In vitro differentiation of human macrophages with enhanced antimycobacterial activity
Guillaume Vogt, Carl Nathan
Guillaume Vogt, Carl Nathan
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Technical Advance Infectious disease

In vitro differentiation of human macrophages with enhanced antimycobacterial activity

Abstract

Mycobacterium tuberculosis causes widespread, persistent infection, often residing in macrophages that neither sterilize the bacilli nor allow them to cause disease. How macrophages restrict growth of pathogens is one of many aspects of human phagocyte biology whose study relies largely on macrophages differentiated from monocytes in vitro. However, such cells fail to recapitulate the phenotype of tissue macrophages in key respects, including that they support early, extensive replication of M. tuberculosis and die in several days. Here we found that human macrophages could survive infection, kill Mycobacterium bovis BCG, and severely limit the replication of M. tuberculosis for several weeks if differentiated in 40% human plasma under 5%–10% (physiologic) oxygen in the presence of GM-CSF and/or TNF-α followed by IFN-γ. Control was lost with fetal bovine serum, 20% oxygen, M-CSF, higher concentrations of cytokines, or premature exposure to IFN-γ. We believe that the new culture method will enable inquiries into the antimicrobial mechanisms of human macrophages.

Authors

Guillaume Vogt, Carl Nathan

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Figure 4

MDM restriction of M. tuberculosis replication in 10% O2: impact of cytokines in the activation and differentiation phases.

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MDM restriction of M. tuberculosis replication in 10% O2: impact of cyto...
(A and B) Role of cytokines in the activation phase. MDMs from 2 donors were each tested twice in independent experiments, one of which is illustrated. MDMs were differentiated for 14 days in 10% O2, treated (right panels) or not (left panels) with IFN-γ (2.5 ng/ml) alone or with the other cytokines indicated, infected on day 16 with M. tuberculosis (MOI of 0.1), and lysed 2 weeks later for determination of CFU. (C) Comparison of GM-CSF and M-CSF in the differentiation phase. MDMs were differentiated as in A, but with M-CSF or GM-CSF (50 ng/ml each) during differentiation. Numbers above bars indicate estimated percentage destruction of the monolayers as recorded in photomicrographs on the day the CFU were determined. Absence of numbers indicates 100% confluent monolayers. Arrow indicates that the adverse effect of M-CSF during the first 14 days was overcome by inclusion of GM-CSF in the next 16 days. Results are from 1 experiment representative of 5 performed.