Viewpoint pubs.acs.org/journal/aidcbc
Cite This: ACS Infect. Dis. XXXX, XXX, XXX−XXX
Lymph Nodes: The Unrecognized Barrier against Pathogens Ania Bogoslowski and Paul Kubes* Calvin, Phoebe & Joan Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta T2N 4N1, Canada Department of Physiology and Pharmacology, Cumming School of Medicine, University of Calgary, Calgary, Alberta T2N 4N1, Canada ABSTRACT: Lymph nodes have been studied for decades as the main site of the adaptive immune response. In this Viewpoint, we outline how the lymph nodes have another less appreciated function as an active innate barrier. Lymph nodes drain lymphatic fluid from tissues that are exposed to the external environment, such as the skin, lung, or gut. Pathogens that travel through lymphatics should be able to enter the circulation, if it were not for the strategic localization of lymph nodes along lymphatics which prevent systemic access. There is growing evidence for several populations of innate immune cells in the lymph node that function to control pathogens. Understanding how the lymph node functions as an active innate barrier can contribute to improving defenses against dissemination of infections in patients.
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Table 1. Pathogens That Disseminate via the Lymphatic System
ymph nodes are strategically located throughout the lymphatic system at key draining points. Their function as collection sites for antigen from the periphery and adaptive immune responses is well established. However, in the past several years, research into a new function of the lymph nodes has been emerging. The presence of localized reservoirs of innate immune cells in the lymph node suggests that this organ may be more than just a hub of adaptive immunity. While adaptive immune cells need specific antigen presentation, costimulation, and then time (days) for clonal expansion, the innate immune system will respond to pathogens instantly and contain pathogens until adaptive immunity is activated. Indeed, it is becoming appreciated that lymph nodes may play an important role as active barriers, preventing dissemination of pathogens via lymphatics.
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pathogen nematode worm (Filariodidea family) Staphylococcus aureus Mycobacterium tuberculosis Streptococcus group A Yersinia pestis Salmonella species Listeria monocytogenes
initial infection site
method of dissemination to lymph node
skin
migration of larva2
skin lung
free (not within immune cells)3 unknown4
skin
free, hyaluronan and Lyve-1 interactions5
skin gut gut
in phagocytes6 in monocytes and granulocytes7 free, expression of In1A (internalin A, a high affinity ligand for E-cadherin)8
A BARRIER TO PATHOGENS
Pathogen dissemination through lymphatics is surprisingly understudied despite it being a common occurrence. For example, lymphatic filariasis is a lymphatic and lymph node trophic disease caused by three species of nematode worms that infect 120 million people globally.1 Many pathogens may spread directly to lymph nodes, carried by immune cells, or by some unknown mechanism (Table 1). The first level of innate immune defense against any pathogen is at the skin and mucus barriers. Once these are breached, innate immune cells are quickly recruited to keep the pathogen contained.9 Because pathogens arrive unexpectedly into tissue, they have the advantage of being there prior to recruited immune cells. As such, there is a possibility that the pathogen could manage to escape into lymphatics or blood. Liver Kupffer cells and spleen macrophages are positioned in blood vessels to capture pathogens, preventing systemic dissemination.10,11 In the case of bacterial dissemination through lymphatics (which are devoid of resident macrophages), one could imagine that this would be a free passage back into the bloodstream. However, the pathogens need to © XXXX American Chemical Society
bypass the lymph nodes, which as we will show in this Viewpoint, function as the next sequential barrier. The gut is an important entry site into the body for pathogens. The gastrointestinal tract is home to a plethora of symbiotic bacteria living in a carefully maintained balance. In the case of translocation of these otherwise beneficial bacteria or pathogens during gastrointestinal infection, the mesenteric lymph nodes serve as essential barriers. Interestingly, a recent study suggests that bacteria that cross the intestinal barrier flow to the lymph nodes and are not able to cross into the bloodstream, either directly or via the lymph nodes.12 Disruption of gut homeostasis with alcohol or medication such as NSAIDS can either alter the microbiome or injure the mucosal barrier, enhancing bacterial translocation.13,14 On the basis of these potential breaches of the gastrointestinal tract by bacteria, it would be logical for the lymph nodes to have innate immune cells at the ready for the onslaught of any Received: May 1, 2018
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DOI: 10.1021/acsinfecdis.8b00111 ACS Infect. Dis. XXXX, XXX, XXX−XXX
ACS Infectious Diseases
Viewpoint
and infected the subcapsular macrophages but spread no further because it was contained by essential IFN-γ responses from the macrophages.17 When these macrophages were removed, the virus infected the neurons in the lymph node and spread to the central nervous system.17 It is evident that the resident macrophages are a key part of the innate-immune barrier function of the lymph node. Another key innate immune cell, the natural killer (NK) cell, resides in the interfollicular zone of lymph nodes.16 In response to certain stimuli, such as Ankara virus, NK cells will change their usual migratory behavior and begin to accumulate in the subcapsular sinus while upregulating activation markers.18 This change in behavior is partially mediated by subcapsular sinus macrophages that secrete type I IFNs to promote antiviral responses in the NK cells.18 Another study showed that NK cells can kill cytomegalovirus-infected lymph node stromal cells to prevent dissemination of murine cytomegalovirus from the lymph node.19 NK cells also carry out antibacterial functions in the lymph node, as demonstrated by their ability to secrete TNF-α, a pro-inflammatory cytokine, to direct the immune response against Yersinia pestis.20 Recently, it has been observed that resident innate-like IL17+ CCR6+ T lymphocytes patrol the lymph node subcapsular sinus and interact with resident macrophages.21 The presence of bacteria activates these innate-like lymphocytes to release IL17, a cytokine with a key antibacterial and antifungal role in the skin.21 The presence of these innate cells and their rapid responses to infections emphasizes the function of innate immune cells in the lymph node as a barrier to disseminating infection. Several papers have identified that neutrophils can be recruited to the draining lymph node following infection with mycobacterium,22 Toxoplasma gondii,23 Pseudomonas aeruginosa,16 and Staphylococcus aureus.24,25 In these studies, huge numbers of neutrophils were recruited to the lymph node very quickly following infection in peripheral tissue. Neutrophils in the lymph node exhibited swarming behavior and clearance of possibly infected macrophages.16,23 Two of these studies identified neutrophils traveling through afferent lymphatic vessels, following pathogens to the lymph node.22,24 In order to understand the mechanisms of neutrophil entry and the role of these cells in acute infection in the context of the lymph node, our group has recently studied neutrophil recruitment following a peripheral S. aureus infection.3 We showed that after S. aureus infection, bacteria are detectable only in the site of infection and the draining lymph node, with no bacterial dissemination to the blood and other organs.3 As soon as 30 min after infection, large amounts of neutrophils begin entering the lymph node via blood vessels in an L-selectin and platelet P-selectin dependent pathway and swarming within the lymph node tissue.3 Finally, if neutrophil recruitment into lymph nodes is blocked, then S. aureus will disseminate, which is greatly exacerbated in the absence of macrophages.3 This demonstrates the key role that neutrophils play in the barrier function of the lymph node. It also emphasizes that this organ is not just a passive barrier but that it can be actively modulated to quickly recruit additional cells in response to an insult. Figure 1 illustrates the rapid changes the innate immune cells of the lymph node undergo to boost defenses in infection.
disseminating bacteria, as well as the ability to quickly activate or recruit additional cells.
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INNATE IMMUNE CELLS IN THE LYMPH NODE Several innate immune cell subsets have been identified in specific areas of the lymph node (Figure 1). At steady state, one
Figure 1. Innate immune cell populations in the lymph node at steady state and after infection. Macrophages reside in the subcapsular sinus and the medullary sinus. Lymphatic fluid brings pathogens, DAMPS, and antigen to macrophages. At steady state, NK cells and innate-like T cells reside in the T cell zone or the interfollicular zone (near, but not in, the B cell zone). There are also neutrophils present in the lymph node at steady state. Following an infectious stimulus, the subcapsular sinus macrophages can activate NK cells and innate-like T cells. Innate-like T cells accumulate in the subcapsular sinus. Rapidly after infection, neutrophils enter the lymph node through the HEV and accumulate in the tissue and subcapsular sinus.
population of resident macrophages lines the subcapsular sinus and another is resident within the medullary sinus. The macrophages of the lymph node have been dubbed “cellular flypaper” because of their ability to bind and/or phagocytose materials from the lymphatic fluid. Lymph borne materials reach the lymph node only a few minutes after subcutaneous injection.15 While small particles will enter the lymph node paracortex using a series of conduits, bigger particles, bacteria, and viruses will be captured by lymph node macrophages. Eliminating these macrophages increases pathogen spread.16,17 Kastenmüller et al. showed IL1-β released from lymph node resident macrophages activated innate-like lymphocytes and contributed to neutrophil recruitment following footpad infection with Pseudomonas aeruginosa.16 Depletion of these resident macrophages increased the bacterial load in the lymph node and the blood.16 Subcapsular sinus macrophages have been shown to be particularly important in viral infection. When a neurotrophic virus, vescicular stomatisis virus, was injected in the footpad, it reached the lymph node
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DESIGNED FOR DEFENSE The location of lymph nodes also supports the hypothesis that these organs play a barrier role. Lymph nodes are organized in B
DOI: 10.1021/acsinfecdis.8b00111 ACS Infect. Dis. XXXX, XXX, XXX−XXX
ACS Infectious Diseases
Viewpoint
series like a pearl necklace, so that if the first lymph node is overwhelmed, redundancy exists in the system. In the case of an overwhelming dissemination from a skin infection, we demonstrated that, even if the defenses in the first draining lymph node were saturated, any pathogens that escaped were detained in the subsequent lymph node.3 Often, it is overlooked that the lymphatic fluid brings much more than just antigen and antigen-presenting cells to the lymph node. The lymphatic fluid is a concentrate of soluble information describing the state of the tissue being drained. From the interstitial fluid of a tissue, there is a mixture of growth factors, cytokines, and serum proteins that make their way to the lymph node. The lymphatic vessels are an information highway bringing a “status report” on specific tissues to the lymph node. To sift through all this information, the complexity of the innate immune cells residing in the lymph node may be much greater than previously appreciated. The types of studies discussed in this Viewpoint help in developing a new paradigm for understanding the lymph node’s innate immune functions. In conclusion, initial studies have shown many different populations of innate immune cells in the lymph node along with an enormous capacity for additional recruitment. When the lymph node is notified of a potential danger in the upstream tissue, the innate immune cells are poised to defend the site against pathogens that disseminate via lymphatics. Knowledge of how innate immune cells behave in the lymph node could be used to boost these natural responses and prevent bacterial dissemination in at-risk patients.
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AUTHOR INFORMATION
Corresponding Author
*Tel: (403) 220-2705. Fax: (403) 270-7516. E-mail: pkubes@ ucalgary.ca. ORCID
Ania Bogoslowski: 0000-0002-1767-2298 Notes
The authors declare no competing financial interest.
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DOI: 10.1021/acsinfecdis.8b00111 ACS Infect. Dis. XXXX, XXX, XXX−XXX
ACS Infectious Diseases
Viewpoint
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DOI: 10.1021/acsinfecdis.8b00111 ACS Infect. Dis. XXXX, XXX, XXX−XXX
