IL-8 Production in Human Lung Fibroblasts and Epithelial Cells Activated by the Pseudomonas Autoinducer N-3-Oxododecanoyl Homoserine Lactone Is Transcriptionally Regulated by NF-κB and Activator Protein-2

Abstract

The destructive pulmonary inflammation associated with Pseudomonas aeruginosa colonization is caused, in part, by the production of the chemokine IL-8, which recruits neutrophils into the lung. The Pseudomonas autoinducer, N-3-oxododecanoyl homoserine lactone (3-O-C12-HSL), is a small lipid-soluble molecule that is essential in the regulation of many P. aeruginosa virulence factors, but little is known about how it affects eukaryotic cells. In this report we demonstrate that 3-O-C12-HSL is a potent stimulator of both IL-8 mRNA and protein from human fibroblasts and epithelial cells in vitro. The IL-8 produced from these 3-O-C12-HSL-stimulated cells was found to be functionally active by inducing the chemotaxis of neutrophils. To determine a mechanism for this IL-8 induction, deletion constructs of the IL-8 promoter were examined. It was found that the DNA region between nucleotides −1481 and −546 and the transcription factor NF-κB were essential for the maximal induction of IL-8 by 3-O-C12-HSL. This was confirmed by EMSAs, where 3-O-C12-HSL induced a shift with both AP-2 and NF-κB consensus DNA. The activation of NF-κB and subsequent production of IL-8 were found to be regulated by a mitogen-activated protein kinase pathway. These findings support the concept that the severe lung damage that accompanies P. aeruginosa infections is caused by an exuberant neutrophil response stimulated by 3-O-C12-HSL-induced IL-8. Understanding the mechanisms of 3-O-C12-HSL activation of lung structural cells may provide a means to help control lung damage during infections with P. aeruginosa.

N-(3-Oxo-acyl)homoserine Lactones Signal Cell Activation through a Mechanism distinct from the Canonical Pathogen-associated Molecular Pattern Recognition Receptor Pathways

“…The abilities of this Gram-negative bacterium to survive and establish chronic infections in the host appear to be associated with functions of C12 (20,21). Interestingly, C12 has also been reported to exhibit effects on eukaryotic cells (15,21) including inhibition of cell proliferation (22,23), induction of apoptosis (23,24), and nuclear translocation of NF-B (15,25). These observations suggest that C12 may function as a PAMP.…”

“…Consistent with the observation that SV40 large T antigen alters the functions of p53 (57, 61), we found that C12 induces Cox-2 mRNA in SV40-transformed WI-38 cells but not in primary cells. Analogous to C12-mediated transcriptional response in WI-38 cells, Cox-2 and IL-8 induction was previously observed after C12 treatment of immortalized or transformed human epithelial cells and lung fibroblasts (25,62). Although the integrity of p53 was not tested in these cells, immortalization and transformation may be a result of mutations in the p53 gene (57,61).…”

Mouse and Human Cell Activation by N -Dodecanoyl- dl -Homoserine Lactone, a Chromobacterium violaceum Autoinducer

“…Apart from its quorum-sensing function, the P. aeruginosa N-3-oxododecanoyl-homoserine lactone (3-oxo-C 12 -HSL) autoinducer has been suggested to modulate the immune responses of the infected host. In this context, it was demonstrated that 3-oxo-C 12 -HSL induces the production of interleukin-8 (IL-8), cyclooxygenase 2, and prostaglandin E2 in human fibroblasts and that it accelerates apoptosis in mammalian macrophages and neutrophils (15,16,18).…”

Pseudomonas aeruginosa Homoserine Lactone Activates Store-operated cAMP and Cystic Fibrosis Transmembrane Regulator-dependent Cl− Secretion by Human Airway Epithelia

“…3O-C12 operates through TLR-and Nod/Ipaf/caterpillarindependent signaling to activate multiple proinflammatory genes that are associated with NF-B signaling, including IL8, Cox2, and MUC5AC in both epithelial and other cell types (7)(8)(9)(10)(11)(12). Some of these proinflammatory effects may be mediated through activation of MAPKs (8,9) or Ca 2ϩ (7,13) or inhibition of peroxisome proliferator-activated receptor ␥ (14).…”