title: HIF-2α attenuates lymphangiogenesis by up-regulating IGFBP1 in hepatocellular carcinoma.
abstract: BACKGROUND
  Tumour-associated lymphangiogenesis was identified as an important clinical determinant for the prognosis of hepatocellular carcinoma (HCC) and significantly influences patient survival. However, in this context, little is known about regulation of lymphangiogenesis by hypoxia-inducible factors (HIF). In HCC, mainly HIF-1α was positively correlated with lymphatic invasion and metastasis, whereas a defined role of HIF-2α is missing.
  
  
  RESULTS
  We created a stable knockdown (k/d) of HIF-1α and HIF-2α in HepG2 cells and generated co-cultures of HepG2 spheroids with embryonic bodies. This constitutes an in vitro tumour model mimicking the cancer microenvironment and allows addressing the role of distinct HIF isoforms in regulating HCC lymphangiogenesis. In co-cultures with a HIF-2α k/d, lymphangiogenesis was significantly increased, whereas the k/d of HIF-1α showed no effect. The HIF-2α-dependent lymphangiogenic phenotype was confirmed in vivo using matrigel plug assays with supernatants of HIF-2α k/d HepG2 cells. We identified and verified insulin-like growth factor binding protein 1 (IGFBP1) as a HIF-2α target gene. The potential of HepG2 cells to induce lymphangiogenesis in two independent functional assays was significantly enhanced either by a k/d of HIF-2α or by silencing IGFBP1. Moreover, we confirmed IGF as a potent pro-lymphatic growth factor with IGFBP1 being its negative modulator.
  
  
  CONCLUSIONS
  We propose that HIF-2α acts as an important negative regulator of hepatic lymphangiogenesis in vitro and in vivo by inducing IGFBP1 and thus, interfering with IGF signalling. Therefore, HIF-2α may constitute a critical target in HCC therapy.
authors.affiliation: Institute of Biochemistry I, Goethe-University Frankfurt, Frankfurt am Main, 60590, Germany.
authors.forename: Theresa
authors.initials: T
authors.lastname: Geis
authors.affiliation: Institute for Vascular Signalling, Centre for Molecular Medicine, Goethe-University Frankfurt, Frankfurt am Main, 60590, Germany.
authors.forename: Rüdiger
authors.initials: R
authors.lastname: Popp
authors.affiliation: Institute for Vascular Signalling, Centre for Molecular Medicine, Goethe-University Frankfurt, Frankfurt am Main, 60590, Germany.
authors.forename: Jiong
authors.initials: J
authors.lastname: Hu
authors.affiliation: Institute for Vascular Signalling, Centre for Molecular Medicine, Goethe-University Frankfurt, Frankfurt am Main, 60590, Germany.
authors.forename: Ingrid
authors.initials: I
authors.lastname: Fleming
authors.affiliation: Institute of Biochemistry I, Goethe-University Frankfurt, Frankfurt am Main, 60590, Germany.
authors.forename: Nina
authors.initials: N
authors.lastname: Henke
authors.affiliation: Institute of Biochemistry I, Goethe-University Frankfurt, Frankfurt am Main, 60590, Germany.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Dehne
authors.affiliation: Institute of Biochemistry I, Goethe-University Frankfurt, Frankfurt am Main, 60590, Germany.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Brüne
mesh_terms: D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics*; D006528:Carcinoma, Hepatocellular / Q000235:genetics*; D002460:Cell Line; D045744:Cell Line, Tumor; D018920:Coculture Techniques; D005260:Female; D056945:Hep G2 Cells; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics; D018970:Insulin-Like Growth Factor Binding Protein 1 / Q000235:genetics*; D008113:Liver Neoplasms / Q000235:genetics*; D042583:Lymphangiogenesis / Q000235:genetics*; D008207:Lymphatic Metastasis / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D015854:Up-Regulation / Q000235:genetics*; D000099162:Endothelial PAS Domain-Containing Protein 1
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D018970:Insulin-Like Growth Factor Binding Protein 1; D000099162:Endothelial PAS Domain-Containing Protein 1; C497442:HIF1A protein, human; C032231:IGFBP1 protein, human
keywords: Insulin-like growth factor-signalling; Spheroid-embryonic body-co-culture; Tumour hypoxia