Unknown,Transcriptomics,Genomics,Proteomics

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Host genomic regions susceptible to damage during H. pylori infection

ABSTRACT: We identify here host genomic regions that are more likely to be damaged during an infection with the human gastirc pathogen Helicobacter pylori and characterize underlying features of them. By ChIP-seq we obtained global information about the localization of the DNA-damage marker H2AXpS139. We used 20 ng of immunoprecipitated DNA of the human gastric cell line AGS to produce 240 mio sequence reads in total and compared the damage-pattern of 6 h and 19 h infected cells with that of 10 Gy irradiated ones. While the overall levesl of DNA damage wetre similar in terms of broken DNA and accumulation of H2AXpS139, infection-induced damage clearly accumulates towards the ends of chromosomal arms, namely telomere-proximal and close to centromeres, it also enriches mainly in genic and especially in transcribed regions. We show, taht this damage pattern is unique to the infection, occurs also in human non-transformed gastric epithelial cells and shows some correlation with previously identified cancer-related genes. Examination of a DNA-damage induced histone modification from four different conditions.

ORGANISM(S): Homo sapiens

SUBMITTER: Thomas Meyer

PROVIDER: E-GEOD-55699 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:We previously established long-term 3D organoid culture systems for several murine tissues (intestine, stomach, pancreas and liver) as well as human intestine and pancreas. Here, we describe culture conditions to generate long-term 3D culture from human gastric stem cells. The technology can be applied to study the epithelial response to infection with Helicobacter pylori. Human gastric cultures can expand indefinitely in 3D Matrigel. Cultures can be generated from normal tissue, from single sorted stem cells, or from tumor tissue. Organoids maintain many characteristics of the respective tissue in terms of histology, marker expression and euploidy. Organoids from normal tissue express markers of four lineages of the stomach and self-organize in gland and pit-domains. They can be directed to specifically express either lineages of the gastric gland, or the gastric pit by addition of Nicotinamide and withdrawal of Wnt. While gastric pit lineages react marginally to bacterial infection, gastric gland lineages mount a strong inflammatory response. The gastric culture system provides a unique tool to study gastric pathologies. We generated 2 sets of experiments. The first set contains organoids in 4 conditions: (1) organoids in expansion condition ENRWFGNiTi ("gland-type organoids") from 3 donors, (2) organoids as in 1 but differentiated for 4 days in differentiation condition ENR_FGNiTi ("pit'type organoids"), (3) organoids as in 1 but infected with Helicobacter pylori strain P12 MOI 50 for 2 h, (4) organoids as in 2 but infected as in 3. All 4 conditions were tested on the same organoid line in parallel. This experiment was conducted independently with cultures from 3 different donors. The second set of experiments compares freshly isolated glands with organoids. Samples from 2 patients were analyzed. Each patient received a total gastrectomy. From each patient, glands from corpus region or pyloric antrum were isolated. From each isolation, one aliquod was stored for microarray analysis and one aliquod used to generate organoids. Organoids and glands were subsequently lysed and analyzed in parallel.

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Host genomic regions susceptible to damage during H. pylori infection

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