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A novel human gastric primary cell culture system for modelling Helicobacter pylori infection in vitro.

ABSTRACT: Helicobacter pylori is the causative agent of gastric diseases and the main risk factor in the development of gastric adenocarcinoma. In vitro studies with this bacterial pathogen largely rely on the use of transformed cell lines as infection model. However, this approach is intrinsically artificial and especially inappropriate when it comes to investigating the mechanisms of cancerogenesis. Moreover, common cell lines are often defective in crucial signalling pathways relevant to infection and cancer. A long-lived primary cell system would be preferable in order to better approximate the human in vivo situation.Gastric glands were isolated from healthy human stomach tissue and grown in Matrigel containing media supplemented with various growth factors, developmental regulators and apoptosis inhibitors to generate long-lasting normal epithelial cell cultures.Culture conditions were developed which support the formation and quasi-indefinite growth of three dimensional (3D) spheroids derived from various sites of the human stomach. Spheroids could be differentiated to gastric organoids after withdrawal of Wnt3A and R-spondin1 from the medium. The 3D cultures exhibit typical morphological features of human stomach tissue. Transfer of sheared spheroids into 2D culture led to the formation of dense planar cultures of polarised epithelial cells serving as a suitable in vitro model of H. pylori infection.A robust and quasi-immortal 3D organoid model has been established, which is considered instrumental for future research aimed to understand the underlying mechanisms of infection, mucosal immunity and cancer of the human stomach.

SUBMITTER: Schlaermann P

PROVIDER: S-EPMC4752654 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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A novel human gastric primary cell culture system for modelling Helicobacter pylori infection in vitro.

Schlaermann Philipp P Toelle Benjamin B Berger Hilmar H Schmidt Sven C SC Glanemann Matthias M Ordemann Jürgen J Bartfeld Sina S Mollenkopf Hans J HJ Meyer Thomas F TF

Gut 20141224 2

<h4>Background and aims</h4>Helicobacter pylori is the causative agent of gastric diseases and the main risk factor in the development of gastric adenocarcinoma. In vitro studies with this bacterial pathogen largely rely on the use of transformed cell lines as infection model. However, this approach is intrinsically artificial and especially inappropriate when it comes to investigating the mechanisms of cancerogenesis. Moreover, common cell lines are often defective in crucial signalling pathway ...[more]

PMID: 25539675

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Project description:ObjectiveInfection with Helicobacter pylori (H pylori), especially cytotoxin-associated gene A-positive (CagA+) strains, has been associated with various gastrointestinal and extragastric diseases. The aim of this study was to characterize H pylori-induced alterations in the gastric and tongue coating microbiota and evaluate their potential impacts on human health.DesignThe gastric mucosa and tongue coating specimens were collected from 80 patients with chronic gastritis, and microbiota profiles were generated by 16S rRNA gene sequencing. Samples were grouped as H pylori negative (n = 32), CagA-negative H pylori infection (n = 13), and CagA-positive H pylori infection (n=35). The comparison of bacterial relative abundance was made using a generalized linear model. Functional profiling of microbial communities was predicted with PICRUSt and BugBase. Microbial correlation networks were produced by utilizing SparCC method.ResultsSignificant alterations of the gastric microbiota were found in the H pylori+/CagA+ samples, represented by a decrease in bacterial diversity, a reduced abundance of Roseburia, and increased abundances of Helicobacter and Haemophilus genera. At the community level, functions involved in biofilm forming, mobile element content, and facultative anaerobiosis were significantly decreased in gastric microbiome of the H pylori+ subjects. The presence of CagA gene was linked to an increased proportion of Gram-negative bacteria in the stomach, thereby contributing to an upregulation of lipopolysaccharide (LPS) biosynthesis. The number of bacterial interactions was greatly reduced in networks of both tongue coating and gastric microbiota of the H pylori+/CagA+ subject, and the cooperative bacterial interactions dominated the tongue coating microbiome.ConclusionsInfection with H pylori strains possessing CagA may increase the risk of various diseases, by upregulating LPS biosynthesis in the stomach and weakening the defense of oral microbiota against microorganisms with pathogenic potential.

Dataset Information

A novel human gastric primary cell culture system for modelling Helicobacter pylori infection in vitro.

Publications

A novel human gastric primary cell culture system for modelling Helicobacter pylori infection in vitro.

Similar Datasets

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