Recognition of conserved pathogen‐associated molecular patterns constitutes a crucial step in the initiation of innate immune responses. We studied the contribution to the host‐pathogen interaction of mycolic acid (MA), a major lipid component of the cell envelope of the macrophage intracellular pathogen Mycobacterium tuberculosis and other mycobacteria. MA administered to the peritoneal cavity or to the airways induced a unique macrophage morphotype, similar to the foamy macrophage derivatives observed in tuberculous granulomas and characterized by intracellular accumulation of neutral lipids and entry into mitosis. When assayed for production of inflammatory mediators, a conditioning rather than a direct activation of the MA‐elicited foamy macrophages was observed. MA enabled production of IFN‐γ and myeloperoxidase, enhanced TNF‐α production and suppressed IL‐10 upon renewed exposure to innate triggers. Intratracheal instillation of MA mimicked additional features of the airway response to M. tuberculosis infection, namely a rapid but transient neutrophil influx and IL‐6 production and a chronic IL‐12 production. These MA‐elicited cellular innate defenses and the accompanying formation of foamy macrophages identify for the first time the foamy macrophage morphotype as part of the host response to a pathogen‐associated structure. Furthermore, these results characterize MA as a direct trigger of innate immunity, distinct from Toll‐like receptor ligands.

Rationale: Mycolic acid (MA) constitutes a major and distinguishing cell wall biolipid from Mycobacterium tuberculosis. MA interferes with the lipid homeostasis of alveolar macrophages, inducing differentiation into foamy macrophages exhibiting increased proinflammatory function. Objectives: We verified the interference of this altered macrophage function with inhaled antigen–triggered allergic airway inflammation and underlying Th2 lymphocyte reactivity. Methods: Using ovalbumin (OVA) as model allergen, C57BL/6 or BALB/C mice were sensitized by OVA-alum immunization. Experimental asthma, triggered subsequently by repetitive nebulized OVA inhalation, was assessed, using as readout parameters eosinophilia, peribronchial inflammation, and Th2 cytokine function. Measurements and Main Results: A single intratracheal treatment of sensitized mice with MA, inserted into liposomes as carriers, prevented the onset of OVA-triggered allergic airway inflammation and promoted unresponsiveness to a secondary set of allergen exposures. The development of this tolerant condition required an 8-d lapse after MA instillation, coinciding with the appearance of foamy alveolar macrophages. MA-conditioned CD11b+F4/80+ macrophages, transferred to the airways, mimicked the tolerogenic function of instilled MA; however, without the 8-d lapse requirement. Indicative of a macrophage-mediated tolerogenic antigen-presenting function, major histocompatibility complex (MHC)–mismatched donor macrophages failed to promote tolerance. Furthermore, Treg markers were strongly increased and established tolerance was lost after in situ depletion of CD25+ Treg cells. Contrary to the interleukin-10 dependence of tolerogenic dendritic cells, IFN-γ deficiency but not interleukin-10 deficiency abrogated the tolerogenic capacity of MA-conditioned macrophages. Conclusions: These results document an innate-driven Mycobacterium tuberculosis MA-triggered immune regulatory mechanism in control of pulmonary allergic responses by converting macrophages into IFN-γ–dependent tolerogenic antigen-presenting cells.

The appearance of drug-resistant strains of Mycobacterium tuberculosis (Mtb) poses a great challenge to the development of novel treatment programmes to combat tuberculosis. Since innovative nanotechnologies might alleviate the limitations of current therapies, we have designed a new nanoformulation for use as an anti-TB drug delivery system. It consists of incorporating mycobacterial cell wall mycolic acids (MA) as targeting ligands into a drug-encapsulating Poly dl-lactic-co-glycolic acid polymer (PLGA), via a double emulsion solvent evaporation technique. Bone marrow-derived mouse macrophages, either uninfected or infected with different mycobacterial strains (Mycobacterium avium, Mycobacterium bovis BCG or Mtb), were exposed to encapsulated isoniazid-PLGA nanoparticles (NPs) using MA as a targeting ligand. The fate of the NPs was monitored by electron microscopy. Our study showed that i) the inclusion of MA in the nanoformulations resulted in their expression on the outer surface and a significant increase in phagocytic uptake of the NPs; ii) nanoparticle-containing phagosomes were rapidly processed into phagolysosomes, whether MA had been included or not; and iii) nanoparticle-containing phagolysosomes did not fuse with non-matured mycobacterium-containing phagosomes, but fusion events with mycobacterium-containing phagolysosomes were clearly observed.