Gram-Negative Pneumonia Augments Non-Small Cell Lung Cancer Metastasis through Host Toll-like Receptor 4 Activation

Abstract

Introduction: Surgery is essential for cure of early-stage non-small cell lung cancer (NSCLC). Rates of postoperative bacterial pneumonias, however, remain high, and clinical data suggests that post-operative infectious complications confer an increased risk for metastasis. Toll-like receptors (TLRs) mediate the inflammatory response to infection by recognizing evolutionarily conserved bacterial structures at the surface of numerous pulmonary cell types; yet, little is known about how host TLR activation influences NSCLC metastasis. TLR4 recognizes gram-negative bacterium lipopolysaccharide activating the innate immune system.

Methods: C57BL/6 and TLR4 knockout murine airways were inoculated with Escherichia coli or lipopolysaccharide. Hepatic metastasis assays and intravital microscopy were performed. Bronchoepithelial conditioned media was generated through coincubation of bronchoepithelial cells with TLR4 activating Escherichia coli or lipopolysaccharide. Subsequently, H59 NSCLC were stimulated with conditioned media and subject to various adhesion assays.

Results: We demonstrate that gram-negative Escherichia coli pneumonia augments the formation of murine H59 NSCLC liver metastases in C57BL/6 mice through TLR4 activation. Additionally, infected C57BL/6 mice demonstrate increased H59 NSCLC in vivo hepatic sinusoidal adhesion compared with negative controls, a response that is significantly diminished in TLR4 knockout mice. Similarly, intratracheal injection of purified TLR4 activating lipopolysaccharide increases in vivo adhesion of H59 cells to murine hepatic sinusoids. Furthermore, H59 cells incubated with bronchoepithelial conditioned medium show increased cell adhesion to in vitro extracellular matrix proteins and in vivo hepatic sinusoids through a mechanism dependent on bronchoepithelial TLR4 activation and interleukin-6 secretion.

Conclusion: TLR4 is a viable therapeutic target for NSCLC metastasis augmented by gram-negative pneumonia.

Keywords: Cancer metastasis; Non–small cell lung cancer; Pneumonia; Toll-like receptors.