Bombesin increases the expression of c-Met in epithelium and of hepatocyte growth factor in fibroblasts
Department of Surgery, The University of Texas Medical Branch, Galveston, TX, United States
Gastrin-releasing peptide (GRP) and its cognate receptor (GRP-R) are aberrantly overexpressed in several cancers, such as colorectal, prostate and lung cancers. Bombesin (BBS), a homolog of GRP, has been shown to be a mitogen in several cell types. We have reported that BBS is a potent stimulator of COX-2 gene expression in intestinal epithelial cells (J Biol Chem 276:22941, 2001). In addition, we recently found that functional GRP-R receptor exists in primary cultures of epithelium and fibroblasts from collagenase-dissociated human colon tumors, suggesting the possibility that BBS may induce stroma-epithelium interaction through stimulating some gene expression in both cell types. Hepatocyte growth factor (HGF) is a potent mitogenic and motogenic stimulator in normal and neoplastic cells and is produced mainly by fibroblasts; whereas its cognate receptor, c-Met, is expressed in epithelium. To date, whether BBS affects the expression of HGF and c-Met has not been reported. AIM. The purpose of this study is to determine the effect of BBS on the expression of HGF gene in 3T3-L1 fibroblasts and the c-Met gene in MC-26 mouse colon cancer cells. METHODS. 3T3-L1 and MC-26 cells were treated with or without BBS (10 nM) in serum-free conditions. Protein levels of HGF and c-Met were determined by Western blotting. The HGF mRNA was examined by cDNA microarray. The c-Met promoter activity was evaluated by transfection with c-Met promoter coupled to a luciferase reported gene and then examined by luciferase assay. RESULTS. BBS increased [Ca2+]i mobilization in both cell lines, indicating the presence of functional GRP-R. BBS treatment significantly increased the level of HGF mRNA and induced a time-dependent increase of HGF protein in 3T3-L1 cells. In MC-26 cells, BBS increased tyrosine phosphorylation and protein levels of c-Met as well as c-Met promoter activity. CONCLUSIONS. BBS stimulated the expression of HGF mRNA and protein in 3T3-L1 fibroblasts and transactivated c-Met, as well as increased c-Met protein levels and promoter activity in MC-26 cells. These data suggest that in colon cancer, GRP-R may play a role in tumor progression through the establishment of a paracrine growth loop between epithelial and stromal cells.
Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Nice, France; February 7 - 10, 2004; in poster session 898 (Oncogenesis).