Psoriasin, one of several new proteins identified in nasal lavage fluid from allergic and non-allergic individuals using 2-dimensional gel electrophoresis and mass spectrometry

BACKGROUND: Extravasation and luminal entry of plasma occurs continuously in the nose. This process is markedly facilitated in patients with symptomatic allergic rhinitis, resulting in an increased secretion of proteins. Identification of these proteins is an important step in the understanding of the pathological mechanisms in allergic diseases. DNA microarrays have recently made it possible to compare mRNA profiles of lavage fluids from healthy and diseased patients, whereas information on the protein level is still lacking. METHODS: Nasal lavage fluid was collected from 11 patients with symptomatic allergic rhinitis and 11 healthy volunteers. 2-dimensional gel electrophoresis was used to separate proteins in the lavage fluids. Protein spots were picked from the gels and identified using mass spectrometry and database search. Selected proteins were confirmed with western blot. RESULTS: 61 spots were identified, of which 21 were separate proteins. 6 of these proteins (psoriasin, galectin-3, alpha enolase, intersectin-2, Wnt-2B and hypothetical protein MGC33648) had not previously been described in nasal lavage fluids. The levels of psoriasin were markedly down-regulated in allergic individuals. Prolactin-inducible protein was also found to be down-regulated, whereas different fragments of albumin together with Ig gamma 2 chain c region, transthyretin and splice isoform 1 of Wnt-2B were up-regulated among the allergic patients. CONCLUSION: The identification of proteins in nasal lavage fluid with 2-dimensional gelelectrophoresis in combination with mass spectrometry is a novel tool to profile protein expression in allergic rhinitis and it might prove useful in the hunt for new therapeutic targets or diagnostic markers for allergic diseases. Psoriasin is a potent chemotactic factor and its down-regulation during inflammation might be of importance for the outcome of the disease

Global Analysis of Quorum Sensing Targets in the Intracellular Pathogen Brucella melitensis 16 M

Many pathogenic bacteria use a regulatory process termed quorum sensing (QS) to produce and detect small diffusible molecules to synchronize gene expression within a population. In Gram-negative bacteria, the detection of, and response to, these molecules depends on transcriptional regulators belonging to the LuxR family. Such a system has been discovered in the intracellular pathogen Brucella melitensis, a Gram-negative bacterium responsible for brucellosis, a worldwide zoonosis that remains a serious public health concern in countries were the disease is endemic. Genes encoding two LuxR-type regulators, VjbR and BabR, have been identified in the genome of B. melitensis 16 M. A DeltavjbR mutant is highly attenuated in all experimental models of infection tested, suggesting a crucial role for QS in the virulence of Brucella. At present, no function has been attributed to BabR. The experiments described in this report indicate that 5% of the genes in the B. melitensis 16 M genome are regulated by VjbR and/or BabR, suggesting that QS is a global regulatory system in this bacterium. The overlap between BabR and VjbR targets suggest a cross-talk between these two regulators. Our results also demonstrate that VjbR and BabR regulate many genes and/or proteins involved in stress response, metabolism, and virulence, including those potentially involved in the adaptation of Brucella to the oxidative, pH, and nutritional stresses encountered within the host. These findings highlight the involvement of QS as a major regulatory system in Brucella and lead us to suggest that this regulatory system could participate in the spatial and sequential adaptation of Brucella strains to the host environment.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe

The Whereabouts of 2D Gels in Quantitative Proteomics

Two-dimensional gel electrophoresis has been instrumental in the development of proteomics. Although it is no longer the exclusive scheme used for proteomics, its unique features make it a still highly valuable tool, especially when multiple quantitative comparisons of samples must be made, and even for large samples series. However, quantitative proteomics using 2D gels is critically dependent on the performances of the protein detection methods used after the electrophoretic separations. This chapter therefore examines critically the various detection methods (radioactivity, dyes, fluorescence, and silver) as well as the data analysis issues that must be taken into account when quantitative comparative analysis of 2D gels is performed

Protein Detection and Quantitation Technologies for Gel-Based Proteome Analysis

Also published as a book chapter: Proteomics, 2009 / J. Reinders and A. Sickmann (eds.), Ch.4 pp.59-82Numerous protein detection and quantitation methods for gel-based proteomics have been devised that can be classified in three major categories: (1) Universal (or "general") detection techniques, which include staining with anionic dyes (e.g., Coomassie brilliant blue), reverse (or "negative") staining with metal cations (e.g., imidazole-zinc), silver staining, fluorescent staining or labeling, and radiolabeling, (2) specific staining methods for the detection of post-translational modifications (e.g., glycosylation or phosphorylation), and (3) differential display techniques for the separation of multiple, covalently tagged samples in a single two-dimensional electrophoresis (2-DE) gel, followed by consecutive and independent visualization of these proteins to minimize methodical variations in spot positions and in protein abundance, to simplify image analysis, as well as to improve protein quantitation by including an internal standard.The most important properties of protein detection methods applied in proteome analysis include high sensitivity (i.e., low detection limit), wide linear dynamic range for quantitative accuracy, reproducibility, cost-efficiency, ease of use, and compatibility with downstream protein identification or characterization technologies, such as mass spectrometry (MS). Regrettably, no single detection method meets all these requirements, albeit fluorescence-based technologies are currently favored for most applications; hence, the major focus of this chapter is on fluorescent-dye-based protein detection and quantitation techniques. Although satisfying results with respect to sensitivity and reproducibility are also obtained by methods based on radioactive labeling of proteins (which is still unsurpassed in terms of sensitivity), radiolabeling is, however, largely impractical for routine proteomic profiling because of the costs and the health and safety concerns associated with handling radioactive compounds.Walter Weiss, Florian Weiland, and Angelika Gör

Quality of bowel cleansing in hospetalized patients undergoing colonoscopy: a multicenter prospective regional study

BACKGROUND: Quality of bowel cleansing in hospitalized patients undergoing colonoscopy is often unsatisfactory. No study has investigated the inpatient or outpatient setting as cause of inadequate cleansing. AIMS: To assess degree of bowel cleansing in inpatients and outpatients and to identify possible predictors of poor bowel preparation in the two populations. METHODS: Prospective multicentre study on consecutive colonoscopies in 25 regional endoscopy units. Univariate and multivariate analysis with odds ratio estimation were performed. RESULTS: Data from 3276 colonoscopies were analyzed (2178 outpatients, 1098 inpatients). Incomplete colonoscopy due to inadequate cleansing was recorded in 369 patients (11.2%). There was no significant difference in bowel cleansing rates between in- and outpatients in both colonic segments. In the overall population, independent predictors of inadequate cleansing both at the level of right and left colon were: male gender (odds ratio, 1.20 [1.02-1.43] and 1.27 [1.05-1.53]), diabetes mellitus (odds ratio, 2.35 [1.68-3.29] and 2.12 [1.47-3.05]), chronic constipation (odds ratio, 1.60 [1.30-1.97] and 1.55 [1.23-1.94]), incomplete purge intake (odds ratio, 2.36 [1.90-2.94] and 2.11 [1.68-2.65]) and a runway time >12h (odds ratio, 3.36 [2.40-4.72] and 2.53 [1.74-3.67]). CONCLUSIONS: We found no difference in the rate of inadequate bowel preparation between hospitalized patients and outpatient