EGFR regulates macrophage activation and function in bacterial infection
Dana M. Hardbower, … , M. Blanca Piazuelo, Keith T. Wilson
Dana M. Hardbower, … , M. Blanca Piazuelo, Keith T. Wilson
Published August 2, 2016
Citation Information: J Clin Invest. 2016;126(9):3296-3312. https://doi.org/10.1172/JCI83585.
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EGFR regulates macrophage activation and function in bacterial infection

Abstract

EGFR signaling regulates macrophage function, but its role in bacterial infection has not been investigated. Here, we assessed the role of macrophage EGFR signaling during infection with Helicobacter pylori, a bacterial pathogen that causes persistent inflammation and gastric cancer. EGFR was phosphorylated in murine and human macrophages during H. pylori infection. In human gastric tissues, elevated levels of phosphorylated EGFR were observed throughout the histologic cascade from gastritis to carcinoma. Deleting Egfr in myeloid cells attenuated gastritis and increased H. pylori burden in infected mice. EGFR deficiency also led to a global defect in macrophage activation that was associated with decreased cytokine, chemokine, and NO production. We observed similar alterations in macrophage activation and disease phenotype in the Citrobacter rodentium model of murine infectious colitis. Mechanistically, EGFR signaling activated NF-κB and MAPK1/3 pathways to induce cytokine production and macrophage activation. Although deletion of Egfr had no effect on DC function, EGFR-deficient macrophages displayed impaired Th1 and Th17 adaptive immune responses to H. pylori, which contributed to decreased chronic inflammation in infected mice. Together, these results indicate that EGFR signaling is central to macrophage function in response to enteric bacterial pathogens and is a potential therapeutic target for infection-induced inflammation and associated carcinogenesis.

Authors

Dana M. Hardbower, Kshipra Singh, Mohammad Asim, Thomas G. Verriere, Danyvid Olivares-Villagómez, Daniel P. Barry, Margaret M. Allaman, M. Kay Washington, Richard M. Peek Jr., M. Blanca Piazuelo, Keith T. Wilson

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Figure 2

EGFR signaling in macrophages is induced by H. pylori infection in murine and human macrophages.

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EGFR signaling in macrophages is induced by H. pylori infection in murin...
(A) Representative Western blot of EGFR p-Y1068 and p-S1046/7 in RAW 264.7 cells at various time points p.i. with H. pylori PMSS1 or stimulation with EGF (5 ng/ml). n = 3 biological replicates. (B) Densitometric analysis of the levels of p-Y1068 compared with levels of t-EGFR at 15 and 30 minutes p.i. n = 3 biological replicates. ***P < 0.001, by 1-way ANOVA with Newman-Keuls post test. (C) Densitometric analysis of p-S1046/7 levels compared with t-EGFR levels at 15 and 30 minutes p.i. n = 3 biological replicates. *P < 0.05, **P < 0.01, and ***P < 0.001, by 1-way ANOVA with Newman-Keuls post test. (D) Representative Western blot of EGFR at p-Y1068 and p-S1046/7 in THP-1 cells at 15 minutes p.i. with H. pylori PMSS1. Monocytes (– PMA) and macrophages (+ PMA) are represented in this blot. n = 3 biological replicates. (E) Representative Western blot of EGFR at p-Y1068 and p-S1046/7 in RAW 264.7 cells at 15 minutes p.i. with H. pylori PMSS1 ± 10 μM gefitinib. n = 3 biological replicates. (F) Representative immunofluorescence images of WT BMmacs infected with H. pylori PMSS1 ± 10 μM gefitinib at the indicated times. EGFR p-Y1068 = green; DAPI = blue. n = 5 biological replicates. Scale bars: 50 μM. (G) Representative immunofluorescence images of WT BMmacs infected with H. pylori PMSS1 ± 10 ng/ml anti–TNF-α or ± 25 ng/ml anti–HB-EGF at 30 minutes p.i. Green = EGFR p-Y1068; blue = DAPI. n = 3 biological replicates. Scale bars: 50 μM. Ctrl, control.