Sonic Hedgehog Acts as a Macrophage Chemoattractant During Gastric Epithelial Regeneration in Response to Injury
Authors: Chakrabarti Jayati , Schumacher Michael , Engevik Amy , Chang Julie , Teal Emma , Hawkins Jennifer , Helmrath Michael , Zavros Yana 
Published: June 2021

Background/Hypothesis

Hedgehog (Hh) signaling not only plays a crucial role in embryonic development, but also controls gastric physiological events including epithelial cell differentiation, regeneration and acid secretion. Sonic Hedgehog (Shh), is secreted from gastric parietal cells in response to injury and contributes to the regeneration of the injured epithelium. Following injury, the recruitment of macrophages plays crucial role both in the regenerative process. The mechanism that regulates macrophage recruitment in response to injury is largely unknown. Thus, here we tested the hypothesis thatShh stimulates macrophage chemotaxis to the injured epithelium and contributes to gastric regeneration.

Methods

A mouse model expressing a myeloid cell‐specific deletion of Smoothened (LysMCre/SmoKO) was generated using transgenic mice bearing loxP sites flanking exon 1 of the Smo gene (Smo loxP) and mice expressing a Cre recombinase transgene from the Lysozyme M locus (LysMCre). Acetic acid injury was induced in the stomachs of both control and KO mice and gastric epithelial regeneration and macrophage recruitment analyzed over a period of 7 days post‐injury. Bone marrow‐derived macrophages (BM‐Mø) were collected from control and KO mice. Migration of BM‐Mø was performed using ibidi μ‐Slide Chemotaxis chambers in response to recombinant murine Shh (rmShh). Murine‐ and human‐derived gastric organoid/macrophage co‐cultures were established and macrophage chemotaxis measured.

Results

1) LysMCre/SmoKO mice exhibit loss of macrophage recruitment and epithelial regeneration in response to injury: Compared to control mice, KO animals exhibited loss of ulcer repair and normal epithelial regeneration, that correlated with decreased macrophage infiltration at the site of injury. 2) In vitro, macrophages from LysMCre/SmoKO mice or organoid/macrophage co‐cultures treated with Vismodegib lose chemotaxis: Forward migration index, a measure of directed movement, was significantly elevated in control BM‐Mø migrating towards rmShh compared to media control. In contrast, Smo‐deficient BM‐Mø did not migrate towards rmShh. While control BM‐Mø migrated toward chemokine CCL2, this chemotactic response was not observed using Smo‐deficient BM‐Mø. In murine‐derived gastric organoid/BM‐Mø co‐cultures, epithelial Shh, stimulated with histamine, correlated with macrophage migration toward the organoid. This response was lost in organoid/BM‐Mø co‐cultures derived from KO mice. Human‐derived organoid/macrophage co‐cultures stimulated with histamine resulted in Shh secretion and macrophage migration toward the gastric organoid, a response that was blocked with Smo‐inhibitor Vismodegib.

Conclusions

Shh signaling acts as a macrophage chemoattractant via a Smo‐dependent mechanism during gastric epithelial regeneration in response to injury.

Support or Funding Information

NIH (NIDDK) 5R01DK083402‐08 grant (YZ)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.