title: Modulation of chemotactic and pro-inflammatory activities of endothelial progenitor cells by hepatocellular carcinoma.
abstract: Endothelial progenitor cells (EPCs) participate in the neovascularization processes in the development of hepatocellular carcinoma (HCC). We investigated whether interactions between EPCs and HCC cells affect chemotactic and pro-inflammatory activities of EPCs. Two distinct phenotypes of circulating EPCs, i.e., myeloid-derived EPCs (colony forming unit-endothelial cells, CFU-ECs) and outgrowth EPCs (endothelial-colony forming cells, ECFCs), were co-cultured with Huh7 and Hep3B cells by using transwell chamber and IBIDI(TM) Culture-Inserts and μ-slide plates. Transwell and horizontal migration/invasion assays and time-lapse microscopy were used to monitor and analyze the migration and invasion of EPCs induced by these HCC cells. A human cytokine antibody array was used to compare protein expression profiles in EPCs and HCC cells. Flow cytometry and electromobility shift analysis were used to detect nuclear factor-κB (NF-κB)-DNA binding activity and pro-inflammatory adhesion molecule expression in EPCs. Ectopic full-length CC chemokine receptor 6 (CCR6) plasmid was used to transfect into ECFCs to investigate the role of CCR6 in HCC-induced EPC migration and invasion. The results show that co-culture with Huh7 and Hep3B cells induces the expression of endothelial cell (EC) markers KDR, Flt1, CD31 and VE-cadherin in CFU-ECs, but down-regulates the expressions of CD31 and VE-cadherin in ECFCs. These HCC cells induce migration and invasion of CFU-ECs, but not ECFCs, and do not affect the cell cycle distribution in these EPCs. Cytokine protein array identifies macrophage inflammatory protein-3α (MIP-3α) produced by HCC cells as a critical factor responsible for the HCC-induced chemotaxis of CFU-ECs, which highly express the specific MIP-3α counterreceptor CCR6. Overexpressing CCR6 in ECFCs significantly increases their chemotaxis in response to HCC cells. Co-culturing EPCs with HCC cells results in decreases in NF-κB binding activity and hence intracellular adhesion molecule-1, vascular cell adhesion molecule-1, and E-selectin expressions in EPCs. Our results indicate that HCC cells exert differential effects on CFU-ECs and ECFCs, with increased chemotaxis for CFU-ECs, but not ECFCs. This HCC-induced chemotaxis of CFU-ECs is mediated by MIP-3α produced by HCC cells, which targets to CCR6 on CFU-ECs. Tumors may provide a humoral microenvironment to attenuate the pro-inflammatory activity of EPCs, which might be associated with the tumor escape mechanism.
authors: Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan.
authors: Yu-Tsung
authors: YT
authors: Shih
authors: Mei-Cun
authors: MC
authors: Wang
authors: Hsin-Hsin
authors: HH
authors: Peng
authors: Ting-Fang
authors: TF
authors: Chen
authors: Linyi
authors: L
authors: Chen
authors: Jang-Yang
authors: JY
authors: Chang
authors: Jeng-Jiann
authors: JJ
authors: Chiu
mesh_terms: D015703:Antigens, CD / Q000378:metabolism; D015820:Cadherins / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000503:physiopathology*; D002465:Cell Movement; D002478:Cells, Cultured; D054418:Chemokine CCL20 / Q000378:metabolism; D002633:Chemotaxis; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms / Q000503:physiopathology*; D016328:NF-kappa B / Q000378:metabolism; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000378:metabolism; D040081:Protein Array Analysis; D054399:Receptors, CCR6 / Q000235:genetics / Q000378:metabolism; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D000099152:Cadherin 5
chemical_list: D015703:Antigens, CD; D015820:Cadherins; D054418:Chemokine CCL20; D016207:Cytokines; D016328:NF-kappa B; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D054399:Receptors, CCR6; D000099152:Cadherin 5; C108355:CCL20 protein, human; C106311:CCR6 protein, human