2 | RESEARCH OVERVIEW ON CAP MECHANISM

Among the key parts of CAP, the effects of vagus nerve and α7nAChR are relatively identified. In CAP, muscarinic receptors (mAChR), microRNAs (miRNAs), β2 adrenergic receptors, spleen also play a role in it (Figure 2).

2.1 | Vagus nerve

Anti-inflammatory effect of CAP depends on the integrity of the vagus nerve. Vagus nerve is the connector of brain and immune system (spleen, liver, etc.). Inflammatory cytokines released by macrophages, dendritic cells, and immune cells could locally activate vagus nerve fibers. Signal is transmitted to nucleus tractus solitarius (NTS) in the medulla oblongata (Tracey, 2009). NTS takes the integrated information to make a decision and project it into efferent vagus nerve in the dorsal motor nucleus, prompting ACh to release at the target organ and activating cholinergic receptors in macrophages, such as α7nAChR, thereby inhibiting the generation of inflammatory cytokines (Ulloa, 2005). Vagus nerve regulation on TNF is one of the most deeply studied anti-inflammatory mechanisms, and it also reduces the levels of IL-1β, IL-6, and high mobility group box 1 (HMGB1). Parasympathetic nerves are involved in controlling the inflammatory responses, while skin and skeletal muscles can also exhibit cholinergic anti-inflammatory regulation without vagal innervation. Studies have indicated that vagus nerve is not a proprietary attribute of inflammatory cholinergic regulation (Pereira & Leite, 2016). Thus, the regulation of other parts of CAP needs further study.

2.2 | Nicotinic receptor

Cholinergic receptors are nicotinic receptors (nAChR) and mAChR, the anti-inflammatory effects of cholinergic transmitters are mainly mediated by nAChR. nAChR is a member of four transmembrane domains superfamilies of neurotransmitter-gated ion channels. These receptors play a central role in regulating synaptic nerve transmission and basic intracellular signaling, and controlling the viability of neuronal and the structure of synaptic. Moreover, α7nAChR is the closest receptor to inflammation and has a high affinity for α-silver snake toxin. It is a homopentamer that composed of five α7 subunits and is extensively distributed among macrophages, endothelial cells, lymphocytes, glial cells, and intestinal epithelial cells, etc. Besides, α7nAChR is associated with obesity, diabetes, AD, arthritis, asthma, atherosclerosis, cancer, and psoriasis (Bencherif, Lippiello, Lucas, & Marrero, 2011). Activation of α7nAChR contained in lymphocytes and macrophages can regulate their activation, and macrophages mediate the physiological control of cytokine production by autocrine/paracrine ACh through nAChR (De Rosa, Dionisio, Agriello, Bouzat, & Esandi Mdel, 2009; Gotti & Clementi, 2004). Activation of α7nAChR can suppress the release of inflammatory cytokines such as TNF, IL-1β, IL-6, IL-8 and HMGB1. ACh is the most direct agonist affecting α7nAChR, and related enzymes such as AChE may have potential therapeutic significance. The inhibition of AChE is a key way to treat diseases such as AD(Y. Jiang, Gao, & Turdu, 2017). Therefore, α7nAChR and ACh-related enzymes have become important targets for inflammation-related diseases treatment.

2.3 | Muscarinic receptors

The muscarinic receptor (mAChR) is a seven-transmembrane domain G protein-coupled receptor composed of five different subtypes, namely M1-M5. It can mediate many biological effects, including the regulation of heart rate, glandular secretion, smooth muscle contraction, and central nervous system (Kruse et al., 2014). The role of peripheral mAChR in inflammation is still controversial. Intravenous injection of mAChR inhibitor atropine methyl nitrate in endotoxemia rats cannot eliminate the inhibitory effect of vagal nerve stimulation on TNF, nor can it inhibit serum TNF(Pavlov et al., 2006). Another study found that M1 and M5 mAChRs are involved in regulating the production of cytokines (such as IL-6), resulting in the switch regulation of antibody class from IgM to IgG1, up-regulating immune response of protein antigens, but not participating in the initial production response of antibodies (Fujii et al., 2007). However, further validation of central mAChR agonists found that the stimulation of nAChR and mAChR significantly reduced the levels of pro-inflammatory cytokines like IL-1β, IL-6, macrophage inflammatory protein 2 (MIP-2), and animal mortality in early sepsis. CAP are regulated mainly by stimulating central mAChR and peripheral 7nAChR (Zabrodskii, Lim, Shekhter, & Kuzmin, 2012). Especially in central inflammatory diseases, it may be closely related to central mAChR, and the mechanism research between them needs to deepen on mAChR and central inflammation.

2.4 | β2 adrenergic receptor

Adrenergic receptors (AR) are a large family of seven transmembrane receptors which respond to catecholamines. Both primary and secondary immune organs are dominated by plenty of sympathetic nerve fibers emitted from the sympathetic postganglionic fibers. When the sympathetic nervous system is excited, the end of the sympathetic nerves that govern each immune system mainly release norepinephrine (NE), which in turn causes a local immune response. Vagus nerve stimulation relies on NE released from splenic sympathetic nerve endings, followed by β2AR activation in T lymphocytes (Vida et al., 2011). High concentrations of NE activate lymphocytes, thereby producing ACh in vitro (Rosas-Ballina et al., 2011). These findings indicate that β2AR activation imitates CAP. At the same time, β2AR activation can also prevent the development of sepsis through additional mechanisms in the innate immune system, such as directly inhibiting cytokine production or increasing bacterial phagocytosis in macrophages (Muthu et al., 2010; Severn, Rapson, Hunter, & Liew, 1992).

2.5 | miRNAs

miRNAs regulate gene expression in various biological processes. About 5000-10000 miRNAs are expressed in mammalian cells, which regulate more than 60% of protein-coding genes. miRNAs rapidly stop mRNA translation and adjust specific genes expression, prevent the ability of mRNA translation and degradation more quickly and stronger. It enable CAP to moderate the acute production of inflammatory cytokines, and limit the potentially excessively enthusiastic secondary response while maintaining controlled inflammation monitoring (Ulloa, 2013). miRNAs do not represent an effective mechanism for completely shutting down gene expression. Instead, they usually function as a “regulatory” mechanism that regulates gene expression within range of 1-4 times. This is a typical feature of the cholinergic anti-inflammatory pathway. It uses chemical instability and short life cycle of ACh and miRNAs to quickly regulate inflammation and avoid tissue damage, but maintains baseline expression of infection. Compared with the immunosuppressive effects of glucocorticoids, these mechanisms have biological advantages. Clinical studies have revealed that the function of macrophages in children with intestinal failure is excessively activated. miRNA-124 directly targets the 3’ untranslated region of STAT3 and AChE mRNA, inhibits macrophage activation, reduces IL-6 and TNF-α release, and alleviates abdominal inflammation (Y. T. Xiao, Wang, Lu, Cao, & Cai, 2016). Nicotinic acid can induce miRNA-124 through α7nAChR to inhibit STAT3 tyrosine phosphorylation and protein expression, and negatively regulate the inflammation caused by Toll-like receptor 4 (TLR4) (Qin, Wang, Su, & Liu, 2016; Y. Sun et al., 2013). miRNA-124a, miRNA-132 and miRNA-205-5b are also the key mediator of cholinergic anti-inflammatory activity (Q. Li, 2011; F. Liu et al., 2015; W. Zhou, Wang, Li, Li, & Sang, 2016). At present, there are not many drugs studied from this angle, and it is necessary to further clarify which miRNAs is more closely related to CAP.

2.6 | Spleen

Anatomically, spleen is necessary for vagus nerve to control inflammatory response (Huston et al., 2008). Spleen is also the main source of serum TNF in endotoxemia, and TNF enters systemic circulation from splenic vein through liver. Vagus nerve regulates cytokines production through two-stage tandem neurons. Preganglionic neurons are located in the dorsal nucleus of vagus motor. Postganglionic neurons come from the superior mesenteric plexus of the abdominal cavity (Agostoni, Chinnock, De Daly, & Murray, 1957). Splenic nerve fibers are composed of catecholamine fibers (Klein, Wilson, Dzielak, Yang, & Viveros, 1982). Similarly, spleen produces ACh, but no ACh nerve fibers have been detected (Nance & Sanders, 2007), and it is still not completely clear whether spleen is innervated by vagus nerve. Studies have found that reserpine consumes catecholamines to eliminate vagal nerve’s inhibition of TNF (Nezhinskaia, Vladykin, & Sapronov, 2010). After spleen resection, vagal nerve stimulation specifically reduces TNF production from macrophages in the red pulp and marginal area (Rosas-Ballina et al., 2008). And the effect of nicotine on reducing the level of HMGB1 in the blood of sepsis animals is inhibited (X. M. Song et al., 2008).