Q J Med 2014; 107:789–792 doi:10.1093/qjmed/hcu005 Advance Access Publication 18 January 2014 Review A novel regulator of lung inflammation and immunity: pulmonary parasympathetic inflammatory reflex X. YANG, C. ZHAO, Z. GAO and X. SU From the Unit of Respiratory Infection and Immunity, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai 200031, China Address correspondence to X. Su, MD, PhD, Unit of Respiratory Infection and Immunity, Institut Pasteur of Shanghai, Chinese Academy of Sciences, B104, Life Science Research Building, 320 Yueyang Road, Shanghai 200031, China. email: [email protected] Summary information integrating center. This modulatory circuit might loop the lungs, immune and nervous systems and play a very important role in regulating lung infection, inflammation and immunity through the neural innervations and signals when the lungs encounter pathogenic challenges. Introduction macrophages express a7 nAChR that are commonly expressed by neurons.2 Moreover, electric stimulation of vagus nerve can trigger release of acetylcholine (ACh), which can activate a7 nAChR in the macrophages and therefore suppress activation of NF-kB and dampen proinflammatory cytokine production.3 Studies further revealed that the spleen plays an important role in mediating anti-inflammatory activity of vagus nerve.4,5 More importantly, Tracey group set forth a new working model for CAP regarding the role of spleen.6 The main conception is that vagal outflow at splenic nerve terminus releases adrenergic agonist-norepinephrine that could activate b2 adrenergic receptors on splenic CD4+CD44highCD62Llow T lymphocytes that contain choline acetyltransferase (CHAT). These unique T lymphocytes synthesize ACh that could act on a7 nAChR-expressing splenic macrophages and suppress NF-kB activity.6 As such, this theory can be used to interpret the modulatory effects of CAP on proinflammatory responses at Since the cholinergic anti-inflammatory pathway (CAP, also called inflammatory reflex) was termed by Kevin Tracey a decade ago,1 much work has been done regarding how the parasympathetic, especially vagus nerve-mediated inflammatory reflex regulates innate immune responses to injury, pathogens and tissue ischemia. The neuroimmune interplay provides the host with a fast, discrete and localized means of controlling the immune response.1 This review will briefly introduce the basic formation of pulmonary parasympathetic inflammatory reflex and its regulatory function on lung infection, inflammation and immunity. The current status of cholinergic anti-inflammatory pathway The remarkable discovery regarding inflammatory reflex is that proinflammatory cells, especially ! The Author 2014. Published by Oxford University Press on behalf of the Association of Physicians. All rights reserved. For Permissions, please email: [email protected] Downloaded from by guest on February 5, 2015 In this review, we first analyzed the current status of cholinergic anti-inflammatory pathway and then put forward a novel regulatory machinery—pulmonary parasympathetic inflammatory reflex, which is composed by lung vagal sensors at afferent arm, a7 nAChR (a7 nicotinic acetylcholine receptors)— expressing cells at efferent arm and the brain 790 X. Yang et al. systemic levels. We reported that dysfunction of cholinergic signaling in the splenocytes might be associated with postoperative cognitive decline under metabolic syndrome.7 However, this theory is challenged by a recent study that vagal efferent neurons in a rat model neither synapse with splenic sympathetic neurons nor drive their ongoing activity.8 Thus, it is imperative to rethink the current CAP working model,9 especially regulatory effects of pulmonary parasympathetic (vagal) inflammatory reflex during lung infection, inflammation and immunity. Possibility of existence of pulmonary parasympathetic inflammatory reflex The afferent arm of pulmonary parasympathetic inflammatory reflex The vagal nerve endings innervate the distal airways or alveolar epithelia11,12 and express variety of receptors that can sense mechanical, chemical, biological and other stimuli and convey the signaling through the afferent vagal nerve to the center of The efferent arm of pulmonary parasympathetic inflammatory reflex Chemical, mechanical or biological stimuli can directly cause damage of lung alveolar epithelial cells. Proinflammatory cytokines that are secreted by alveolar macrophages and infiltrated proinflammatory cells (e.g. monocytes and neutrophils) also contribute to proinflammatory cascades and further lung injury. Alveolar macrophages, epithelial cells and inflammatory infiltrated neutrophils express a7 nAChR and could be the players at efferent arm of pulmonary parasympathetic inflammatory reflex.13,14 Pulmonary vagal nerve endings innervate lungs and secrete ACh that could activate a7 nAChR-expressing proinflammatory cells at the efferent arm of lung vagal inflammatory reflex, suppress NF-kB activation and secretion of proinflammatory cytokines therefore lessen the extent of lung inflammation and injury. This notion has been strongly supported by the studies that in acid, LPS and Escherichia Coli-induced acute lung injury mouse models (by sensing chemical and biological stimuli), activation of a7 nAChR by its agonists could attenuate, on the contrary, vagotomy and deficiency of a7 nAChR deteriorate the degree of lung injury.13,14 In a ventilator-induced lung injury mouse model (by sensing mechanical stretch), vagotomy worsens proinflammatory responses in the lung.23 Downloaded from by guest on February 5, 2015 The vagus nerve is the 10th cranial nerve and contains sensory (afferent) and motor (efferent) fibers.10 Both right and left vagus nerves descend from the brain in the carotid sheath, lateral to the carotid artery and then innervate the lungs. Electrical microscopy has shown that vagus nerve fibers were present in the human lung, especially in the alveoli.11,12 Interestingly, inflammatory responses of the lungs vary when the pathogenic challenges were stimulated systemically or locally. In an endotoxemia mouse model (systemic challenge), activation of a7 nAChR by vagus nerve stimulation reduced the levels of TNF-a in the plasma, liver and spleen, but not in the lung.4 However, in an acute lung injury model (local challenge), expression of a7 nAChR in alveolar macrophages and neutrophils was increased. Administration of a7 nAChR agonists could inhibit NF-kB activity in the BAL (bronchoalveolar lavage) proinflammatory cells and reduce both TNF-a and MIP-2 levels in the airspaces of the lung. Vagotomy and deficiency of a7 nAChR reversed the above mentioned pathological processes.13,14 These findings indicate a vagal inflammatory circuit may exist between the lung and nervous system and exert modulatory effects on lung infection and immunity. brain (e.g. NTS, nucleus tractus solitorius).15 For example, vagal C-fiber receptors can sense mechanical and chemical irritant stimuli.16 These vagal nociceptors can also be activated by proinflammatory cytokines and therefore transmit immune signals from the lung to the brain.17 Vagal afferent nerve endings express ion channel-transient potential channels,18 which can detect the pungent odor, hot sensations and ion changes. Toll-like receptors 2, 3, 4 and 7 are expressed in the nerve endings or airway epithelial cells,19 that can respond to bacterial and viral pathogens. In addition, pulmonary sensory neurons and epithelial cells are reported to express inflammatory cytokine receptors (e.g. TNF-a receptor)20,21 which may be the components of vagal afferent arm of pulmonary vagal inflammatory reflex. Furthermore, bacteria and their products could activate sensory neurons and modulate inflammation that supports the role of nervous system in host–pathogen interactions.22 Pulmonary parasympathetic inflammatory reflex Brain center of pulmonary parasympathetic inflammatory reflex It is assumed that vagal sensors recognize local proinflammatory cytokines or other mediators and then transmit the information to brain center. The information is further processed in the brain, and reflexively acts through the vagal efferent to suppress proinflammatory cytokine production of inflammatory cells through release of ACh and activation of a7 nAChR.24 Anatomically, vagus afferent fibers reside in the nodose ganglion and terminate primarily within the dorsal vagal complex (DVC) of the medulla oblongata. The DVC consists of the NTS, the dorsal motor nucleus of the vagus (DMN) and the area postrema.25 Stimulation of capsaicin-sensitive vagal afferents could alter c-fos expression in the NTS in a mouse model if inoculated with bacteria,26 suggesting that NTS might be the center at brain that could integrate the information from the vagal sensory fibers, but this hypothesis requires further study. Airway epithelia and periepithelial tissue are vagally innervated.11,12 Classically, vagal nerve endings can directly release ACh upon inflammatory stimulation. During lung viral infection and inflammation, CHAT mRNA is upregulated in the lung that could promote alternative synthesis of ACh in an a7 nAChR-dependent manner (Su X, et al. unpublished data). These results indicate that vagal nerve endingsreleased ACh might trigger synthesis and secretion of ACh by the lung cells (possibly epithelial cells or neuroendocrine cells). This positive feedback between ACh and its receptor maintains a constant generation of ACh in the alveoli, which can inhibit NF-kB activity in the a7 nAChR-expressing alveolar macrophages and infiltrated proinflammatory cells and therefore dampen lung inflammation and injury.27 Due to the rapid hydrolysis of ACh (half-life, 2 min), measurement of the concentration of ACh in the BAL requires prevention of ACh hydrolysis and advanced techniques (e.g. high performance liquid chromatography). Indirect way to evaluate ACh is through measuring AChE activity and choline (a metabolite of ACh) levels. Study has shown that AChE activity in proinflammatory cells and BAL choline concentration were markedly increased in the E. coli pneumonia mice compared with normal mice.13 Taken together, vagus nerve, lung epithelial cells and BAL proinflammatory cells may collaboratively participate in the synthesis and hydrolysis of ACh during lung infection and inflammation. Responses of pulmonary parasympathetic inflammatory reflex to different pathogens The regulatory effects of pulmonary parasympathetic inflammatory reflex on lung infection and immunity are pattern recognition receptors dependent. Gram-positive bacterial infection can be recognized by TLR2. Activation of a7 nAChR by nicotine increased Streptococcus Pneumoniae-induced lung infection and lung inflammation.28 TLR4 is the receptor of endotoxin, a component of Gram-negative bacteria. Activation of a7 nAChR by nicotine reduces mortality of E. coli pneumonia; on the contrary, cervical vagotomy and a7 nAChR deficiency increases mortality of E. coli pneumonia.14 In addition, TLR3, TLR7 and MAVS-RIG-I systems play key roles in mediating influenza viral lung infection and inflammation.29 In vitro experiments have shown that in the lung epithelial cells, activation of a7 nAChR promotes influenza viral replication. Deficiency of a7 nAChR protects mice from influenza virus-induced lung infection and inflammation (Su X, et al. unpublished data). Thus, pulmonary parasympathetic (vagal) inflammatory reflex is functionally versatile in response to the challenges of different pathogens. Perspectives and conclusion The interaction between nervous and immune systems has been studied for many years, but there are many key scientific questions we cannot answer. Particularly, we need to understand how vagus nerve terminals sense the invading pathogens, how the infection signals transmit to the brain center and how the brain responds to infection and regulates the immune cells to eliminate pathogens. There is more work to do; nevertheless, this study supports the view that pulmonary parasympathetic inflammatory reflex might be present and function as a requisite regulator for lung infection, inflammation and immunity. Funding The National Natural Science Foundation of China (Grant No.81270139 to X.S.); The Key Project of Science and Technology of Shanghai (Grant No. 12JC1408900 to X.S.); One Hundred Person Downloaded from by guest on February 5, 2015 Sources and metabolism of ACh in the lung 791 792 X. Yang et al. Project of the Chinese Academy of Sciences (Grant No. Y316P21209 to X.S.); The Knowledge Innovation Program of the Chinese Academy of Sciences (Grant No. Y114P11209 to X.S.). Conflict of interest: None declared. References 1. Tracey KJ. The inflammatory reflex. Nature 2002; 420:853–9. 2. Wang H, Yu M, Ochani M, Amella CA, Tanovic M, Susarla S, et al. Nicotinic acetylcholine receptor alpha7 subunit is an essential regulator of inflammation. Nature 2003; 421:384–8. 3. Borovikova LV, Ivanova S, Zhang M, Yang H, Botchkina GI, Watkins LR, et al. Vagus nerve stimulation attenuates the systemic inflammatory response to endotoxin. Nature 2000; 405:458–62. 4. Huston JM, Ochani M, Rosas-Ballina M, Liao H, Ochani K, Pavlov VA, et al. Splenectomy inactivates the cholinergic antiinflammatory pathway during lethal endotoxemia and polymicrobial sepsis. J Exp Med 2006; 203:1623–8. 6. Rosas-Ballina M, Olofsson PS, Ochani M, Valdes-Ferrer SI, Levine YA, Reardon C, et al. Acetylcholine-synthesizing T cells relay neural signals in a vagus nerve circuit. Science 2011; 334:98–101. 7. Su X, Feng X, Terrando N, Yan Y, Chawla A, Koch LG, et al. Dysfunction of inflammation-resolving pathways is associated with exaggerated postoperative cognitive decline in a rat model of the metabolic syndrome. Mol Med 2012; 18:1481–90. 8. Bratton BO, Martelli D, McKinley MJ, Trevaks D, Anderson CR, McAllen RM. Neural regulation of inflammation: no neural connection from the vagus to splenic sympathetic neurons. Exper Physiol 2012; 97:1180–5. 9. Romanovsky AA. The inflammatory reflex: the current model should be revised. Exper Physiol 2012; 97:1178–9. 15. Ganchrow D, Ganchrow JR, Cicchini V, Bartel DL, Kaufman D, Girard D, et al. The nucleus of the solitary tract in the C57BL/6J mouse: Subnuclear parcellation, chorda tympani nerve projections and brain stem connections. J Comp Neurol 2013; doi: 10.1002/cne.23484. 16. Sant’Ambrogio G, Widdicombe J. Reflexes from airway rapidly adapting receptors. Respir Physiol 2001; 125:33–45. 17. Yu J, Lin S, Zhang J, Otmishi P, Guardiola JJ. Airway nociceptors activated by pro-inflammatory cytokines. Respir Physiol Neurobiol 2007; 156:116–9. 18. Nassenstein C, Kwong K, Taylor-Clark T, Kollarik M, Macglashan DM, Braun A, et al. Expression and function of the ion channel TRPA1 in vagal afferent nerves innervating mouse lungs. J Physiol 2008; 586:1595–604. 19. Barajon I, Serrao G, Arnaboldi F, Opizzi E, Ripamonti G, Balsari A, et al. Toll-like receptors 3, 4, and 7 are expressed in the enteric nervous system and dorsal root ganglia. J Histochem Cytochem 2009; 57:1013–23. 20. van Houwelingen AH, Kool M, de Jager van Heuven-Nolsen D, Kraneveld AD, derived TNF-alpha primes sensory nerve monary hypersensitivity reaction. J 168:5297–302. SC, Redegeld FA, et al. Mast cellendings in a pulImmunol 2002; 21. Saperstein S, Chen L, Oakes D, Pryhuber G, Finkelstein J. IL-1beta augments TNF-alpha-mediated inflammatory responses from lung epithelial cells. J Interferon Cytokine Res 2009; 29:273–84. 22. Chiu IM, Heesters BA, Ghasemlou N, Von Hehn CA, Zhao F, Tran J, et al. Bacteria activate sensory neurons that modulate pain and inflammation. Nature 2013; 501:52–7. 23. dos Santos CC, Shan Y, Akram A, Slutsky AS, Haitsma JJ. Neuroimmune regulation of ventilator-induced lung injury. Am J Respir Crit Care Med 2011; 183:471–82. 24. Pavlov VA, Wang H, Czura CJ, Friedman SG, Tracey KJ. The cholinergic anti-inflammatory pathway: a missing link in neuroimmunomodulation. Mol Med 2003; 9:125–34. 25. Berthoud HR, Neuhuber WL. Functional and chemical anatomy of the afferent vagal system. Auton Neurosci 2000; 85:1–17. 10. Gallowitsch-Puerta M, Pavlov VA. Neuro-immune interactions via the cholinergic anti-inflammatory pathway. Life Sci 2007; 80:2325–9. 26. Riley TP, Neal-McKinney JM, Buelow DR, Konkel ME, Simasko SM. Capsaicin-sensitive vagal afferent neurons contribute to the detection of pathogenic bacterial colonization in the gut. J Neuroimmunol 2013; 257:36–45. 11. Fox B, Bull TB, Guz A. Innervation of alveolar walls in the human lung: an electron microscopic study. J Anatomy 1980; 131:683–92. 27. Su X. Leading neutrophils to the alveoli: who is the guider? Am J Respir Crit Care Med 2012; 186:472–3. 12. Hertweck MS, Hung KS. Ultrastructural evidence for the innervation of human pulmonary alveoli. Experientia 1980; 36:112–3. 28. Giebelen IA, Leendertse M, Florquin S, van der Poll T. Stimulation of acetylcholine receptors impairs host defence during pneumococcal pneumonia. Eur Respir J 2009; 33:375–81. 13. Su X, Lee JW, Matthay ZA, Mednick G, Uchida T, Fang X, et al. Activation of the alpha7 nAChR reduces acid-induced acute lung injury in mice and rats. Am J Respir Cell Mol Biol 2007; 37:186–92. 29. Pothlichet J, Meunier I, Davis BK, Ting JP, Skamene E, von Messling V, et al. Type I IFN triggers RIG-I/TLR3/NLRP3dependent inflammasome activation in influenza A virus infected cells. PLoS Pathog 2013; 9:e1003256. Downloaded from by guest on February 5, 2015 5. Rosas-Ballina M, Ochani M, Parrish WR, Ochani K, Harris YT, Huston JM, et al. Splenic nerve is required for cholinergic antiinflammatory pathway control of TNF in endotoxemia. Proc Natl Acad Sci USA 2008; 105:11008–13. 14. Su X, Matthay MA, Malik AB. Requisite role of the cholinergic alpha7 nicotinic acetylcholine receptor pathway in suppressing Gram-negative sepsis-induced acute lung inflammatory injury. J Immunol 2010; 184:401–10.