Glycosylation-related gene expression in HT29-MTX-E12 cells upon infection by Helicobacter pylori

Abstract

AIMTo identify glycosylation-related genes in the HT29 derivative cell line, HT29-MTX-E12, showing differential expression on infection with Helicobacter pylori (H. pylori).METHODSPolarised HT29-MTX-E12 cells were infected for 24 h with H. pylori strain 26695. After infection RNA was isolated from both infected and non-infected host cells. Sufficient infections were carried out to provide triplicate samples for microarray analysis and for qRT-PCR analysis. RNA was isolated and hybridised to Affymetrix arrays. Analysis of microarray data identified genes significantly differentially expressed upon infection. Genes were grouped into gene ontology functional categories. Selected genes associated with host glycan structure (glycosyltransferases, hydrolases, lectins, mucins) were validated by real-time qRT-PCR analysis.RESULTSInfection of host cells was confirmed by the isolation of live bacteria after 24 h incubation and by PCR amplification of bacteria-specific genes from the host cell RNA. H. pylori do not survive incubation under the adopted culture conditions unless they associate with the adherent mucus layer of the host cell. Microarray analysis identified a total of 276 genes that were significantly differentially expressed (P < 0.05) upon H. pylori infection and where the fold change in expression was greater than 2. Six of these genes are involved in glycosylation-related processes. Real-time qRT-PCR demonstrated significant downregulation (1.8-fold, P < 0.05) of the mucin MUC20. REG4 was heavily expressed and significantly downregulated (3.1-fold, P < 0.05) upon infection. Gene ontology analysis was consistent with previous studies on H. pylori infection.CONCLUSIONGene expression data suggest that infection with H. pylori causes a decrease in glycan synthesis, resulting in shorter and simpler glycan structures.

Bifidobacterium longum subsp. infantis ATCC 15697 and Goat Milk Oligosaccharides Show Synergism In Vitro as Anti-Infectives against Campylobacter jejuni

“…Previously, we demonstrated an increase in B. infantis adherence to HT-29 cells after pre-exposure to GMO [11]. The HT-29 cells are extensively used as a model of the gastrointestinal tract, particularly as in vitro intestinal models of bacterial colonisation [41,42]. These cells exhibit classical characteristics that model small intestinal absorptive epithelial cells upon reaching confluence [43] and are a useful indicator of the structural landscape of the intestinal epithelium [44].…”

Intestinal mucus is capable of stabilizing supersaturation of poorly water-soluble drugs

“…Lastly, the type and form of mucus in the S-P vs. transport experiments were different, in that (porcine) mucus was mixed into the buffer in the S-P experiments, whereas (human) mucus was present as a discrete and continuous layer immediately adjacent to the cell monolayer in the transport experiments. Mucus that is secreted by HT29-MTX cell lines has been well-characterized in the literature, and consists mainly of MUC5AC and MUC1, as well as MUC2, MUC3A, and MUC5B mucins [43][44][45][46] . While the main secreted gel-forming mucin produced by HT29-MTX cultures is MUC5AC, the major gel-forming mucin in the small intestine is MUC2.…”

Comprehensive Integration of Genome-Wide Association and Gene Expression Studies Reveals Novel Gene Signatures and Potential Therapeutic Targets for Helicobacter pylori-Induced Gastric Disease

“…Only datasets with available raw data were included. After a thorough search and excluding datasets as specified above, four datasets for Human samples (GSE27411, GSE60427, GSE60662, and GSE5081) and four cell line infection datasets (GSE60661, GSE70394, GSE74577, and GSE74492) ( 7 , 21 – 27 ) were selected for further analysis. A total of 98 human samples were considered for downstream analysis of the discovery steps, containing data from 72 helicobacter-infected/gastritis/atrophy/metaplasia patients, and 26 healthy controls.…”