Gastroenterology

Gastroenterology

Volume 142, Issue 5, May 2012, Pages 1150-1159.e6
Gastroenterology

Original Research
Basic and Translational—Alimentary Tract
Gastric Sonic Hedgehog Acts as a Macrophage Chemoattractant During the Immune Response to Helicobacter pylori

https://doi.org/10.1053/j.gastro.2012.01.029Get rights and content

Background & Aims

Macrophages mediate the epithelial response to Helicobacter pylori and are involved in the development of gastritis. Sonic Hedgehog (Shh) regulates gastric epithelial differentiation and function, but little is known about its immunoregulatory role in the stomach. We investigated whether gastric Shh acts as a macrophage chemoattractant during the innate immune response to H pylori infection.

Methods

Mice with parietal cell–specific deletion of Shh (PC-ShhKO) and control mice were infected with H pylori. Levels of gastric Shh, cytokines, and chemokines were assayed by quantitative reverse-transcriptase polymerase chain reaction or by a Luminex-based multiplex assay 2, 7, or 180 days after infection. Circulating concentrations of Shh were measured by enzyme-linked immunosorbent assay. Bone marrow chimera experiments were performed with mice that have myeloid cell–specific deletion of the Hedgehog signal transduction protein Smoothened (LysMCre/SmoKO). Macrophage recruitment was measured in gastric tissue and peripheral blood by fluorescence-activated cell sorting analysis.

Results

Control mice infected with H pylori for 6 months developed an inflammatory response characterized by infiltration of CD4+ T cells and increased levels of interferon gamma and interleukin 1β in the stomach. PC-ShhKO mice did not develop gastritis, even after 6 months of infection with H pylori. Control mice had increased concentrations of Shh, accompanied by the recruitment of CD11b+F4/80+Ly6Chigh macrophages 2 days after infection. Control mice that received bone marrow transplants from control mice had an influx of macrophages to the gastric mucosa in response to H pylori infection; this was not observed in H pylori–infected control mice that received bone marrow transplants from LysMCre/SmoKO mice.

Conclusions

H pylori induces release of Shh from the stomach; Shh acts as a macrophage chemoattractant during initiation of gastritis.

Section snippets

Animal Models

A mouse model expressing a parietal cell–specific deletion of Shh (formerly known as HKCre/ShhKO, referred to as PC-ShhKO in the current studies) was generated as previously reported.14 Age-matched Shh loxP (homozygous for the loxP sites without the Cre transgene) and HKCre littermates were used as controls.

PC-ShhKO mice were crossed with gastrin-deficient mice (GKO) to generate double-knockout PC-ShhKO/GastrinKO mice. Gastrin deficiency was confirmed by radioimmunoassay and polymerase chain

H pylori–Infected PC-ShhKO Mice Fail to Develop Gastritis

To determine the role of gastric Shh signaling in the initiation of gastritis, control and PC-ShhKO mice were infected with H pylori and analyzed 6 months after inoculation. Histologic examination revealed that H pylori generated a significant inflammatory response in the control group (Figure 1B) compared with the uninfected animals (Figure 1A). Interestingly, PC-ShhKO mice did not develop an inflammatory response after 6 months of H pylori infection (Figure 1C and D). The characteristic

Discussion

Hedgehog signaling components Ptch, Smo, and Gli are all expressed in macrophages.19, 23 Furthermore, pathway analysis of lipopolysaccharide-stimulated macrophages revealed that Hedgehog signaling genes were up-regulated within 24 hours, showing a capacity for stimulated macrophages to be Hedgehog responsive.34 Bone marrow chimera experiments using LysMCre/SmoKO mice showed the role of gastric Shh as a macrophage chemoattractant. First, the targeted deletion of Hedgehog signaling within the

Acknowledgments

The authors thank Dr Marsha Wills–Karp and Alyssa Sproles of the Cincinnati Cytokine/Chemokine/Mediator Measurement Core at the Digestive Health Center of Cincinnati Children's Medical Health Center for assistance and Dr Kathryn A. Eaton (Department of Microbiology and Immunology, University of Michigan, Ann Arbor, MI) for her discussion and suggestions.

References (42)

  • M.C. Martínez et al.

    Transfer of differentiation signal by membrane microvesicles harboring hedgehog morphogens

    Blood

    (2006)
  • B.J. Marshall et al.

    Unidentified curved bacilli in the stomach of patients with gastritis and peptic ulceration

    Lancet

    (1984)
  • K.A. Eaton et al.

    The role of T cell subsets and cytokines in the pathogenesis of Helicobacter pylori gastritis in mice

    J Immunol

    (2001)
  • Y. Zavros et al.

    Chronic gastritis in the hypochlorhydric gastrin-deficient mouse progresses to adenocarcinoma

    Oncogene

    (2005)
  • Y. Zavros et al.

    Treatment of Helicobacter gastritis with IL-4 requires somatostatin

    Proc Natl Acad Sci U S A

    (2003)
  • H.M. Algood et al.

    Host response to Helicobacter pylori infection before initiation of the adaptive immune response

    FEMS Immunol Med Microbiol

    (2007)
  • R.L. Ferrero et al.

    NF-kappaB activation during acute Helicobacter pylori infection in mice

    Infect Immun

    (2008)
  • S.H. Chu et al.

    Role of NF-kappaB and AP-1 on Helicobater pylori-induced IL-8 expression in AGS cells

    Dig Dis Sci

    (2003)
  • H. Mutoh et al.

    Monocyte chemoattractant protein-1 is generated via TGF-beta by myofibroblasts in gastric intestinal metaplasia and carcinoma without H. pylori infection

    Cancer Sci

    (2010)
  • T. Watanabe et al.

    Role of neutrophils in a rat model of gastric ulcer recurrence caused by interleukin-1 beta

    Am J Pathol

    (1997)
  • M. Kaparakis et al.

    Macrophages are mediators of gastritis in acute Helicobacter pylori infection in C57BL/6 mice

    Infect Immun

    (2008)
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    Conflicts of interest The authors disclose no conflicts.

    Funding Supported by National Institutes of Health grant 1R01DK083402-01 (to Y.Z.) and in part by the Digestive Health Center of Cincinnati Children's Medical Health Center (DHC: Bench to Bedside Research in Pediatric Digestive Disease) grant CHTF/SUB DK078392. Monica DeLay, manager of the Research Flow Cytometry Core in the Division of Rheumatology at Cincinnati Children's Hospital Medical Center, provided assistance and was supported in part by National Institutes of Health grant AR-47363. All flow cytometric data were acquired using equipment maintained by the Research Flow Cytometry Core in the Division of Rheumatology at Cincinnati Children's Hospital Medical Center, which was supported in part by National Institutes of Health grant AR-47363.

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