Helicobacter pylori Neutrophil-Activating Protein Directly Interacts with and Activates Toll-like Receptor 2 to Induce the Secretion of Interleukin-8 from Neutrophils and ATRA-Induced Differentiated HL-60 Cells.

Shao-Hsuan Wen,Zhi-Wei Hong,Han-Wen Chang,Chung-Chu Chen,Hua-Wen Fu

doi:10.3390/ijms222111560

Abstract

Helicobacter pylori neutrophil-activating protein (HP-NAP)-induced production of reactive oxygen species (ROS) by neutrophils and monocytes is regulated by pertussis toxin (PTX)-sensitive G proteins, whereas HP-NAP-induced cytokine secretion by monocytes is mediated by Toll-like receptor 2 (TLR2). However, it is unclear whether TLR2 participates in HP-NAP-induced cytokine secretion by neutrophils. Here, all-trans retinoic acid (ATRA)-induced differentiated HL-60 cells were first employed as a neutrophil model to investigate the molecular mechanisms underlying neutrophil responses to HP-NAP. HP-NAP-induced ROS production in ATRA-induced differentiated HL-60 cells is mediated by the PTX-sensitive heterotrimeric G protein-dependent activation of extracellular signal-regulated kinase 1/2 and p38-mitogen-activated protein kinase, which is consistent with the findings reported for human neutrophils. Next, whether TLR2 participated in HP-NAP-induced secretion of interleukin-8 (IL-8) was investigated in neutrophils and ATRA-induced differentiated HL-60 cells. In both cells, TLR2 participated in HP-NAP-induced IL-8 secretion but not HP-NAP-induced ROS production. Interestingly, PTX-sensitive G proteins also contributed to the HP-NAP-induced secretion of IL-8 from neutrophils and the differentiated HL-60 cells. Our ELISA-based binding assay further revealed the competitive binding of Pam3CSK4, a TLR2 agonist, and HP-NAP to TLR2, which suggests the presence of specific and direct interactions between HP-NAP and TLR2. Thus, HP-NAP directly interacts with and activates TLR2 to induce IL-8 secretion in neutrophils and ATRA-induced differentiated HL-60 cells.

Highlights

Helicobacter pylori (H. pylori), a Gram-negative and microaerophilic bacterium, is found in the gastric mucosa of humans
In all-trans retinoic acid (ATRA)-induced differentiated HL-60 cells, HPNAP induced a significant increase in the number of reactive oxygen species (ROS)-producing cells from 7 ± 1% to 23 ± 8%, whereas in dimethyl sulfoxide (DMSO)-induced differentiated HL-60 cells, Helicobacter pylori neutrophil-activating protein (HP-NAP) induced no significant increase in the number of ROS-producing cells from 14 ± 4% to 17 ± 5% (Figure 1A)
ATRA-induced differentiated HL-60 cells were chosen as a neutrophil model to investigate the cell signal responses that are triggered by HP-NAP

Summary

Introduction

Helicobacter pylori (H. pylori), a Gram-negative and microaerophilic bacterium, is found in the gastric mucosa of humans. H. pylori infects over half of the human population and its infection causes various gastroduodenal diseases including chronic gastritis and peptic ulcers [1]. Chronic gastric inflammation caused by H. pylori infection leads to neutrophil infiltration of the gastric mucosa [2,3]. The extent of neutrophil infiltration is correlated to the degree of mucosal damage in patients with H. pylori infection [4]. Neutrophil infiltration of the infection site of H. pylori can be attributed to the bacterial factors with chemotactic properties specific to neutrophils. H. pylori neutrophil-activating protein (HP-NAP), a key virulence factor of H. pylori, plays such a role because of its capability to attract and activate neutrophils

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Conclusion