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clinical implications of basic research

The Granuloma in Tuberculosis Friend or Foe?

Eric J. Rubin, M.D., Ph.D. Granulomas are cellular aggregates that are the pathologic hallmarks of tuberculosis. These chronic inflammatory lesions have long been considered to be necessary for containment of infection. A recent study by Davis and Rama krishnan,1 however, suggests that granulomas may help to promote infection, rather than sim ply contain it. The vast majority of persons infected with Mycobacterium tuberculosis remain asymptomatic for life; at least 90% of infected adults never be come ill. What is the basis for resistance to tuber culosis? The high rates of clinical tuberculosis among persons infected with the human immu nodeficiency virus and also among persons re ceiving cytotoxic therapy point to a critical role of the intact adaptive immune system. However, the most important cellular player may be the macrophage, which has two mutually contradic tory roles in tuberculosis. On the one hand, acti vated macrophages are capable of killing or at least controlling the growth of M. tuberculosis. Granulomas are present in persons with intact immunity but absent or poorly formed in persons with poor immune responses; this observation supports the hypothesis that they are critical for limiting bacterial growth. On the other hand, macrophages provide the primary growth niche for this intracellular organism; throughout infec tion, mycobacteria are largely intracellular. The hypothesis that granulomas limit bacte rial growth is based largely on animal models that do not permit the observation of infection continuously over time. Granulomas are located deep in tissues; most models require that infect ed animals be killed to permit observation of the interaction between bacteria and host structures. Thus, we have had to draw conclusions about a dynamic process from analyses at single time points. Although the zebrafish cannot be infected with M. tuberculosis, it is a natural host for the fish (and occasionally human) pathogen M. marinum. More importantly, the zebrafish embryo (which is also a host for M. marinum) is trans parent, allowing easy visualization of living bac teria in a living host. Davis and Ramakrishnan infected zebrafish with M. marinum that expressed fluorescent proteins. Phagocytic cells, the fish equivalent of human macrophages, then took up the labeled bacteria, allowing investigators to follow the fate of both infected and uninfected phagocytes over time by means of microscopy (Fig. 1). Infected cells appear to recruit uninfect ed phagocytes. As infected cells die, apparently via apoptosis, they are taken up by previously un infected cells that themselves become infected. These cells provide a new growth niche for the pathogen and permit renewed bacterial growth. The authors observed that M. marinum mu tants with a disabling mutation at the ESX1 lo cus, a chromosomal region critical to the virulence of M. tuberculosis,2 are able to infect phagocytes but do not efficiently recruit uninfected cells for further rounds of infection. Thus, growth of the M. marinum mutant was restricted by a completely unexpected mechanism involving the attraction, by bacterial factors, of uninfected cells in order to create a favorable growth environment for the pathogen. How well does this model translate to human tuberculosis? There are important differences in both the pathogens and the hosts. For example, whereas M. tuberculosis resides largely within vacu oles in the infected cell, M. marinum can escape into the cytoplasm and probably takes advantage of efficient mechanisms of celltocell spread.3 In addition, the embryo of the zebrafish lacks an adaptive immune system, and the human adaptive immune system is critical to contain ing M. tuberculosis infection at later stages. Re modeling of the granuloma in humans with contributions from immune cells and the cyto kines they release might produce an environment that is less hospitable for the pathogen than for the fish granuloma.
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Mycobacterium marinum Bacterial infection

Phagocytic cell

Cell death

Recruitment of phagocytic cells

Egress of infected cells produces new granulomas

Further growth of pathogen

Infected cell ingested

ESX1-deficient Mycobacterium marinum Bacterial infection

Phagocytic cell

Cell death

Less recruitment of phagocytic cells

Egress of fewer infected cells results in fewer and smaller new granulomas

Further growth of pathogen

Infected cell ingested

COLOR FIGURE

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infected macrophages The New England Journal of Medicine ME Downloaded from nejm.org on December 8, 2012. For personal use only. No other uses without permission. DE Phimister Copyright 2009 Massachusetts Medical Society. All rights reserved. Artist Knoper

Bacterial growth in

clinical implications of basic research

Figure 1 (facing page). The Mycobacterium and the Macrophage. Davis and Ramakrishnan1 recently observed that growth of Mycobacterium marinum in the phagocytic cell of zebra fish (the zebrafish equivalent of the human macrophage) eventually leads to cell death (Panel A). Infected cells re cruit uninfected cells, some of which ingest dead infected cells; this provides a niche for further growth of the patho gen and permits egress of infected cells to produce new granulomas. Infection with bacteria that do not have the critical virulence region ESX1 (Panel B) results in less recruitment of uninfected cells and, consequently, fewer bacteria, smaller lesions, and fewer new granulomas.

mechanisms to eradicate early infection. The study by Davis and Ramakrishnan suggests that interfering with signaling between the host and pathogen might tip the scales in favor of clear ance of infection.
No potential conflict of interest relevant to this article was re ported. From the Harvard School of Public Health, Harvard University, Boston. pansion and dissemination of early tuberculous infection. Cell 2009;136:3749. 2. Pym AS, Brodin P, Brosch R, Huerre M, Cole ST. Loss of RD1 contributed to the attenuation of the live tuberculosis vaccines Mycobacterium bovis BCG and Mycobacterium microti. Mol Microbiol 2002;46:70917. 3. Stamm LM, Morisaki JH, Gao LY, et al. Mycobacterium mari num escapes from phagosomes and is propelled by actinbased motility. J Exp Med 2003;198:13618. 4. Ewer K, Millington KA, Deeks JJ, Alvarez L, Bryant G, Lalvani A. Dynamic antigenspecific Tcell responses after pointsource exposure to Mycobacterium tuberculosis. Am J Respir Crit Care Med 2006;174:8319.
Copyright 2009 Massachusetts Medical Society.

1. Davis JM, Ramakrishnan L. The role of the granuloma in ex

Sand castles topple, however. For example, the longheld view that an initial encounter between M. tuberculosis and lung macrophages inevitably leads to chronic infection has been challenged. Newer and more specific tests indicate that the contacts of tuberculosis patients often mount an immune response that subsequently resolves,4 a finding that suggests that humans have innate

collections of articles on the journals web site

The Journals Web site (NEJM.org) sorts published articles into more than 50 distinct clinical collections, which can be used as convenient entry points to clinical content. In each collection, articles are cited in reverse chronologic order, with the most recent first.

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