Abstract
The human gastric pathogen Helicobacter pylori activates human epithelial cells by a particular combination of mechanisms, including NOD1 and ALPK1-TIFA activation. These mechanisms are characterized by a strong participation of the bacterial cag pathogenicity island, which forms a type IV secretion system (CagT4SS) that enables the bacteria to transport proteins and diverse bacterial metabolites, including DNA, glycans, and cell wall components, into human host cells. Building on previous findings, we sought to determine the contribution of lipopolysaccharide inner core heptose metabolites (ADP-heptose) in the activation of human phagocytic cells by H. pylori. Using human monocyte/macrophage-like Thp-1 cells and human primary monocytes and macrophages, we were able to determine that a substantial part of early phagocytic cell activation, including NF-κB activation and IL-8 production, by live H. pylori is triggered by bacterial heptose metabolites. This effect was very pronounced in Thp-1 cells exposed to bacterial purified lysates or pure ADP-heptose, in the absence of other bacterial MAMPs, and was significantly reduced upon TIFA knock-down. Pure ADP-heptose on its own was able to strongly activate Thp-1 cells and human primary monocytes/macrophages. Comprehensive transcriptome analysis of Thp-1 cells co-incubated with live H. pylori or pure ADP-heptose confirmed a signature of ADP-heptose-dependent transcript activation in monocyte/macrophages. Bacterial enzyme-treated lysates (ETL) and pure ADP-heptose–dependent activation differentiated monocytes into macrophages of predominantly M1 type. In Thp-1 cells, the active CagT4SS was less required for the heptose-induced proinflammatory response than in epithelial cells, while active heptose biosynthesis or pure ADP-heptose was required and sufficient for their early innate response and NF-κB activation. The present data suggest that early activation and maturation of incoming and resident phagocytic cells (monocytes, macrophages) in the H. pylori–colonized stomach strongly depend on bacterial LPS inner core heptose metabolites, also with a significant contribution of an active CagT4SS.
Highlights
The bacterium Helicobacter pylori is a chronic human pathogen of major global importance, since about half of the world population carry this bacterial species in their stomach [1]
Various wild type strains at the same multiplicities of infection (MOI) activated the Thp-1 cells to a significantly different extent at 20 h post co-incubation (p.c.) (Figures 1C–E). cagA mutants did not show a significant difference in cell activation in comparison to wild type bacteria in any of the settings, while DcagPAI mutants or T4SS-inactive mutants showed a significantly reduced activation potential (IL-8 release) as compared to the parental wild type strain (Figure 1)
The aim of this study was to investigate the role of LPS heptosemediated signaling by H. pylori in human monocyte-like cells, which can be differentiated to a macrophage-like phenotype, exemplified by the model cell line Thp-1 [47, 53, 67]
Summary
The bacterium Helicobacter pylori is a chronic human pathogen of major global importance, since about half of the world population carry this bacterial species in their stomach [1]. TLR4 and TLR5 cellular receptors, widely present on epithelial cells, appear to be less activated by the bacteria, since the respective bacterial surface molecules (microbeassociated molecular patterns [MAMP]) of H. pylori, lipopolysaccharide (LPS/lipid A) and flagellins, have evolved to low activation potential [10, 23,24,25,26] Another very well-studied cell interaction module of H. pylori with respect to gastric epithelial cells is the cag pathogenicity island (cagPAI). The Cag secretion system is expressed by H. pylori in the stomach [33, 34] and can translocate various small molecules into gastric epithelial cells, including bacterial DNA [12], the NOD1 innate receptor ligand ieDAP [13] and lipopolysaccharide (LPS) heptose precursors, HBP and ADP-heptose [17, 21] These small molecule metabolites contribute to different extents and at different times to the epithelial cell activation and modulation by H. pylori [13, 16, 20, 35, 36]. At least two CagT4SS transported molecules, the oncogene CagA [37,38,39,40] and the peptidoglycan metabolite ieDAP [13, 16], provide signals of sustained, late cell activation [16, 41]
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