Comparative analysis of two-dimensional electrophoresis maps (2-DE) of Helicobacter pylori from Brazilian patients with chronic gastritis and duodenal ulcer: a preliminary report

Abstract

SUMMARYHelicobacter pylori is a bacterium recognized as the major cause of peptic ulcer and chronic gastritis. Recently, a proteomebased approach was developed to investigate pathogenic factors related to H. pylori. In this preliminary study, H. pylori strains were isolated from gastric biopsies of patients with chronic gastritis and duodenal ulcers. A partial proteomic analysis of H. pylori strains was performed by bacterial lyses and proteins were separated by two-dimensional gel electrophoresis (2-DE). A comparative analysis was performed to verify a differential protein expression between these two 2-DE maps. These data should be useful to clarify the role of different proteins related to bacterial pathogenesis. This study will be completed using a larger number of samples and protein identification of H. pylori by MALDI-TOF mass spectrometry. KEYWORDS:Helicobacter pylori; Proteome; Bidimensional electrophoresis; Chronic gastritis; Peptic ulcers.Helicobacter pylori is a pathogenic bacterium associated with the etiopathogenesis of universally spread chronic gastritis, peptic ulcers, gastric adenocarcinoma and MALT-lymphoma, mainly in developing countries 1,5 . Several of its biological aspects have not been revealed yet, in spite of molecular studies about this bacterium. A strategy to improve these studies consists in analyzing the whole protein expressed by the organism in its microenvironment. This is defined as proteome and the methodology is based around the technique of two-dimensional electrophoresis, which is directed towards display of proteins expressed inside gels, followed by protein identification using Peptide Mass Fingerprinting mass spectrometry (MALDI-TOF-MS) 7,8,9 .Recently, some reports have described a comparison among 2-DE proteome maps of clinical isolates obtained from patients who had peptic ulcers and chronic gastritis. It has been found that the main proteins have a different expression in these 2-DE maps, including GroEL (a protein that increases urease enzyme activity) and SodF (an enzyme involved in free radical and hydrogen peroxide catabolism) 2,3 . In this work, we carried out a comparative analysis of 2-DE maps of H. pylori from Brazilian patients, in order to find molecular targets related to its pathogenic mechanism in clinical isolates derived from duodenal ulcers and gastritis.H. pylori strains were isolated from gastric mucosa biopsies of two patients who had not been previously treated, submitted to upper gastrointestinal endoscopy at Gastrocentro, UNICAMP. Bacterial isolation and protein extraction were performed according to our previous reports 6,8 .The samples were submitted to isoelectrofocusing (IEF) solubilized with 8M urea, 4% CHAPS, 70 mM DTT, ampholine linear pH gradient 4-7 (pH 4-7L) 1.5% and 0.001% bromophenol blue. At this stage, 64 μg of each protein sample were applied to polyacrylamide dried strips, pH gradient 4-7 L (immobilized pH gradient -IPG). The first dimension of 2-DE was performed on an Amersham Biosciences Electrophoresis System accumulating 96 ...

Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) Applications in Bacteriology: brazilian contributions

“…Numerous publications reported MALDI-TOF MS applications on identification of diverse bacterial groups, genus or species such as Enterobacteriaceae (BIBERG et al, 2015), non-fermenting bacteria (MELLMANN et al, 2008), Staphylococcus spp. (DUBOIS et al, 2010), Neisseria (ILINA et al, 2009, Clostridia (GROSSE-HERRENTHEY et al, 2008), Mycobacteria (PIGNONE et al, 2006), Helicobacter pylori (PEREIRA et al, 2006), Campylobacter (MARTINY et al, 2011), Mycoplasma hominis (PAILHORIÈS et al, 2014) and some Haemophilus species (FRICKMANN et al, 2013).…”

“…In 2006, the MALDI-TOF MS analysis of Helicobacter pylori strains from chronic gastritis and duodenal ulcers was useful to identify bacterial proteins like GroEL and clarify the role of different proteins related to H. pylori pathogenesis (PEREIRA et al, 2006).…”

Peptic Ulcer Disease

“…A proteome analysis of H. pylori isolates from two patients ( Pereira et al, 2006 ), one with gastritis and the other with DU showed that four spots were uniquely associated with each condition and six spots had variable expression. Further work is required to confi rm this study and to identify the proteins.…”

Comparative Proteomic Analysis of <i>Helicobacter pylori</i> Strains Isolated from Chinese Patients

“…Jungblut et al reported that there were certain differences in the positions of protein spots in 2-DE maps of H. pylori strains 26695 and J99 [11]. Using comparative analyses of 2-DE maps from H. pylori isolates derived from duodenal ulcers and gastritis, Perei-ra et al reported that H. pylori isolates differentially expressed certain proteins [12]. However, whether gastrointestinal diseases are related to different patterns of H. pylori protein expression is still debated and further studies must be performed.…”