title: HGF regulates VEGF expression via the c-Met receptor downstream pathways,             PI3K/Akt, MAPK and STAT3, in CT26 murine cells.
abstract: In the present study, we assessed the involvement of hepatocyte growth factor (HGF)/c-Met signalling with vascular endothelial cell growth factor (VEGF) and hypoxia inducible factor (HIF)-1α expression in the downstream pathways phosphatidylinositol 3-kinase (PI3K)/Akt, mitogen-activated protein kinase (MAPK) and signal transducer and activator of transcription 3 (STAT3) in CT26 cells, to determine the mechanisms of the potent anti-angiogenic effect of NK4. We established genetically modified CT26 cells to produce NK4 (CT26-NK4). VEGF expression in subcutaneous CT26 tumours in vivo and in culture supernatants in vitro was determined by ELISA. HIF-1α expression in nuclear extracts was evaluated by western blot analysis. VEGF and HIF-1α mRNA levels were examined by real-time reverse transcription-polymerase chain reaction (RT-PCR). The DNA binding activity of HIF-1α was evaluated using an HIF-1α transcription factor assay kit. Our results demonstrated that VEGF expression was reduced in homografts of CT26-NK4 cells, compared to those of the control cells. In vitro, VEGF expression, which was induced by HGF, was inhibited by anti-HGF antibody, NK4 and by kinase inhibitors (PI3K, LY294002; MAPK, PD98059; and STAT3, Stattic). HGF‑induced HIF‑1α transcriptional activity was also inhibited by the kinase inhibitors. Real-time RT-PCR demonstrated that HGF‑induced HIF‑1α mRNA expression was not inhibited by LY294002 and PD98059, but was inhibited by Stattic. These data suggest that the PI3K/Akt, MAPK and STAT3 pathways, downstream of HGF/c‑Met signalling, are involved in the regulation of VEGF expression in CT26 cells. HGF/c‑Met signalling may be a promising target for anti-angiogenic strategies.
authors.affiliation: Division of Digestive Surgery, Department of Surgery, Kyoto Prefectural                     University of Medicine, Kamigyo-ku, Kyoto 602-8566, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Matsumura
authors.forename: Takeshi
authors.initials: T
authors.lastname: Kubota
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Taiyoh
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Fujiwara
authors.forename: Kazuma
authors.initials: K
authors.lastname: Okamoto
authors.forename: Daisuke
authors.initials: D
authors.lastname: Ichikawa
authors.forename: Atsushi
authors.initials: A
authors.lastname: Shiozaki
authors.forename: Shuhei
authors.initials: S
authors.lastname: Komatsu
authors.forename: Masayoshi
authors.initials: M
authors.lastname: Nakanishi
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Kuriu
authors.forename: Yasutoshi
authors.initials: Y
authors.lastname: Murayama
authors.forename: Hisashi
authors.initials: H
authors.lastname: Ikoma
authors.forename: Toshiya
authors.initials: T
authors.lastname: Ochiai
authors.forename: Yukihito
authors.initials: Y
authors.lastname: Kokuba
authors.forename: Takahiro
authors.initials: T
authors.lastname: Nakamura
authors.forename: Kunio
authors.initials: K
authors.lastname: Matsumoto
authors.forename: Eigo
authors.initials: E
authors.lastname: Otsuji
mesh_terms: D000230:Adenocarcinoma / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D000818:Animals; D003110:Colonic Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology; D015972:Gene Expression Regulation, Neoplastic; D017228:Hepatocyte Growth Factor / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics; D020935:MAP Kinase Signaling System / Q000235:genetics; D051379:Mice; D058990:Molecular Targeted Therapy; D051058:Oncogene Protein v-akt / Q000235:genetics / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D019859:Proto-Oncogene Proteins c-met / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
chemical_list: C553307:HGF protein, mouse; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D050796:STAT3 Transcription Factor; C494087:Stat3 protein, mouse; D042461:Vascular Endothelial Growth Factor A; C467480:vascular endothelial growth factor A, mouse; D017228:Hepatocyte Growth Factor; D019869:Phosphatidylinositol 3-Kinases; D019859:Proto-Oncogene Proteins c-met; D051058:Oncogene Protein v-akt