A proteomic approach combining MS and bioinformatic analysis for the detection and identification of biomarkers of administration of exogenous human growth hormone in humans

An integrated MS-based proteomic approach is described that combines MALDI-MS and LC-MS with artificial neural networks for the identification of protein and peptide biomarkers associated with recombinant human growth hormone (rhGH) administration. Serum from exercised males administered with rhGH or placebo was analysed using ELISA to determine insulin-like growth factor-I concentrations. Diluted serum from rhGH- and placebo-treated subjects was analysed for protein biomarkers by MALDI-MS, whereas LC-MS was used to analyse tryptically digested ACN-depleted serum extracts for peptide biomarkers. Ion intensities and m/z values were used as inputs to artificial neural networks to classify samples into rhGH- and placebo-treated groups. Six protein ions (MALDI-MS) correctly classified 96% of samples into their respective groups, with a sensitivity of 91% (20 of 22 rhGH treated) and specificity of 100% (24 of 24 controls). Six peptide ions (LC-MS) were also identified and correctly classified 93% of samples with a sensitivity of 90% (19 of 21 rhGH treated) and a specificity of 95% (20 of 21 controls). The peptide biomarker ion with the highest significance was sequenced using LC-MS/MS and database searching and found to be associated with leucine-rich α-2-glycoprotein.     

Expression of tropomyosin alpha 4 chain is increased in esophageal squamous cell carcinoma as evidenced by proteomic profiling by two-dimensional electrophoresis and liquid chromatography-mass spectrometry/mass spectrometry

To identify proteins associated with esophageal carcinogenesis, we performed protein profiling of 16 esophageal squamous cell carcinomas (ESCCs) and paired noncancerous tissues by 2-DE and MS/MS. In cancerous tissues, three spots showed significant up-regulation in the amount of protein, while eight spots were significantly down-regulated. The identities of the spots were determined by PMF with LC-MS/MS and were confirmed by immunoblotting. The up-regulated proteins were tropomyosin alpha 4 chain, transgelin, and pyruvate kinase. The down-regulated proteins were serum albumin precursor, isoforms of annexin A1, tropomyosin beta chain, 14-3-3 protein sigma, and isoforms of serotransferrin precursor. In all 16 cases, up-regulation of the tropomyosin alpha 4 chain was confirmed by immunoblotting. Localization of the tropomyosin alpha 4 chain in ESCC cells and adjacent fibroblasts was confirmed by immunohistochemistry.     

Proteome analysis of human foetal, aged and advanced nuclear cataract lenses

The most complete proteome of human lenses has been compiled using 2-D LC-MS/MS analysis of foetal, aged normal and advanced nuclear cataract lenses. A total of 231 proteins were identified across all lens groups, including 112 proteins that have not been reported previously. Proteins were grouped according to their PANTHER molecular function classification in order to facilitate comparisons. Previously unreported N-terminal acetylation was detected in a number of proteins, with the majority being associated with the prior removal of a methionine residue. This pattern of proteolysis may indicate that methionine aminopeptidase activity is present in human lenses. Acetylation is likely to aid in the stability of proteins that are present in the lens for many decades. Protein sequences were also used to interrogate the three human lens cDNA libraries publicly available. Surprisingly, 84 proteins we identified were not present in the cDNA libraries.     

Towards a comprehensive proteome of normal and malignant human colon tissue by 2-D-LC-ESI-MS and 2-DE proteomics and identification of S100A12 as potential cancer biomarker

The aim of this study was to characterize the proteome of normal and malignant colonic tissue. We previously studied the colon proteome using 2‐DE and MALDI‐MS and identified 734 proteins (Roeßler, M., Rollinger, W., Palme S., Hagmann, M.‐L., et al.., Clin. Cancer Res. 2005, 11, 6550–6557). Here we report the identification of additional colon proteins from the same set of tissue samples using a complementary nano‐flow 2‐D‐LC‐ESI‐MS. In total, 484 proteins were identified in colon. Of these, 252 had also been identified by the 2‐DE/MALDI‐MS approach, whereas 232 proteins were unique to the 2‐D‐LC‐ESI‐MS analysis. Comparing protein expression in neoplastic and normal colon tissue indicated elevated expression of several proteins in colorectal cancer, among them the well established tumor marker carcinoembryonic antigen, as well as calnexin, 40S ribosomal protein S15a, serpin H1, and S100A12. Overexpression of these proteins was confirmed by immunoblotting. Serum levels of S100A12 were determined by ELISA and were found to be strongly elevated in colorectal cancer patients compared to healthy individuals. We conclude, that 2‐D‐LC‐ESI‐MS is a powerful approach to identify and compare protein profiles of tissue samples, that it is complementary to 2‐DE/MALDI‐MS approaches and has the potential to identify novel biomarkers.     

Thymosin β4 is differentially expressed in the cerebrospinal fluid of Creutzfeldt‐Jakob disease patients: a MALDI‐TOF MS protein profiling study

MALDI‐TOF protein profiling analysis permits the detection of peptides and small proteins in complex protein mixtures with great accuracy. We applied this analysis to cerebrospinal fluid (CSF) from 15 patients affected by Creutzfeldt‐Jakob disease (CJD). We compared the levels of the normalized ion signals of 11 sporadic and 4 genetic CJD forms with those from ten healthy control subjects and eight non‐CJD relapsing‐remitting multiple sclerosis patients. In so doing, we detected 61 differentially expressed ion signals in CJD samples compared to controls. Among the 61 signals, 3 signals had significantly increased levels with high statistical significance (p <0.0001) and were located at 3238.3 m/z, 4963.7 m/z, and 8565.3 m/z. We characterized the 5.0 and 8.6 kDa proteins as thymosin β4 N‐acetylated and free ubiquitin, respectively, while the 3.2‐kDa peptide remained uncharacterized. Although elevated ubiquitin levels have previously been described in CJD, we have demonstrated for the first time the involvement of thymosin β4 in a neurodegenerative disease. To support the validity of thymosin β4 levels obtained by MALDI‐TOF analysis, an independent enzyme immunoassay analysis was performed. Moreover, a validation cohort consisting of CSF from three CJD patients, five healthy subjects, and six non‐CJD relapsing‐remitting multiple sclerosis patients was analyzed in a similar way, yielding superimposable results. We propose that thymosin β4 is a potential new candidate marker for the ante mortem diagnosis of CJD disease.     

Subjective measures of presence and discomfort in viewers of color‐separation‐based stereoscopic cinema

Abstract— The experience of various people related to a real 3‐D cinema visit has been studied, and the results will be discussed. This study has two distinct parts, which includes the comfort of the viewers and their sense of presence. Eighty‐four viewers filled out questionnaires about comfort and visual strain. Forty‐one subjects described their presence experience during the movie presentation. A majority of the people felt comfortable after the movie viewing; they experienced only mild‐eyestrain‐related symptoms. People evaluated the movie world as highly realistic, but they did not feel that they were actors in the virtual‐movie world. Most of the participants would recommend 3‐D cinema to friends because it was a very entertaining experience.     

Characterization of 2450‐MHz microwave thermal coagulation zone based on characteristic length growth model and shape variation factor

It is critical for microwave ablation (MWA) treatment planning to evaluate the changes of thermal coagulation zones. In MWA procedures, the shapes and sizes of thermal coagulation zones are gradually evolving over time. To this end, a novel characterization and mapping method of thermal coagulation zones is presented in this article. Firstly, finite element method (FEM) models of temperature distributions for 40, 45, 50, 55, and 60 W microwave ablations were built to derive thermal ablation data of ex vivo porcine livers and were compared with experimental results. Secondly, growth models of characteristic lengths were fitted. Finally, characterization functions of thermal coagulation zones were developed using these growth models. In addition, shape variation factors were incorporated to handle the minor shape variations of thermal coagulation zones. Experimental results showed that these characterization functions could accurately represent the changes of thermal coagulation zones. The standard deviations between prediction results and simulated values were less than 1 mm. The comparative results were statistically analyzed by paired t test (P > 0.05), indicating no significant differences. The proposed method can simply and effectively predict the changes of MWA coagulation zones with time, thus providing reliable coagulation dimensions for the thermal ablation therapies.     

Parameter estimation of the 2D‐GTD model and RCS reconstruction based on an improved 2D‐ESPRIT algorithm

The two‐dimensional estimating signal parameter via rotational invariance techniques (2D‐ESPRIT) algorithm is a classical method to estimate parameters of the two‐dimensional geometric theory of diffraction (2D‐GTD) model. While as signal‐to‐noise‐ratio (SNR) decreases, the parameter estimation performance of 2D‐ESPRIT algorithm is severely influenced. To solve this problem, a performance‐enhanced 2D‐ESPRIT algorithm is proposed in this article. The improved 2D‐ESPRIT algorithm combines the conjugate data with the original back‐scattered data and obtains a novel covariance matrix by squaring the original total covariance matrix. Simulation results indicate that the improved algorithm has a better noise robustness and a more stable parameter estimation performance than the classical ESPRIT algorithm and the classical TLS‐2D‐ESPRIT algorithm. To further validate the superiority of the improved 2D‐ESPRIT algorithm, reconstructed radar cross section (RCS) is presented in this article. Compared with the classical 2D‐ESPRIT algorithm, the proposed algorithm presents higher RCS fitting precision. Furthermore, the impacts of other factors on parameter estimation, such as matrix pencil parameters and paring parameters, are also studied in this article.