Amplification of Lipopolysaccharide-Induced Cytokine Synthesis in Non–Small Cell Lung Cancer/Neutrophil Cocultures

Abstract

Proinflammatory cytokines are centrally involved in tumor progression and survival in non-small cell lung cancer, and both the presence of infiltrating neutrophils and bacterial infection in the lung may indicate a poor prognosis. Against this background, we investigated the effect of the bacterial cell wall component lipopolysaccharide (LPS) on interleukin (IL)-6 and IL-8 synthesis in the non-small cell lung cancer line A549 and in A549-neutrophil cocultures. The LPS induced a dose-dependent and time-dependent release of IL-8 from A549 cells, whereas IL-6 could not be detected. Interestingly, in A549-neutrophil cocultures, IL-8 synthesis was massively amplified and IL-6 was also released, compared with the respective monocultures. The A549 cells were identified as the primary cellular source of these cytokines, as enhanced cytokine mRNA transcription was detected in this cell type, although not in neutrophils in the coculture system. Experiments done in transwells indicated that direct cell-cell contact was a prerequisite for the increased cytokine generation. Inhibition of tumor necrosis factor-α bioactivity by neutralizing antibodies and blocking cyclooxygenase-2 activity blunted the enhanced cytokine generation in the coculture system. Amplification of LPS-induced cytokine secretion could be reproduced when the small cell lung cancer cell line H69 was cocultured with neutrophils. When the Gram-positive cell wall component lipoteichoic acid was used instead of LPS, cytokine synthesis was also amplified in A549-neutrophil cocultures, to a similar extent to that observed with LPS. These data indicate that interaction between bacterial pathogens, neutrophils, and tumor cells might amplify the release of proinflammatory cytokines which may promote tumor growth in vivo.

Endotoxin induces proliferation of NSCLC in vitro and in vivo: role of COX-2 and EGFR activation

“…Direct activation of this receptor system and downstream signaling events such as ERK and JNK activation in response to LPS have been described [55, 56]. In addition, we and others have demonstrated that LPS activates IL-8 synthesis in A549 cells [57, 58], and this cytokine is known to transactivate EGFR in NSCLC cell lines as an alternative pathway [59]. …”

Lipoteichoic acids from Staphylococcus aureus stimulate proliferation of human non-small-cell lung cancer cells in vitro

“…At the end of the incubation period, cell supernatants were harvested, cell debris was removed by centrifugation at 13.000× g and samples were stored at −20 °C until further processing. Release of IL-8 was determined in a direct sandwich ELISA, as described previously [33]. To normalize the data, IL-8 was expressed as ΔIL-8, meaning that baseline levels of IL-8 secreted from unstimulated controls were subtracted from those induced by stimulation with LTA.…”

Morphine Attenuates Endothelial Cell Adhesion Molecules Induced by the Supernatant of LPS-Stimulated Colon Cancer Cells

“…Furthermore, IL-8, IL-6, and vascular endothelial growth factor are promoted by LPS in ovarian cancer cells (3) and IL-8 synthesis is increased by LPS stimulation in non-small cell lung cancer cells (4). Moreover, it is known that IL-1β and IL-6 are highly induced by tumors that metastasize to the liver, such as breast cancer and colon carcinoma (18).…”

“…A large reservoir of LPS is available in the colon, because bacteria like Escherichia coli typically produce copious amounts of LPS, and thus inflammation around colon cancer cells can cause much LPS release. This LPS stimulates to secrete various cytokines and pro-inflammatory mediators (3, 4). The effects of cytokines and pro-inflammatory mediators on adhesion molecules expressions are not clear, but in a previous study, it was demonstrated that the supernatant of LPS-stimulated HT-29 colon cancer cells up-regulates adhesion molecules on endothelial cells.…”