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We assessed the effect of voriconazole (VRC) on the expression and release of selected cytokines and chemokines in the THP-1 human monocytic cell line in response toAspergillus fumigatushyphal fragments (HF) by cDNA microarray analysis, reverse transcriptase (RT) PCR, and enzyme-linked immunosorbent assay. Stimulation of THP-1 cells by HF alone caused a significant up-regulation ofCCL4(MIP1B) andCCL16, whileCCL2(MCP1) was down-regulated. By comparison, in the presence of VRC, a large number of genes such asCCL3(MIP1A),CCL4(MIP1B),CCL5(RANTES),CCL7(MCP3),CCL11(EOTAXIN),CCL15(MIP1Δ),CXCL6, andCXCL13were strongly up-regulated in THP-1 cells challenged by HF, whereasCCL20(MIP3A) andCCL21(MIP2) were down-regulated. Among five genes differentially expressed in THP-1 cells,IL12A,IL12B, andIL-16were down-regulated whereasIL-11andTGFB1were significantly up-regulated in the presence of VRC. The inflammation-related genesIFNγ,IL1R1, andTNFAwere also up-regulated in THP-1 cells exposed to HF only in the presence of VRC. RT-PCR of four selected genes validated the results of microarrays. The release of interleukin 1β (IL-1β) and IL-12 was significantly increased from monocytes stimulated either by HF alone (P< 0.05) or in the presence of VRC (P< 0.01 andP< 0.05, respectively). In contrast, tumor necrosis factor alpha release from monocytes was enhanced only in the presence of VRC (P< 0.01). The chemokines monocyte chemoattractant protein 1 and macrophage inflammatory protein 1β were decreased under both conditions (P< 0.01). These results demonstrate that in the presence of VRC, HF induces a more pronounced profile of gene expression in THP-1 cells than HF alone, potentially leading to more-efficient host resistance toA. fumigatus.

Pseudomonas aeruginosa, an opportunistic pathogen, causes infections associated with a high incidence of morbidity and mortality in immunocompromised hosts. Production of tumor necrosis factor alpha (TNF-α), primarily by cells of monocytic lineage, is a crucial event in the course of these infections. During in vivo infections withP. aeruginosa, both lipopolysaccharide (LPS) and extracellular slime glycolipoprotein (GLP) produced by mucoid and nonmucoid strains are released. In the present study, we sought to explore the relative contributions of these two bacterial products to TNF-α production by human monocytes. To this end, fresh human monocytes and THP-1 human monocytic cells were stimulated withP. aeruginosaLPS or GLP. GLP was found to be a more potent stimulus for TNF-α production (threefold higher) by human monocytes than LPS. Moreover, its effect was comparable to that of viable bacteria. Quantitative mRNA analysis revealed predominantly transcriptional regulation. Electrophoretic mobility shift assays and transfection assays demonstrated activation of NF-κB and activator protein 1 (AP-1). NF-κB activation by GLP was rapid and followed the same time course as that by viable bacteria, suggesting that bacteria could directly activate NF-κB through GLP. MoreoverP. aeruginosaGLP induced the formation of AP-1 complex with delayed kinetics compared with NF-κB but much more efficiently than the homologous LPS. These results identify GLP as the most important stimulant for TNF-α production by human monocytes. Activation of NF-κB and AP-1 byP. aeruginosaGLP may be involved not only in TNF-α induction but also in many of the inflammatory responses triggered in the course of infection withP. aeruginosa.