title: Muscarinic receptor agonists stimulate human colon cancer cell migration and invasion.
abstract: Muscarinic receptors (CHRM) are overexpressed in colon cancer. To explore a role for muscarinic receptor signaling in colon cancer metastasis, we used human H508 and HT29 colon cancer cells that coexpress epidermal growth factor (ERBB) and CHRM3 receptors. In a wound closure model, following 8-h incubation of H508 cells with 100 μM ACh we observed a threefold increase in cell migration indistinguishable from the actions of epidermal growth factor (EGF). Atropine blocked the actions of ACh but not of EGF. In SNU-C4 colon cancer cells that express ERBB but not CHRM, EGF caused a threefold increase in migration; ACh had no effect. ACh-induced cell migration was attenuated by chemical inhibitors of ERBB1 activation, by anti-ERBB1 antibody, and by inhibitors of ERK and phosphatidylinositol 3-kinase (PI3K) signaling. Consistent with matrix metalloproteinase-7 (MMP7)-mediated release of an ERBB1 ligand, heparin binding epidermal growth factor-like growth factor (HBEGF), ACh-induced migration was inhibited by an MMP inhibitor and by anti-MMP7 and -HBEGF antibodies. ACh-induced cell migration was blocked by inhibiting RhoA and ROCK, key proteins that interact with the actin cytoskeleton. ACh-induced RhoA activation was attenuated by agents that inhibit ERBB1, ERK, and PI3K activation. Collectively, these findings indicate that ACh-induced cell migration is mediated by MMP7-mediated release of HBEGF, an ERBB ligand that activates ERBB1 and downstream ERK and PI3K signaling. In a cell invasion model, ACh-induced HT29 cell invasion was blocked by atropine. In concert with previous observations, these findings indicate that muscarinic receptor signaling plays a key role in colon cancer cell proliferation, survival, migration, and invasion.
authors.affiliation: Division of Gastroenterology and Hepatology, Univ. of Maryland School of Medicine, Baltimore, 21201, USA.
authors.forename: Angelica
authors.initials: A
authors.lastname: Belo
authors.forename: Kunrong
authors.initials: K
authors.lastname: Cheng
authors.forename: Ahmed
authors.initials: A
authors.lastname: Chahdi
authors.forename: Jasleen
authors.initials: J
authors.lastname: Shant
authors.forename: Guofeng
authors.initials: G
authors.lastname: Xie
authors.forename: Sandeep
authors.initials: S
authors.lastname: Khurana
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Raufman
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology; D000199:Actins / Q000378:metabolism; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects*; D003110:Colonic Neoplasms / Q000473:pathology*; D003599:Cytoskeleton / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000502:physiology; D018773:Genes, erbB-1 / Q000235:genetics; D006801:Humans; D020783:Matrix Metalloproteinase 7 / Q000378:metabolism; D018721:Muscarinic Agonists / Q000494:pharmacology*; D009218:Myosins / Q000378:metabolism; D009361:Neoplasm Invasiveness / Q000473:pathology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013268:Stimulation, Chemical; D054460:rho-Associated Kinases / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
chemical_list: D000199:Actins; D018721:Muscarinic Agonists; D019869:Phosphatidylinositol 3-Kinases; D054460:rho-Associated Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D020783:Matrix Metalloproteinase 7; D009218:Myosins; D020742:rhoA GTP-Binding Protein; D000109:Acetylcholine