Angiopoietin-2 and the Vascular Endothelial Growth Factor Promote Migration and Invasion in Hepatocellular Carcinoma- and Intrahepatic Cholangiocarcinoma-Derived Spheroids
Adriana Romanzi,
Fabiola Milosa,
Gemma Marcelli,
Rosina Maria Critelli,
Simone Lasagni,
Isabella Gigante,
Francesco Dituri,
Filippo Schepis,
Massimiliano Cadamuro,
Gianluigi Giannelli,
Luca Fabris,
Erica Villa
Affiliations
Adriana Romanzi
Department of Biomedical, Metabolic and Neural Sciences, Clinical and Experimental Medicine Program, University of Modena and Reggio Emilia, 41125 Modena, Italy
Fabiola Milosa
Chimomo Department, Gastroenterology Unit, University of Modena and Reggio Emilia, 41125 Modena, Italy
Gemma Marcelli
Chimomo Department, Gastroenterology Unit, University of Modena and Reggio Emilia, 41125 Modena, Italy
Rosina Maria Critelli
Chimomo Department, Gastroenterology Unit, University of Modena and Reggio Emilia, 41125 Modena, Italy
Simone Lasagni
Department of Biomedical, Metabolic and Neural Sciences, Clinical and Experimental Medicine Program, University of Modena and Reggio Emilia, 41125 Modena, Italy
Isabella Gigante
National Institute of Gastroenterology IRCCS “Saverio de Bellis”, Research Hospital, 70013 Castellana Grotte, Italy
Francesco Dituri
National Institute of Gastroenterology IRCCS “Saverio de Bellis”, Research Hospital, 70013 Castellana Grotte, Italy
Filippo Schepis
Chimomo Department, Gastroenterology Unit, University of Modena and Reggio Emilia, 41125 Modena, Italy
Massimiliano Cadamuro
Department of Molecular Medicine, School of Medicine, University of Padua, 35121 Padua, Italy
Gianluigi Giannelli
National Institute of Gastroenterology IRCCS “Saverio de Bellis”, Research Hospital, 70013 Castellana Grotte, Italy
Luca Fabris
Department of Molecular Medicine, School of Medicine, University of Padua, 35121 Padua, Italy
Erica Villa
Chimomo Department, Gastroenterology Unit, University of Modena and Reggio Emilia, 41125 Modena, Italy
Aggressive hepatocellular carcinoma (HCC) overexpressing Angiopoietin-2 (ANG-2) (a protein linked with angiogenesis, proliferation, and epithelial–mesenchymal transition (EMT)), shares 95% of up-regulated genes and a similar poor prognosis with the proliferative subgroup of intrahepatic cholangiocarcinoma (iCCA). We analyzed the pro-invasive effect of ANG-2 and its regulator vascular endothelial growth factor (VEGF) on HCC and CCA spheroids to uncover posUsible common ways of response. Four cell lines were used: Hep3B and HepG2 (HCC), HuCC-T1 (iCCA), and EGI-1 (extrahepatic CCA). We treated the spheroids with recombinant human (rh) ANG-2 and/or VEGF and then observed the changes at the baseline, after 24 h, and again after 48 h. Proangiogenic stimuli increased migration and invasion capability in HCC- and iCCA-derived spheroids and were associated with a modification in EMT phenotypic markers (a decrease in E-cadherin and an increase in N-cadherin and Vimentin), especially at the migration front. Inhibitors targeting ANG-2 (Trebananib) and the VEGF (Bevacizumab) effectively blocked the migration ability of spheroids that had been stimulated with rh-ANG-2 and rh-VEGF. Overall, our findings highlight the critical role played by ANG-2 and the VEGF in enhancing the ability of HCC- and iCCA-derived spheroids to migrate and invade, which are key processes in cancer progression.
