Three-dimensional growth as multicellular spheroid activates the proangiogenic phenotype of colorectal carcinoma cells via LFA-1-dependent VEGF: implications on hepatic micrometastasis.
BACKGROUND
The recruitment of vascular stromal and endothelial cells is an early event occurring during cancer cell growth at premetastatic niches, but how the microenvironment created by the initial three-dimensional (3D) growth of cancer cells affects their angiogenesis-stimulating potential is unclear.


METHODS
The proangiogenic profile of CT26 murine colorectal carcinoma cells was studied in seven-day cultured 3D-spheroids of <300 mum in diameter, produced by the hanging-drop method to mimic the microenvironment of avascular micrometastases prior to hypoxia occurrence.


RESULTS
Spheroid-derived CT26 cells increased vascular endothelial growth factor (VEGF) secretion by 70%, which in turn increased the in vitro migration of primary cultured hepatic sinusoidal endothelium (HSE) cells by 2-fold. More importantly, spheroid-derived CT26 cells increased lymphocyte function associated antigen (LFA)-1-expressing cell fraction by 3-fold; and soluble intercellular adhesion molecule (ICAM)-1, given to spheroid-cultured CT26 cells, further increased VEGF secretion by 90%, via cyclooxygenase (COX)-2-dependent mechanism. Consistent with these findings, CT26 cancer cells significantly increased LFA-1 expression in non-hypoxic avascular micrometastases at their earliest inception within hepatic lobules in vivo; and angiogenesis also markedly increased in both subcutaneous tumors and hepatic metastases produced by spheroid-derived CT26 cells.


CONCLUSIONS
3D-growth per se enriched the proangiogenic phenotype of cancer cells growing as multicellular spheroids or as subclinical hepatic micrometastases. The contribution of integrin LFA-1 to VEGF secretion via COX-2 was a micro environmental-related mechanism leading to the pro-angiogenic activation of soluble ICAM-1-activated colorectal carcinoma cells. This mechanism may represent a new target for specific therapeutic strategies designed to block colorectal cancer cell growth at a subclinical micrometastatic stage within the liver.
Valcárcel
María
M
Dept, Cell Biology and Histology, Basque Country University School of Medicine & Dentistry, Bizkaia-48940, Spain. valcarcelcuesta@yahoo.es
Arteta
Beatriz
B
Jaureguibeitia
Arrate
A
Lopategi
Aritz
A
Martínez
Iñigo
I
Mendoza
Lorea
L
Muruzabal
Francisco J
FJ
Salado
Clarisa
C
Vidal-Vanaclocha
Fernando
F
D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects; D015179:Colorectal Neoplasms / Q000098:blood supply* / Q000201:enzymology / Q000473:pathology; D051546:Cyclooxygenase 2 / Q000378:metabolism; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000494:pharmacology; D008113:Liver Neoplasms / Q000556:secondary*; D016169:Lymphocyte Function-Associated Antigen-1 / Q000378:metabolism*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009389:Neovascularization, Pathologic / Q000201:enzymology / Q000473:pathology*; D010641:Phenotype; D011994:Recombinant Proteins / Q000494:pharmacology; D012995:Solubility / Q000187:drug effects; D018874:Spheroids, Cellular / Q000187:drug effects / Q000201:enzymology / Q000473:pathology*; D014407:Tumor Cells, Cultured; D014410:Tumor Stem Cell Assay; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*
D016861:Cyclooxygenase Inhibitors; D016169:Lymphocyte Function-Associated Antigen-1; D011994:Recombinant Proteins; D042461:Vascular Endothelial Growth Factor A; D018799:Intercellular Adhesion Molecule-1; D051546:Cyclooxygenase 2