(537) GM-CSF is Essential for Effective Macrophage Killing of Nontuberculous Mycobacteria

Background: People with CF (PwCF) are at increased risk for developing pulmonary nontuberculous mycobacteria (pNTM) infections. Eradication of pNTM is often unsuccessful, even with months of antibiotic therapy. A better understanding of host factors that predispose PwCF to pNTM will identify those at risk of pNTM and could reveal targets for immunomodulatory therapies to enhance traditional antimicrobials. Critical knowledge gaps exist surrounding acute events that occur when inhaled NTM initially interact with airway macrophages. In the lung, macrophages can be grouped into two main categories: resident airspace macrophages (AMs) and recruited monocyte-derived macrophages (MDMs). Most macrophages in bronchiectatic airways of PwCF are MDMs recruited from the bloodstream, a compartment with low levels of GM-CSF. In contrast, macrophages that first interface with NTM in healthy lungs are AMs, which are exposed to high levels of GM-CSF produced by alveolar epithelial cells. This study investigates the hypothesis that macrophages must be programmed by GM-CSF to effectively kill NTM, and MDMs recruited to the airways in PwCF are less effective at killing NTM than resident AMs due to insufficient exposure to GM-CSF.

Methods: Human MDMs were cultured in M-CSF or GM-CSF and then assayed for ability to ingest and kill Mycobacterium abscessus (MAbsc). GM-CSF knock-out (KO) mice and wild type (WT) were infected by oropharyngeal (op) inoculation with MAbsc. WT mice were infected op with MAbsc-embedded agar beads, which results in small airways infection, and compared over 7 weeks to mice infected with non-bead embedded “planktonic” MAbsc.

Results: MDMs matured in M-CSF demonstrated minimal ability to kill MAbsc, but MDMs exposed to GM-CSF effectively kill MAbsc

(p < 0.001). At 6 days post infection, GM-CSFKO mice contained > 3 log more bacteria in their lungs than WT mice (p < 0.0005), despite recruitment of both macrophages and neutrophils to the airspaces. Cytospins of day 6 bronchoalveolar lavage (BAL) cells revealed that macrophages from GM-CSF KO mice ingested MAbsc, but failed to control bacterial replication, whereas WT BAL macrophages contained copious MAbsc at day 3 post infection, but were almost devoid of MAbsc by day 6. MAbsc
-embedded agar beads lodged in small airways allowed local diffusion of bacteria resulting in recruitment of neutrophils and macrophages to the airway. As bacterial burden increased, chronic infection was established with higher bacterial burden at all time points than mice infected with planktonic MAbsc (p < 0.0001). Small airway MAbsc bead infection also resulted in significant differences in proportions and absolute numbers of resident AMs and recruited MDMs in the lungs.

Conclusions: These data support a novel model of human susceptibility to pNTM in which bronchiectactic airways recruit macrophages that are incapable of effective killing of inhaled NTM, leading to chronic infection. These data additionally suggest that immunomodulatory therapy with GM-CSF could enhance bacterial killing by MDMs recruited to the airways and assist with eradication of pNTM that persists despite intense antimicrobial therapies.

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