We previously found that the tryptophan catabolite picolinic acid (PA) is a costimulus for the activation of macrophage effector functions. In this study, we have investigated the ability of PA to modulate the expression of chemokines in macrophages. We demonstrate that PA is a potent activator of the inflammatory chemokines MIP (macrophage inflammatory protein)-1α and MIP-1β (MIPs) mRNA expression in mouse macrophages in a dose- and time-dependent fashion and through a de novo protein synthesis-dependent process. The induction by PA occurred within 3 h of treatment and reached a peak in 12 h. The stimulatory effects of PA were selective for MIPs because other chemokines, including monocyte chemoattractant protein-1, RANTES, IFN-γ-inducible protein-10, MIP-2, and macrophage-derived chemokine, were not induced under the same experimental conditions and were not an epiphenomenon of macrophage activation because IFN-γ did not affect MIPs expression. Induction of both MIP-1α and MIP-1β by PA was associated with transcriptional activation and mRNA stabilization, suggesting a dual molecular mechanism of control. Iron chelation could be involved in MIPs induction by PA because iron sulfate inhibited the process and the iron-chelating agent, desferrioxamine, induced MIPs expression. We propose the existence of a new pathway leading to inflammation initiated by tryptophan catabolism that can communicate with the immune system through the production of PA, followed by secretion of chemokines by macrophages. These results establish the importance of PA as an activator of macrophage proinflammatory functions, providing the first evidence that this molecule can be biologically active without the need for a costimulatory agent.

Activation of murine macrophages (Mφ) requires the collaboration of signals derived from the immune system and the environment. In this study, we engineered a murine Mφ cell line to become activated in response to an environmental signal, hypoxia, as the sole stimulus. Hypoxia is a condition of low oxygen tension, occurring in several pathological tissues, which acts in synergy with IFN-γ to induce full Mφ activation. We transfected the ANA-1 murine Mφ cell line with a construct containing the IFN-γ gene controlled by a synthetic promoter inducible by hypoxia (HRE3x-Tk), and we characterized the cellular and molecular biology of the engineered Mφ under normoxia or hypoxia. Engineered Mφ in normoxia expressed basal levels of IFN-γ mRNA and protein that were strongly augmented by shifting the cells to hypoxia. Furthermore, they responded to the synthesized IFN-γ with induction of IFN-responsive factor-1 and 2′-5′-oligoadenylate synthase expression. Under normoxic conditions, the engineered Mφ had a significant constitutive level of Ia Ags and Fc receptors. Hypoxia induced further augmentation of Ia and Fc expression. Finally, hypoxia induced inducible NO synthase expression, and subsequent reoxygenation led to the production of NO. In conclusion, the engineered Mφ, which produce IFN-γ in an inducible manner, express new biochemical and functional properties in response to low oxygen environment as the sole stimulus, thereby circumventing the need for costimulation by other immune system-derived signals.

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