Regulation of CD44 expression by bone morphogenetic protein (BMP) signaling and inflammatory stimuli

Abstract

The adhesion molecule CD44 is involved in inflammation-mediated gastric carcinogenesis. Lgr5 marks populations of gastric stem cells. Lgr5-expressing cells can be identified in the antrum, along the lesser curvature of the oxyntic mucosa, and in gastric carcinomas. We reported that inflammation and inhibition of BMP signaling, achieved by expression of the BMP inhibitor noggin (Nog) in the oxyntic epithelium (H+/K+-Nog mice), and by crossing Lgr5-Cre mice to mice with floxed alleles of BMP receptor 1A (Lgr5-Cre;BMPR1Aflox-flox mice), activate aberrant Lgr5 cells that give rise to dysplastic, proliferating lineages. We tested the hypothesis that BMP signaling regulates the fate of Lgr5 cells and the expression of CD44 during gastric inflammation and carcinogenesis. To conduct lineage tracing in the presence of inhibition of BMP signaling we crossed H+/K+-Nog mice and BMPR1Aflox-flox mice to Rosa26-tdTomato (Tom) reporter mice to generate H+/K+-Nog;Rosa26-tdTom and BMPR1Aflox-flox;Rosa26-tdTom mice. Both mice were crossed to Lgr5-Cre mice to generate Lgr5-Cre;H+/K+-Nog;Rosa26-tdTom and Lgr5-Cre;BMPR1Aflox-flox;Rosa26tdTom mice. Infection with H. felis (HF) was used to induce inflammation. Animals were analyzed 3 months after HF inoculation. Distribution of Tom-, CD44- and BrdU-positive cells was analyzed by immunostaining. Cultures of human organoids derived from antral and fundic biopsies were used to assess the effect of IFNγ (30 ng/ml) on the expression of CD44 and Lgr5. Expression of CD44, Lgr5, IFNγ, TNF-α, IL-1-β, and of the BMP signaling inhibitors Smad6, Smad7 and Bambi was measured in 16 human tumors of the lesser curvature and in an equal number of cancers of the body using QRT-PCR. Inhibition of BMP signaling and infection with HF induced the expression of CD44. HF-infected Lgr5-Cre;H+/K+-Nog;Rosa26-tdTom mice and Lgr5-Cre;BMPR1Aflox-flox;Rosa26-tdTom mice exhibited Tom expression in glands of the antrum and of the lesser curvature that positively stained with anti-CD44 and -BrdU antibodies. IFNγ induced the expression of CD44 and Lgr5 in antral, but not in fundic organoids. Expression of CD44, Lgr5, IFNγ, TNF-α, IL-1-β, Smad6, Smad7 and Bambi was increased in tumors of the lesser curvature but not of the body. Inflammation and inhibition of BMP signaling induce CD44 and Lgr5 in vivo and in vitro, in both mouse and human tissues. Lineage tracing studies demonstrate the presence of aberrant Lgr5 cells located along the lesser curvature that give rise to proliferating lineages that express CD44. Tumors of the lesser curvature but not of the body exhibit increased expression of CD44, Lgr5, BMP inhibitors, and pro-inflammatory molecules. These findings underscore the importance of inflammation and BMP signaling in the development of neoplastic lesions of the stomach.