Up-Regulation of Tumor Interleukin-8 Expression by Infiltrating Macrophages

Up-Regulation of Tumor Interleukin-8 Expression by Infiltrating Macrophages

Its Correlation with Tumor Angiogenesis and Patient Survival in Non-Small Cell Lung Cancer1

Institute of Biomedical Sciences and Molecular Biology, National Chung Hsing University, Taichung, Taiwan 402 [J. J. W. C.]; Departments of Medical Research [J. J. W. C.], Internal Medicine [P-L. Y., A. Y., P-C. Y.], Surgery [Y-C. L.], and Forensic Medicine [C. T. S.], School of Pharmacy [T-M. H.], and Institute of Toxicology [M-L. K.], National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, Taiwan 100; and Institute of Biomedical Sciences, Academia Sinica [P-C. Y.] and National Health Research Institute [P-C. Y.], Taipei, Taiwan 115, Republic of China

Abstract

Purpose: To evaluate the interaction between tumor-infiltrating macrophages and cancer cells and its effect on the expression of a potent angiogenic factor, interleukin-8 (IL-8), tumor angiogenesis, and patient outcome in non-small cell lung cancer (NSCLC).

Experimental Design: We measured tumor IL-8 mRNA expression (by real-time quantitative reverse transcription-PCR), intratumor microvessel counts, and tumor-infiltrating macrophage density (by immunohistochemical staining) in 35 NSCLC surgical specimens and correlated with the patient’s clinical outcome. We then investigated the interaction between macrophages (cell line THP-1) and six different human cancer cell lines (four NSCLCs, one osteosarcoma, and one hepatoma) and its effect on IL-8 mRNA expression using a macrophage/cancer cell coculture system, IL-8 mRNA expression in lung cancer cells, and macrophages being measured separately after coculture in the presence or absence of six anti-inflammatory agents, i.e., pentoxifylline, aspirin, indomethacin, dexamethasone, celecoxib (a selective cyclooxygenase-2 inhibitor), and pyrrolidine dithiocarbamate, a specific nuclear factor κB (NF-κB) inhibitor. NF-κB transcriptional activity and protein levels were measured by reporter gene assay and Western blot.

Results: The tumor-infiltrating macrophage density correlated significantly and positively with tumor IL-8 mRNA expression and intratumor microvessel counts and significantly and negatively with patient survival. In addition, after cell–cell interaction in cancer cell:macrophage cocultures, marked IL-8 mRNA expression was induced in lung cancer cells (∼270-fold) and, to a lesser degree, in macrophages (4.5-fold). The increase in IL-8 mRNA expression correlated with the in vitro metastatic potential of the cancer cells. All six anti-inflammatory agents suppressed induction of IL-8 mRNA expression in lung cancer cells by >90%, four (pentoxifylline, celecoxib, pyrrolidine dithiocarbamate, and dexamethasone) having a dose-dependent effect. NF-κB transcriptional regulation and protein levels were simultaneously increased in the nuclei of cancer cells in macrophage/cancer cell cocultures, this effect also being suppressed by all six anti-inflammatory agents.

Conclusions: The interaction between infiltrating macrophages and cancer cells up-regulates IL-8 mRNA expression, especially in the cancer cells; this may contribute greatly to the increased tumor angiogenesis and adverse outcome in NSCLC patients with a high density of tumor-infiltrating macrophages. Anti-inflammatory agents can suppress the induction of IL-8 mRNA expression seen in lung cancer cells after coculture with macrophages, and this suppression is mediated, in part, through the NF-κB pathway.

Footnotes

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↵1 Supported by NSC 91-3112-P-005-008-Y (Taipei, Taiwan, ROC).
↵2 J. J. W. C., P-L. Y., and A. Y. contributed equally to this study.
↵3 To whom requests for reprints should be addressed, at Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan 100, Republic of China. Phone: 886-2-2356-2116; Fax: 886-2-2358-2867; E-mail: pcyang{at}ha.mc.ntu.edu.tw
↵4 The abbreviations used are: IL, interleukin; NF-κB, nuclear factor κB; NSCLC, non-small cell lung cancer; FBS, fetal bovine serum; VEGF, vascular endothelial growth factor; RTQ-RT-PCR, real-time quantitative reverse transcription-PCR; IHC, immunohistochemical staining; PDTC, pyrrolidine dithiocarbamate; TBP, TATA box binding protein; TNF, tumor necrosis factor; ATCC, American Type Culture Collection; CT, threshold cycle; COX, cyclooxygenase; CI, confidence interval; NSAID, nonsteroidal anti-inflammatory drug.
- Accepted October 16, 1902.
- Received May 7, 1902.
- Revision received October 10, 1902.