Inflammation and Gli2 Suppress Gastrin Gene Expression in a Murine Model of Antral Hyperplasia

Abstract

Chronic inflammation in the stomach can lead to gastric cancer. We previously reported that gastrin-deficient (Gast−/−) mice develop bacterial overgrowth, inflammatory infiltrate, increased Il-1β expression, antral hyperplasia and eventually antral tumors. Since Hedgehog (Hh) signaling is active in gastric cancers but its role in precursor lesions is poorly understood, we examined the role of inflammation and Hh signaling in antral hyperplasia. LacZ reporter mice for Sonic hedgehog (Shh), Gli1, and Gli2 expression bred onto the Gast−/− background revealed reduced Shh and Gli1 expression in the antra compared to wild type controls (WT). Gli2 expression in the Gast−/− corpus was unchanged. However in the hyperplastic Gast−/− antra, Gli2 expression increased in both the mesenchyme and epithelium, whereas expression in WT mice remained exclusively mesenchymal. These observations suggested that Gli2 is differentially regulated in the hyperplastic Gast−/− antrum versus the corpus and by a Shh ligand-independent mechanism. Moreover, the proinflammatory cytokines Il-1β and Il-11, which promote gastric epithelial proliferation, were increased in the Gast−/− stomach along with Infγ. To test if inflammation could account for elevated epithelial Gli2 expression in the Gast−/− antra, the human gastric cell line AGS was treated with IL-1β and was found to increase GLI2 but decrease GLI1 levels. IL-1β also repressed human GAST gene expression. Indeed, GLI2 but not GLI1 or GLI3 expression repressed gastrin luciferase reporter activity by ∼50 percent. Moreover, chromatin immunoprecipitation of GLI2 in AGS cells confirmed that GLI2 directly binds to the GAST promoter. Using a mouse model of constitutively active epithelial GLI2 expression, we found that activated GLI2 repressed Gast expression but induced Il-1β gene expression and proliferation in the gastric antrum, along with a reduction of the number of G-cells. In summary, epithelial Gli2 expression was sufficient to stimulate Il-1β expression, repress Gast gene expression and increase proliferation, leading to antral hyperplasia.

Inhibition of Hedgehog signaling in the gastrointestinal tract: Targeting the cancer microenvironment

“…In contrast to inflammation-dependent regulation of Gli1 in the corpus 102 , we found using LacZ reporter mice that Gli2 but not Gli1 expression increased in hyperplastic antrums of gastrin null mice 75 . Loss of gastrin predisposes to tumor formation in the antrum but not the corpus 107 .…”

“…Further, the pro-inflammatory cytokine IL-1β stimulates expression of Gli2 instead of Gli1 in the AGS gastric cell line. Thus, the Hh-related transcription factor in the antrum initiating preneoplastic changes appears to be epithelial Gli2 and not Gli1 , whose expression remained exclusively in the stroma 75 . This study uncovered the possibility that transformation in the gut can be mediated by epithelial expression of Gli2 independent of Hh ligand.…”

“…It was previously reported that both the TGF and Ras-Map kinase pathways directly activate Gli1 independent of Smo 73, 74 . Moreover, it is now evident that cytokines activate Gli1 75 . Das et al.…”

“…Indeed, mutations in Ptch1 and Smo are infrequent in gastric cancers 105 . Acutely, Shh ligand acts as a chemoattractant recruiting inflammatory cells to the stomach 100 , but chronically myeloid cells do not appear to be responsive to canonical signaling indicating that non-canonical Hh signaling modulates Gli1 activity during chronic gastritis 6, 75, 102 . Thus the use of Hh inhibitors in human subjects chronically infected with Helicobacter might effectively prevent pre-neoplastic mucosal changes, i.e., SPEM, from which gastric adenocarcinomas can arise 106 .…”

Recapitulating Human Gastric Cancer Pathogenesis: Experimental Models of Gastric Cancer

“…Tumors in this mouse model are associated with bacterial overgrowth [137] and inflammation [136, 138]. We reported multiple inflammatory mediators, such as IL-1β, IL-11, and the Tgfβ pathway components activin A and follistatin with epithelial Gli2 appear to be important epithelial drivers of the histologic changes during antral transformation in the Gast −/− mice [139, 140]. Takashi et al investigated the role of gastrin in H. felis -infected hypergastrinemic transgenic (INS-GAS) mice, GAS-KO mice, and C57BL/6 wild-type mice on a uniform C57BL/6 genetic background housed under specific-pathogen-free conditions [141].…”