Melanoma Inhibits Macrophage Activation by Suppressing Toll-like Receptor 4 Signaling

Background

Activated macrophages defend against tumors by secreting cytokines to recruit secondary immune cells, presenting antigen to T cells, and by direct tumor cytotoxicity. Peritoneal macrophages harvested from melanoma-bearing mice are less cytotoxic to melanoma cells, and produce less superoxide, nitric oxide, and tumor necrosis factor-α (TNF-α) than those from nontumor-bearing mice. Similar impairment of macrophage activation occurs in vitro using media harvested from cultured melanoma cells. Stimulation of Toll-like receptor 4 (TLR-4) activates macrophages and results in the release of TNF-α. We hypothesized that melanoma inhibits macrophage activation by suppressing TLR-4 signaling.

Study design

Melanoma conditioned media (MCM) was generated from B16 melanoma cells. Peritoneal macrophages from TLR-4 competent or TLR-4 incompetent mice were exposed to control or MCM for 24hours; then stimulated with lipopolysaccharide. TNF-α secretion, TNF-α mRNA production, nuclear factor-κB (NF-κB) activation, and TLR-4 surface expression were measured.

Results

Peritoneal macrophages exposed to MCM produced considerably less TNF-α in response to stimulus than controls (691 pg/mL versus 2,066 pg/mL, p < 0.001). TNF-α production by TLR-4 incompetent macrophages was not affected by MCM (454 pg/mL versus 480 pg/mL). Stimulated TNF-α mRNA and activated NF-κB were decreased in MCM treated C57BL/6 macrophages (by 38% and 33%, respectively). TLR-4 surface expression, however, was not decreased by exposure to MCM.

Conclusions

Melanoma inhibits macrophage activation by suppressing TLR-4 signaling downstream of the TLR-4 receptor.

Abbreviations and Acronyms:  αMSH, alpha-melanocyte stimulating hormone , DMEM, Dulbecco’s modified Eagle’s medium , FBS, fetal bovine serum , LPS, lipopolysaccharide , MCM, melanoma conditioned media , NF-κB, nuclear factor-κB , NO, nitric oxide , RNA, Ribonucleic acid , TLR, Toll-like receptor , TNF-α, tumor necrosis factor-α