers for NSC in vitro derivation. By contrast, Cpsf-6 virtually disappears with differentiation with an 11-fold drop in NSC, highlighting this protein as a novel marker for undifferentiated ESC.     

关于第二类Fornasini-Marchesini模糊的2-D动态规划

为了求解用正常或奇异的第二类Fornasini-Marchesini模型(FMM Ⅱ)描述的2-D线性离散系统的最优控制问题, 首先将2-D FMM Ⅱ用变结构1-D形式表示, 再利用1-D动态规划的方法, 给出使给定指标函数最小的2-D控制序列、最优轨线及性能指标的最优值的计算方法.     

Duality of 2-D singular systems of Roesser models

In this paper, the properties and concepts of dual systems of the two-dimensional singular Roesser models (2-D SRM) are studied. Two different concepts of the dual systems are proposed for the 2-D SRM. One is derived from the duality defined for two-dimensional singular general models (2-D SGM)-called the S-dual systems; the other one is defined based on 2-D SRM in a traditional sense-called the T-dual systems. It is shown that if a 2-D SRM is jump-mode free or jump-mode reachable, then it can be equivalently transformed into a canonical form of a 2-D SRM, for which the T-duality and the S-duality are equivalent. This will be of some perspective applications in the robust control of 2-D SRM.     

Duality of 2-D singular systems of Roesser models

In this paper, the properties and concepts of dual systems of the two-dimensional singular Roesser models （2-D SRM） are studied. Two different concepts of the dual systems are proposed for the 2-D SRM. One is derived from the duality defined for two-dimensional singular general models （2-D SGM）-called the S-dual systems; the other one is defined based on 2-D SRM in a traditional sense-called the T-dual systems. It is shown that if a 2-D SRM is jump-mode free or jump-mode reachable, then it can be equivalently transformed into a canonical form of a 2-D SRM, for which the T-duality and the S-duality are equivalent. This will be of some perspective applications in the robust control of 2-D SRM.     

二维可分Roesser模型的模能控(观)性

本文讨论了二维可分Roesser模型(RM)的模能控(观)性的判定问题,得出了相应的充要条件和数值稳定的算法,同时给出了该算法具有数值鲁棒性的充要条件和算法累积误差最大容许上界的显式估计,最后对二维可分RM的模能观性在状态空间中给出了一种几何解释.     

2-D系统模能控性和模能观性判据

本文指出并纠正了文［１～３］关于２－Ｄ系统模能控性和模能观性判据的错误。