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The extravasation of cancer cells, a key step for distant metastasis, is thought to be initiated by disruption of the endothelial barrier by malignant cancer cells. An endothelial covering-type extravasation of cancer cells in addition to conventional cancer cell invasion-type extravasation was dynamically visualized in a zebrafish hematogenous metastasis model. The inhibition of VEGF-signaling impaired the invasion-type extravasation via inhibition of cancer cell polarization and motility regulated by an intracellular signaling. Paradoxically, the inhibition of VEGF-signaling showed the promotion, rather than the inhibition, of the endothelial covering-type extravasation of cancer cells, with structural changes in the endothelial walls. These findings may be a clue to the full understanding of the metastatic process as well as the metastatic acceleration by antiangiogenic reagents observed in preclinical studies.
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Assessing the morphological Recovery of VEGF-depleted Cells by Cancer Cell-cultured Medium
Twenty-four h after the siRNA transfection, culture medium was replaced with fresh medium (left) or cultured medium in which normal RFP-HeLa cells were cultured for 36 h (right), and then siRNA-treated cells were cultured for another 24 h. Phase contrast images show the RFP-HeLa cells in the polymer-bottom dishes 48 h after the siRNA transfection. Bar, 20 μm.
Effects of Sunitinib Treatment on Cancer Cell Properties and Extravasation
(A) Phase contrast images of control and sunitinib-treated RFP-HeLa cells. (B) Quantitative evaluation of chemotactic migration in control and sunitinib-treated RFP-HeLa cells (mean ± SD, n=3). (C) Confocal microscopic images of control and sunitinib-treated RFP-Hep2 cells showing the distributions of vinculin (green) and DNA (blue). (D) Representative images after 11 h observation of RFP-HeLa cells that formed severe emboli with and without sunitinib treatment. Red colors in the bottom images show the area of extravasated RFP-HeLa cells. Bars, 20 μm (A, C), 40 μm (D).