c KaplanCMeier curves showing that high expression of IL-8 in two microarray data sets are positively associated with poor patient survival. with hypoxic CRC cells or treated with hypoxic CRC cell-derived CM, normoxic CRC cells possessed increased metastatic capacity. Furthermore, hypoxic CRC cell-derived CM was enriched in interleukin 8. Hypoxic CRC cell-derived CM and recombinant human IL-8 both USP7/USP47 inhibitor enhanced the metastatic capacity of normoxic cells by increasing the phosphorylation of p65 and then by inducing epithelial-mesenchymal transition. Knockdown USP7/USP47 inhibitor of IL-8 in hypoxic CRC cells or the use of an anti-IL-8 antibody attenuated the CM- or rhIL-8-induced prometastatic capacity of normoxic CRC cells. Inhibition or knockdown of p65 abrogated IL-8-induced prometastatic effects. Most importantly, hypoxia-treated xenograft tumors enhanced the metastasis of normoxic CRC cells. Hypoxic CRC cell-derived IL-8 promotes the metastatic capacity of normoxic cells, and novel therapies targeting the positive interactions between hypoxic and normoxic cells should be developed. USP7/USP47 inhibitor test for two groups. Where more than two groups were compared, one-way analysis of variance was used. A value of P? LIPO that HSS CRC cells expressed higher mRNA levels of matrix metalloproteinase, such as MMP1, MMP2, and membrane type 1-matrix metalloproteinase 1 (MT1MMP) than normoxic CRC cells (i.e., Control) USP7/USP47 inhibitor (Fig. S1B). We then performed Transwell invasion assays and demonstrated that hypoxic CRC cells possessed increased invasive capacity (Fig. ?(Fig.1c).1c). Next, we injected hypoxic and normoxic CRC cells into the tail vein of the NOD/SCID mice. Eight weeks later, hypoxic CRC cells were found to have formed more metastatic lesions than normoxic CRC cells in the lungs of the mice (Fig. ?(Fig.1d).1d). Thus, our findings suggest that hypoxic CRC cells possess high lung metastatic capacity. Open in a separate window Fig. 1 Hypoxic CRC cells possess higher metastatic capacity than normoxic CRC cells.a Schematic of the in vitro physical hypoxic treatment of CRC cells. b Immunoblot analysis of HIF1 in hypoxic CRC cells. Normoxic CRC cells as control, and -actin for loading control. c Transwell invasion assays. In all, 4??104 hypoxic (HSS) and normoxic (Control) CRC cells were incubated, invaded cells were quantified. Scale bars: 200?m. Mean??SD from triple experiments. *P?P?n?=?5 per group). Data are presented as mean??SD. ***P?n?=?3). *P?P?P?n?=?3), ***P?n?=?4 per.