The Stemness-High Human Colorectal Cancer Cells Promote Angiogenesis by Producing Higher Amounts of Angiogenic Cytokines via Activation of the Egfr/Akt/Nf-κB Pathway.

purposeCancer stem cells (CSCs) are responsible for cancer metastasis by stimulating tumor angiogenesis via various mechanisms. To elucidate the potential of the stemness-high human colorectal cancer (CRC) cells (i.e., CRCSCs) in activating angiogenesis, effects of the GATA6-overexpressing HCT-116 and HT-29 human CRC clones established previously by us in promoting the angiogenesis of human umbilical vein endothelial cells (HUVECs) were examined.

methodsAngiogenesis-promoting effects (i.e., migration, invasion, DNA synthesis, and tube formation) in HUVECs of the conditioned media (CM) from various human CRC clones were analyzed. MMP activities were assessed using a zymography assay. Western blotting and selective inhibitors were used to dissect the signaling pathway involved. IHC was used to examine the vascular density in tumor xenografts.

resultsWe found that the conditioned media (CM) collected from the GATA6-overexpressing clones enhanced angiogenesis of HUVECs more effectively which might be attributed partly to a higher MMP-9 production by HUVECs. Subsequently, elevated levels of IL-8 and VEGF-A were detected in the CM whose tube formation-enhancing activities were abolished by the co-treatment with either a VEGFR2 inhibitor or an IL-8 neutralizing antibody. Interestingly, increased production of these cytokines in the GATA6-overexpressing clones was due to an EGFR/AKT-mediated activation of NF-κB. Furthermore, not only were the levels of CD31 and endomucin but also the blood vessel density was much higher in the xenograft tumors grown from these clones.

conclusionOur findings demonstrate that human CRCSCs promote a stronger angiogenesis by producing higher amounts of angiogenic factors through activation of the EGFR/AKT/NF-κB pathway.