Outer Membrane Vesicle Production by Helicobacter pylori Represents an Approach for the Delivery of Virulence Factors CagA, VacA and UreA into Human Gastric Adenocarcinoma (AGS) Cells.

Hsin-Yu Chung,Po-Yi Lin,Yongyu Chew,Mou-Chieh Kao,Deng-Chyang Wu,Shau-Ku Huang

doi:10.3390/ijms22083942

Abstract

Helicobacter pylori infection is the etiology of several gastric-related diseases including gastric cancer. Cytotoxin associated gene A (CagA), vacuolating cytotoxin A (VacA) and α-subunit of urease (UreA) are three major virulence factors of H. pylori, and each of them has a distinct entry pathway and pathogenic mechanism during bacterial infection. H. pylori can shed outer membrane vesicles (OMVs). Therefore, it would be interesting to explore the production kinetics of H. pylori OMVs and its connection with the entry of key virulence factors into host cells. Here, we isolated OMVs from H. pylori 26,695 strain and characterized their properties and interaction kinetics with human gastric adenocarcinoma (AGS) cells. We found that the generation of OMVs and the presence of CagA, VacA and UreA in OMVs were a lasting event throughout different phases of bacterial growth. H. pylori OMVs entered AGS cells mainly through macropinocytosis/phagocytosis. Furthermore, CagA, VacA and UreA could enter AGS cells via OMVs and the treatment with H. pylori OMVs would cause cell death. Comparison of H. pylori 26,695 and clinical strains suggested that the production and characteristics of OMVs are not only limited to laboratory strains commonly in use, but a general phenomenon to most H. pylori strains.

Highlights

Helicobacter pylori is a flagellate spiral-shaped Gram-negative microaerophilic bacterium which is associated with several upper gastrointestinal disorders, such as gastritis, peptic ulcer disease, gastric mucosa-associated lymphoid tissue (MALT) lymphoma, and even gastric cancer [1]
Many reports have suggested that a complex interplay between H. pylori and host factors has evolved many bacterial factors, and some of them, including a number of adherence factors and virulence factors involved in the signaling pathways of host cells, have been identified by the scientific efforts made during the past several decades
The exact pathophysiological mechanisms of many gastric diseases are still not fully understood. Recent progress in this area has revealed that outer membrane vesicles (OMVs), which are constantly released from the surface of Gram-negative bacteria, can provide an additional mechanism for the pathogenicity of bacterial infection as they have the potential to cross the gastric epithelial barrier and may mediate delivery of virulence factors to host immune cells [41,42]

Summary

Introduction

Helicobacter pylori is a flagellate spiral-shaped Gram-negative microaerophilic bacterium which is associated with several upper gastrointestinal disorders, such as gastritis, peptic ulcer disease, gastric mucosa-associated lymphoid tissue (MALT) lymphoma, and even gastric cancer [1]. H. pylori releases several virulence factors such as cytotoxin-associated gene A (CagA), vacuolating cytotoxin (VacA) and urease, whose corresponding genes are frequently found in strains with enhanced pathogenicity. CagA is the best characterized virulence factor in H. pylori. It is an approximately 140-kDa immunedominant protein encoded by the cagA gene localized at the cag pathogenicity island (PAI). H. pylori attaches to the epithelial cells of host gastric tissues, CagA is rapidly injected into the cells through T4SS and phosphorylated by host c-Src family kinases at the tyrosine residue of its multiple Glu-Pro-Ile-Tyr-Ala (EPIYA) sites [3,4,5,6]. The phosphorylated CagA binds an eukaryotic phosphatase SHP-2 and induces the cytoskeleton remodeling, which causes the elongation of gastric epithelial cells, known as the hummingbird phenotype [6]

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