Additional Supporting Information may be found in the online version of this article.

HEP_25907_sm_SuppFig1.tif1735KSupporting Information Figure 1. CXCL5 promotes migration and invasion of HCC cells in vitro. (A) In wound-healing migration assays, microscopic examination at 24 hours post-wounding revealed a significant delay in the wound closure rate of HCCLM3-shRNA-CXCL5 and MHCC97H-shRNA-CXCL5 cells compared with the control cell lines, whereas, HepG2-CXCL5 and PLC/PRF/5-CXCL5 cells had a significant increase in the wound closure rate compared with control cell lines (B). (C) In vitro invasive assays showed that the numbers of invasive HCCLM3-Mock and MHCC97H-Mock cells were significantly higher than those of HCCLM3-shRNA-CXCL5 and MHCC97H-shRNA-CXCL5 cells. (D) The numbers of invasive HepG2-CXCL5 and PLC/PRF/5-CXCL5 cells were significantly higher than those of HepG2-Mock and PLC/PRF/5-Mock cells. Magnification, 100× (A-B) and 200× (C-D).
HEP_25907_sm_SuppFig2.tif197KSupporting Information Figure 2. CXCL5 attracts neutrophils in vitro. (A) In vitro transwell assays show that CXCL5 induces concentration-dependent migration of neutrophils. (B) Western blot analysis of p-Akt and p-p65 in response to CXCL5.
HEP_25907_sm_SuppFig3.tif378KSupporting Information Figure 3. Statistical analysis of tumor volume, intratumoral neutrophil number, and pulmonary metastases in xenografts from nude mice. (A) The tumor volume of the xenograft in each nude mouse group. (B) The number of intratumoral neutrophils in each group of xenograft nude mice. (C-F) All grades of metastatic nodules in lungs of all xenograft nude mice groups.
HEP_25907_sm_SuppFig4.tif9004KSupporting Information Figure 4. Representative images of Gr1+ intratumoral neutrophils with magnification of the selected areas in Fig. 4. Magnification, 200×.
HEP_25907_sm_SuppFig5.tif271KSupporting Information Figure 5. Kaplan-Meier analysis of OS rates (A-B) and cumulative recurrence rates (C-D) in 94 HCCs (cohort 1) with different CXCL5 expression levels.
HEP_25907_sm_SuppFig6.tif315KSupporting Information Figure 6. The correlation and prognostic value of CXCL5 and intratumoral CD66b+ neutrophils in HCC samples (cohort 3, n=502). (A-C) Prognostic value of CXCL5 expression and intratumoral CD66b by Kaplan-Meier analysis. I, CXCL5Low/intratumoral CD66bLow; II, CXCL5Low/intratumoral CD66bHigh and CXCL5High/intratumoral CD66bLow; III, CXCL5High/intratumoral CD66bHigh. (D) Significant positive correlation between CXCL5 expression and intratumoral CD66b in cancerous tissues, shown with a scatter plot.
HEP_25907_sm_SuppFig7.tif1900KSupporting Information Figure 7. Representative images of intratumoral lymphocytes. (A) CD3+, (B) CD4+, (C) CD8+, (D) CD45RO+, (E) Foxp3+, and (F) granzyme B+. Magnification, 200×.
HEP_25907_sm_SuppTab1.doc41KSupporting Information Table 1. Up-regulated at least one-fold of protein phosphorylation after CXCL5 treatment in human neutrophils.
HEP_25907_sm_SuppTab2.doc43KSupporting Information Table 2. Clinicopathologic characteristics of three cohorts of patients with hepatocellular carcinoma.
HEP_25907_sm_SuppTab3.doc51KSupporting Information Table 3. Correlation between the factors and clinicopathologic characteristics in HCC (cohort 1, n=94).
HEP_25907_sm_SuppTab4.doc50KSupporting Information Table 4. Univariate and multivariate analyses of prognostic factors in HCC (cohort 1, n=94).
HEP_25907_sm_SuppTab5.doc48KSupporting Information Table 5. Correlation between the factors and clinicopathologic characteristics in HCC (cohort 3, n=502).
HEP_25907_sm_SuppTab6.doc60KSupporting Information Table 6. Univariate and multivariate analyses of prognostic factors in HCC (cohort 3, n=502).
HEP_25907_sm_SuppTab7.doc42KSupporting Information Table 7. Univariate analyses of intratumoral lymphocytes in HCC (cohort 2, n=323 and cohort 3, n=502).
HEP_25907_sm_SuppTab8.doc40KSupporting Information Table 8. Descriptive statistics of immunohistochemical variables of intratumoral lymphocytes in HCC and correlations with CD66b/CXCL5 (cohort 2, n=323 and cohort 3, n=502).
HEP_25907_sm_SuppInfo.doc97KSupporting Information

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