One of the main features of the immune response to
Almost 20 people develop tuberculosis and four people die from the disease every minute, somewhere in the world [
The prognosis of the disease depends on the ability of the host to eliminate the bacillus. The respiratory route is the principal route of infection. The disease starts when droplets from actively or latently infected individuals reach the respiratory tract of healthy individuals. These droplets contain a small number of bacilli that enter the lung, where they infect primarily alveolar macrophages, type 2 pneumocytes, and polymorphonuclear neutrophils (PMNs) [
Many studies have suggested mechanisms for the initial events in the lung, but most were based on experiments carried out
We know that the infected alveolar macrophages in the lung release various cytokines to recruit different populations of cells, including more macrophages, to the infection site. Dendritic cells are important because they present antigens to T cells in the lymph nodes, in which a T-cell response can subsequently be developed. These signalling events lead to the formation of a granuloma, the hallmark of tuberculosis. This structure is developed by the host to contain the infection and eliminate the bacteria. However, the bacteria persist in a latent state within the granuloma, often for decades. They subsequently reactivate in 10% of the latently infected individuals. The death of the infected cells causes the formation of a necrotic zone in the centre of the granuloma, which eventually disintegrates, releasing the bacteria into the lung, and thence into the environment (see Figure
Formation and maturation of lung tuberculous granulomas. Following inhalation of contaminated aerosols,
The granuloma, the hallmark of tuberculous disease, creates an immune microenvironment in which the infection can be controlled. However, it also provides the mycobacterium with a niche in which it can survive, modulating the immune response to ensure its survival without damage over long periods of time [
The granuloma contains mostly blood-derived macrophages, epithelioid cells (differentiated macrophages) and multinucleated giant cells (also known as Langhans giant cells), surrounded by T lymphocytes [
Some lymphoid clusters organised similarly to the follicular centres of lymph nodes are also associated with granulomas (see Section
Many different chemokines are involved in granuloma formation (see Table
CCL19 and, possibly, CCL21 are involved in the recruitment and priming of IFN-
Animal models are often used to study granulomatous structures. These models re-produce many of the processes occurring in humans, although differences are frequently observed. It is difficult to study biopsy samples from the lung, to which access is often limited. This has led to the widespread use of animal models, which have been improved over the years to reproduce more closely the progression of the disease observed in humans. A large number of mouse models of infection have been generated, but the most relevant is probably that based on the intranasal infection route, because this is the route involved in natural infections in humans [
However, murine granuloma models that more closely resemble granulomas in humans have recently been described. Nos 2−/− mice infected with
Rabbits and guinea pigs, which form granulomas largely similar to those in humans, have also been studied [
Computer models have also recently been developed, to describe or to make predictions based on general information about tuberculosis disease [
Zucchi et al. established a model of tuberculosis in the central nervous system (CNS), in which the injection of mycobacteria into the cerebellum induces granuloma formation. This model is useful for studies of Tb meningitis and of other types of extrapulmonary tuberculosis. The principal drawback of this model is the infection route, which differs radically from the natural route of infection. However, as a model of granuloma formation, it is a useful tool for studying the physiopathology of brain infection and pathogen/host dynamics [
Models for studying Mtb infection and the granulomatous response.
Model | Advantages | Drawbacks |
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Monkey | Granuloma similar to humans. | Difficult to handle. |
Expensive. | ||
| ||
Guinea pigs/rabbits | Granuloma similar to humans. | Restricted availability of reagents. |
Easy to handle. | Genetic manipulation difficult. | |
| ||
Mice | Easy to handle. | Granulomas often differ in many ways from |
Model of choice for genetic studies. | human granulomas (e.g., cellular composition and progression to necrosis). | |
| ||
Zebrafish embryo | Easy to handle. |
|
Good for real-time experiments and live imaging (the larvae are transparent). | ||
Good for studies of the initial steps of granuloma formation and the role of innate immunity. | No lymphocytes present in the embryo. | |
| ||
|
Mimics the physiological granuloma. | Some important elements present in the lung compartment but not in PBMCs may be lacking. |
Possible to study the early steps of granuloma formation. | ||
Flexible (e.g., use of various strains of bacteria, easy addition of cells, cytokines, drugs). | ||
| ||
|
Not expensive. | Highly dependent on the initial parameter settings and cannot take previously unknown information into account. |
Study of the early steps of granuloma formation possible. | ||
Flexible. |
Main chemokines and cytokines involved in the granulomatous response.
Chemokines/cytokines | Main producers | Targets/role |
---|---|---|
CXCL8 (IL-8) | Alveolar macrophages. | Recruitment of neutrophils. |
Epithelial cells of the lung. | ||
| ||
CCL2 (MCP-1) | Monocytes and alveolar macrophages. | Recruitment of macrophages and other immune cells. |
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CCL3 (MIP-1a), CCL4 (MIP-1b) CCL5 (RANTES) | Alveolar macrophages. | Recruitment of macrophages and other immune cells. |
| ||
CXCL9, CXCL10 (IP-10), CXCL11 | Bronchial epithelial cells. | Recruitment of immune cells. |
| ||
CCL19/CCL21 | Stromal cells of the lymph nodes. | Recruitment and priming of IFN- |
Migration of DC from the lung to draining lymph nodes. | ||
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CXCL13 | Dendritic cells, stromal cells of the lymph nodes. | Recruitment of B cells and formation of the granuloma-associated follicular structures. |
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IL-12/IL-23 | Dendritic cells, macrophages. | Th1 polarisation of CD4+ T cells. |
| ||
IFN- |
CD4+ (Th1) and CD8+ T cells, NK. | Activation of macrophages. |
Induction of NO synthesis and bacterial killing. | ||
| ||
TNF- |
CD4+ T cells (Th1), macrophages. | Proinflammatory. |
Induction of chemokine production. | ||
Activation of macrophages. | ||
Critical for granuloma formation. | ||
| ||
IL-1 | Macrophages, DCs. | Proinflammatory. |
Recruitment and activation of phagocytes. | ||
| ||
IL-17 | LT |
Proinflammatory. |
Involved in neutrophil recruitment and macrophage activation. | ||
| ||
IL-10 | Tregs, B-1 cells, AAM. | Anti-inflammatory. |
Polarisation of macrophages towards the AAM type. | ||
| ||
TGF- |
Tregs, AAM. | Anti-inflammatory. |
Most of the macrophages involved in granuloma formation are epithelioid macrophages. These cells are activated and have an abundant cytoplasm. After initial infection, the macrophages (monocytes when immature) migrate to the infection site from the blood. They have various innate immune receptors in their membranes, allowing them to recognise, bacteria, take them up by phagocytosis, and secrete various cytokines. These receptors belong to four main classes: opsonizing receptors (e.g., Fc
As pointed out above, macrophages are the principal cells found in granulomas, but not all the macrophages in the granuloma are infected. The uninfected cells seem to help to contain infection and contribute to cytokine secretion. Some studies have suggested that there are two kinds of macrophages: classically activated macrophages (CAM), which differentiate in response to cytokine signals, or alternatively activated macrophages (AAM). CAMs, which are induced by the secretion of Th1 cytokines, are able to kill bacteria. In murine models, these cells produce iNOS. This enzyme catalyses the synthesis of nitric oxide (NO), a potent antimicrobial compound. The production of iNOs is strongly induced by IFN-
Recent data from mouse models suggest that macrophages of the AAM type are also found in the tuberculous granuloma. The TLR signalling triggered by mycobacteria may lead to the induction of arginase production by macrophages, through the MyD88-dependent production of IL-10, IL-6, and granulocyte colony-stimulating factor (G-CSF). Switching off arginase expression has been shown to be beneficial for host survival [
Macrophages may also fuse to generate multinucleated Langhans giant cells (MGCs), which are characteristic of tuberculosis. The ontogeny of these cells has only recently been described [
mDCs have been found in the granulomas of tuberculosis patients [
Foamy macrophages are also classically found in human tuberculous granulomas. Their foamy appearance results from the accumulation of intracellular lipids within lipid bodies (LB) or droplets. The differentiation of macrophages into foamy cells can be triggered
Foamy cells are found within granulomatous structures in both animal and human models. We have shown that foamy macrophages have lost their phagocytic and bactericidal activities and that they allow Mtb persistence in a dormant state [
Neutrophils are also involved in the granulomatous response. These cells have been described as the first line of defence against tuberculosis. They are activated directly by Mtb products, such as lipoarabinomannan (LAM) in particular [
T lymphocytes account for 15 to 50% of the leukocytes in mouse granulomas. About 60–70% of the T cells present are CD4+, 15–30% are CD8+
The essential role of CD4+ T cells in the control of mycobacterial infection has been highlighted by many studies in knockout mice [
Ordway et al. depleted mice of CD4+ T cells at different stages of infection by Mtb and showed that this caused disorganisation of the granulomatous lesion at all stages [
APC stimulates CD4+ T cells via TCR engagement, together with CD40-CD40L interaction and the production of IL-12. This leads to Th1 polarisation and the strong production of IFN-
A study of CD40L−/− mice showed that engagement of the costimulatory receptor was also required for the efficient recruitment of CD4+ T cells to granulomatous lesions. By contrast, CD28-mediated costimulation is not required, despite the smaller numbers of splenic CD4+ T cells and the lower level of activation of these cells in CD28−/− mice. Some granulomas are formed in CD40L−/− mice, but they fail to control bacterial growth effectively, resulting in a phenotype similar to that of IFN-
A genetic syndrome called Mendelian susceptibility to mycobacterial infection has been described in humans. It is characterized by disseminated infections after vaccination with BCG or contact with nonvirulent mycobacteria (e.g.,
Other Th1-polarising cytokines, such as IL-27 (a member of the IL-12 family), have been described. As expected, the CD4+ T cells of mice with impaired IL-27 signalling produce lower levels of IFN-
CD4+ T cells have been shown to have cytotoxic activity against
Gallegos et al. carried out a series of adoptive transfers of T cells with different genetic alterations in mice of various genetic backgrounds and found that IFN-
In addition to the classical Th1 response, a Th17-type response is also triggered by mycobacterial infection. IL-17 is a proinflammatory cytokine driving the recruitment of effector cells, such as neutrophils, and participating in the activation of macrophages. Some IL-17-producing CD4+ T-cells are present in mycobacterial granulomas, but
The CD4+CD25+FoxP3+ regulatory T-cell compartment is expanded both in patients with active TB [
Mice lacking CD8+ T cells (ß2m−/− MHC-I-deficient mice [
In mouse lung granuloma, CD8+ T cells are initially found towards the periphery, migrating deeper into the structure as the disease progresses [
By immune reconstitution of athymic mice with IFN-
Ordway et al. also suggested another role for CD8+ T cells in the granuloma. They found that, during chronic infection, activated CD8+ T cells produced the chemokine XCL1 (lymphotactin), which negatively regulates IFN-
These cells were first described in granulomas more than 20 years ago and seem to be involved in the formation and development of these structures [
Saunders et al. reported different results for
In mice,
NKT cells, which express both an
However, granuloma formation in the lungs of NKT KO mice infected intranasally with Mtb seemed to be as efficient as that in wild-type mice [
Further studies are required to determine the exact ligands of NKT cells and the effect of these cells on the immune response in the context of mycobacterial infection.
The response to mycobacterial infection is based mostly on cellular immunity, with the role of humoral immunity in protection against TB remains a matter of debate [
The lungs of mice with no B cells contain fewer granulomas than those of wild-type mice, and these granulomas are much smaller with little cellular infiltrate. However, they display higher levels of neutrophil and CD8+ T-cell recruitment. These changes are not correlated with differences in the ability to contain the infection, as the lungs of wild-type mice and mice with no B cells contain similar numbers of bacteria [
Cells of the B-1 subset are present in the peritoneal and pleural cavities. Their function remains unclear, but they may serve as a first line of defence, as described for
The granuloma tends to become necrotic in susceptible individuals, facilitating bacterial spread by coughing. The necrotic tissue has a characteristic “caseating” appearance [
There are good reasons to think that neutrophils also play a role in necrosis. It remains unclear whether these cells are protective or damaging, but, when Mtb-infected animals are repeatedly challenged with mycobacterial antigens, the lesions become necrotic and contain a higher proportion of granulocytes [
Early diagnosis and appropriate treatment of TB are important, to reduce transmission and favour the elimination of the bacterium. Tuberculosis is diagnosed on the basis of laboratory tests, chest X-rays, CT/MRI scans, microbiologic smears, and cultures of body fluids, including sputum in some cases, histological analysis, and biopsy. Diagnosis and follow up have recently been improved by the development of imaging techniques using radiopharmaceutical compounds. FDG-PET can be used not only for diagnosis, but also for monitoring throughout tuberculosis treatments, particularly in cases of multidrug-resistant infections. Soussan et al. identified two different patterns of pulmonary TB by FDG-PET/CT [
We now have various experimental models that should help to unveil the mysteries of the complex host-pathogen relationships taking place in the mycobacterial granuloma. Granuloma formation seems to be primarily a host-defence mechanism for containing the bacteria, but it also shelters the bacteria, providing them with a niche in which they can persist in a latent form until an opportunity arises for re-activation and spread. An understanding of the physiopathology of granulomas is critical for the design of new vaccines and antituberculous drugs.
Granulomas are not restricted to mycobacterial infections, being found in many different kinds of bacterial, fungal or viral infections, and even in noninfectious inflammatory diseases [
The authors would like to thank the Institut National de la Santé et de la Recherche Médicale (INSERM), the Agence Nationale pour la Recherche (ANR), and European Framework Program 7 (FP7) for funding their work. We also thank Jérémy Segard for his artistic representation of granuloma formation and maturation.