Roles of a Mast Cell–Specific Receptor MRGPRX2 in Host Defense and Inflammation

Abstract

Mast cells are multifunctional immune cells that are found most abundantly at host-environment interfaces, such as the skin, respiratory tract, and oral/gastrointestinal mucosa. Not surprisingly, mast cells act as sentinel cells that sense microbial attacks and initiate a protective immune response and promote healing. Although mast cells share many features with other innate immune effector cells, such as neutrophils and macrophages, they uniquely interact closely with blood vessels and release an extensive set of mediators for the recruitment of innate and adaptive immune cells. A novel human G protein-coupled receptor (GPCR), known as Mas-related GPCR-X2 (MRGPRX2, mouse ortholog, MrgprB2), has recently been identified, which is expressed on mast cells but not neutrophils and macrophages. Interestingly, activation of MrgprB2 by bacteria-derived quorum-sensing peptides inhibits bacterial growth, prevents biofilm formation, and leads to the recruitment of neutrophils to effectively clear bacteria. Furthermore, host defense antimicrobial peptides and small-molecule peptide mimetics also activate mast cells via MRGPRX2/B2. MrgprB2-mediated activation of local mast cells also clears cutaneous bacterial infection, promotes healing, and protects against reinfection. In addition to their role in host defense, mast cells contribute to a number of chronic inflammatory diseases such as periodontitis, neurogenic inflammation, and inflammatory pain likely via the activation of MRGPRX2. In this review, we discuss the roles of MRGPRX2/B2 in the clearance of bacterial infection, wound healing, periodontal disease, neurogenic inflammation, and inflammatory pain. We propose that harnessing mast cells’ host defense and immunomodulatory properties via the activation of MRGPRX2 may lead to novel approaches for the treatment of drug-resistant bacterial infections. On the other hand, increased MRGPRX2 expression on mast cells and their inappropriate activation may contribute to periodontitis, neurogenic inflammation, and inflammatory pain. Thus, targeting MRGPRX2 could provide novel approaches to modulate these conditions.

Introduction

Mast cells are multifunctional tissue-resident granulocytes of hematopoietic origin. While best known as key effectors in IgE-mediated allergic reactions, mast cells have recently been recognized for their protective role in host defense against bacterial pathogens (Abraham and St John 2010; Piliponsky et al. 2010; Matsuguchi 2012; Trivedi et al. 2013; Zimmermann et al. 2019). Studies using mast cell–deficient mice demonstrated that their absence is associated with enhanced bacterial burden, decreased neutrophil recruitment, decreased immune responses, and increased mortality (Echtenacher et al. 1996; Malaviya et al. 1996; Gendrin et al. 2015; Arifuzzaman et al. 2019; Zimmermann et al. 2019). For example, in the cecal ligation and puncture (CLP) model of sepsis, mast cells are crucial for initiating innate immune responses to enterobacteria, particularly through their ability to recruit neutrophils to promote bacterial clearance (Echtenacher et al. 1996).

Mast cells possess several unique properties that make them ideal for host defense function. First, they are widely distributed throughout the body and are particularly found at the host-environment interfaces, such as the skin (Fig. 1A), respiratory tract, and oral/gastrointestinal mucosa. In the oral cavity, mast cells are found in the gingiva, periodontal ligament, and dental pulp (Gaje et al. 2016). Given their strategic location, mast cells ideally serve as professional sentinel cells and one of the first responders to encounter bacterial invasion (Metz and Maurer 2007; Abraham and St John 2010; Piliponsky et al. 2010; Trivedi et al. 2013; Cardamone et al. 2016; Zimmermann et al. 2019). Second, mast cells express a variety of receptors that enable them to recognize and respond to a broad range of pathogenic stimuli (Abraham and St John 2010; Trivedi et al. 2013). Third, upon activation, mast cells rapidly release a plethora of preformed mediators such as histamine, proteases, and prestored tumor necrosis factor–α (TNF-α) from their cytoplasmic granules (Fig. 1B), followed by generation and secretion of de novo synthesized mediators, including cytokines, chemokines, and growth and angiogenic factors (Metz and Maurer 2007; Abraham and St John 2010; Cardamone et al. 2016). These mast cell–derived mediators exert potent antimicrobial and immunomodulatory properties, which facilitate both bacterial clearance and promote healing.

Figure 1.

Mast cell localization in the skin and its ultrastructure. (A) Whole-mount immunofluorescence confocal image of adult mouse ear skin stained with Texas Red Avidin (red, mast cells) and Alexa Fluor 488–conjugated CD31 (green, endothelial cells). Bar = 50 µm. (B) Freshly isolated peritoneal lavage cells from mice were used for transmission electron microcopy to visualize mast cell granules. “N” is depicted as mast cell nucleus and “G” is depicted as mast cell granules. Bar = 1 μm.

Periodontitis is a chronic inflammatory disease that affects the tooth-supporting structures, resulting in progressive destruction of the periodontal ligament and alveolar bone (Darveau 2010). Bacterial plaque has been implicated as the primary etiological factor in the pathogenesis of periodontitis. A subset of specific gram-negative periodontal bacteria, so-called red complex pathogens, including Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola, are frequently isolated together in disease sites and strongly associated with disease (Holt and Ebersole 2005). Of these, P. gingivalis is considered the “keystone pathogen” of periodontitis (Hajishengallis et al. 2012). However, while the presence of specific pathogenic bacteria can trigger microbial dysbiosis characteristics and play a crucial role in the initiation and progression of periodontitis, emerging evidence shows that the host response to bacterial challenge is central to tissue destruction and alveolar bone resorption, which is the hallmark of periodontitis (Darveau 2010; Hajishengallis et al. 2012; Hajishengallis 2014). Among the cells found in the periodontal tissues, mast cells have been detected in both healthy and inflamed gingiva. Mast cell numbers are increased in gingiva of patients with chronic periodontitis, and the degree of degranulation and proinflammatory cytokine production from these cells correlates with disease severity (Huang et al. 2013; Agrawal et al. 2016; Malcolm et al. 2016; Gupta et al. 2017; Tada et al. 2019). Furthermore, mast cell–deficient mice are protected from P. gingivalis–induced bone loss when compared to wild-type (WT) mice (Malcolm et al. 2016), indicating a potential role of mast cells in the pathogenesis of periodontitis.

In addition to periodontitis, inappropriate activation of mast cells has been implicated in neurogenic inflammation and pain. Mast cells are found in close proximity to nerve endings and express receptors for a range of neurotransmitters (Forsythe 2019). Upon activation by neuropeptides such as substance P (SP), mast cells release a broad range of proinflammatory cytokines and chemokines, including TNF-α, CCL2, and CCL3, that alter vascular permeability and promote the immune cell recruitment to the affected site (Green et al. 2019). The recruitment of immune cells, including neutrophils and monocytes, mediated by mast cells amplifies local inflammatory responses and facilitates peripheral nociceptor sensitization, contributing to neurogenic inflammation and inflammatory pain (Ren and Dubner 2010; Green et al. 2019).

One of the most recent developments in mast cell biology research has been the discovery of a novel G protein-coupled receptor (GPCR) known as Mas-related GPCR-X2 (MRGPRX2; mouse counterpart MrgprB2). MRGPRX2/B2 is expressed predominantly in connective tissue mast cells (Tatemoto et al. 2006; Motakis et al. 2014; McNeil et al. 2015; Varricchi et al. 2019). Emerging evidence suggests that the activation of mast cells via MRGPRX2/B2 is pivotal to controlling bacterial infection and promoting wound healing (Arifuzzaman et al. 2019; Pundir et al. 2019). On the other hand, inappropriate mast cell activation contributes to chronic inflammatory diseases and inflammatory pain, likely via MRGPRX2/B2 (Subramanian et al. 2016). Our recent study demonstrated that the number of MRGPRX2-expressing mast cells is increased in patients with chronic periodontitis (Gupta et al. 2017). Thus, the main objectives of this article are to discuss the recent advances in our understanding of the roles of MRGPRX2/B2 expressed on mast cells in host defense to bacterial infection, periodontitis, and inflammatory pain.

Discovery of a Novel Mast Cell Receptor MRGPRX2

Unlike other immune cells, mast cells display considerable heterogeneity (Welle 1997; Galli et al. 2011). In humans, mast cells are generally categorized into 2 subtypes based on the protease contents in their granules: mast cells that contain both tryptase and chymase (MC_TC) are predominantly found in connective tissues such as the skin and gingiva, while those that contain only tryptase (MC_T) are mostly found in the lungs (Welle 1997; Gurish and Austen 2012; Gupta et al. 2017). MC_TC contain abundant heparin in their granules, but MC_T do not.

Mast cells are sometimes referred to as constitutive (innate) and mucosal (adaptive) not only based on their tissue distribution and granularity but also on their development patterns and expression of cell surface receptors. MC_TC are considered innate mast cells as they are present constitutively in connective tissues and are generally unaffected by T cells. By contrast, adaptive mast cells are found predominantly in intraepithelial tissues such as the lungs and are induced by T-cell-dependent inflammation (Dwyer et al. 2016). Furthermore, MC_TC express MRGPRX2 on the cell surfaces, whereas MC_T do not (Subramanian et al. 2016) (Table). Thus, while both human mast cell subtypes can be activated via high-affinity IgE receptors (FcεRI), only MC_TC are responsive to a diverse range of cationic substances, collectively called basic secretagogues, through the activation of MRGPRX2 (Tatemoto et al. 2006; Fujisawa et al. 2014).

Table.

Different Properties of Mast Cell Subtypes.

Characteristics	Mast Cell Subtypes
Human	MCTC (tryptase/chymase)	MCT
Rodent	Connective tissue-type mast cells	Mucosal-type mast cells
Tissue	Skin, gingiva	Lungs
Type	Innate	Adaptive
T cell	Independent	Dependent
Function	Homeostasis/host defense	Immunologic
Receptors	FcεRI and MRGPRX2/B2	FcεRI

Initial identification of MRGPRX2 as a mast cell–specific receptor for basic secretagogues was established from the study by Tatemoto et al. (2006). They demonstrated that a number of basic secretagogues, such as neuropeptide substance P, pituitary adenylate cyclase–activating polypeptide, and compound 48/80, activate CD34⁺ cell-derived mast cells through this receptor. Subsequent studies showed that MRGPRX2 is also expressed in human skin and gingival mast cells (Fujisawa et al. 2014; Gupta et al. 2017). The presence of 22 potential MRG coding genes in the mouse genome initially made it difficult to identify the putative MRGPRX2 ortholog in mice. However, in 2015, a seminal article by McNeil et al. (2015) demonstrated that mouse peritoneal mast cells solely express MrgprB2 messenger RNA (mRNA), and that MrgprB2 serves as the murine ortholog of human MRGPRX2. Emerging evidence now suggests that activation of mast cells via MRGPRX2/B2 plays a major role in the immune response to bacterial infection (Pundir et al. 2019).

MRGPRX2/B2 Detects Bacterial Quorum Sensing Molecules and Mediates Antibacterial Immunity

Several Gram-positive and Gram-negative bacteria secrete an extracellular chemical signal, termed an autoinducer, which, upon reaching a threshold concentration, leads to an increase in bacterial cell population. In a particular environment, these autoinducers, collectively called quorum-sensing molecules (QSMs), synchronize the bacterial behavior to function as a multicellular organism (Whiteley et al. 2017). Bacteria use QSMs to communicate and modulate their behavior changes in antibiotic resistance, biofilm formation, and virulence (Whiteley et al. 2017; Pundir et al. 2019). A recent study by Pundir et al. (2019) showed that a number of Gram-positive bacterial QSMs such as competence-stimulating peptide-1 (CSP-1) from Streptococcus pneumoniae, Entf metabolite from Enterococcus faecium, and Streptin-1 from Streptococcus pyogenes are recognized by mast cells through MRGPRX2/B2. Of these, CSP-1 is the most potent candidate to induce antibacterial host response via MRGPRX2/B2-mediated mast cell activation (Pundir et al. 2019). Mast cell activation by QSMs leads to neutrophil recruitment and production of reactive oxygen species (ROS), which eliminates the bacterial infection and reduces biofilm biomass. These antibacterial effects are mediated through MRGPRX2 as the levels of ROS and cytokine/chemokine production are reduced upon knocking down MRGPRX2 in human mast cells. Similarly, WT mouse peritoneal mast cells (PMCs) but not MrgprB2^MUT PMCs induced a similar antibacterial effect (Pundir et al. 2019). CSP-precursor Com is required for virulence during nasopharyngeal colonization. Mutant D39 strain devoid of either comC (dComC) or autoinducer 2 (dLuxS) is resistant to mast cell (MRGPRX2/B2)–mediated killing. Intranasal inoculation with S. pneumoniae, intraperitoneal infection with E. faecium, or skin infection by S. pyogenes cause enhanced bacterial clearance in WT mice compared to MrgprB2^MUT mice (Pundir et al. 2019). Finally, injection of MRGPRX2/B2 agonist CSP-1 or compound 48/80 locally at the site of bacterial infection significantly reduce in vivo local bacterial counts, establishing MRGPRX2/B2 as a pharmacological candidate for the modulation of bacterial infection (Fig. 2).

Figure 2.

MRGPRX2/B2 act as receptors for quorum-sensing molecules (QSMs) and host defense peptides to mediate host defense. QSMs act as an extracellular signal for bacteria to facilitating cell-cell communication. Mast cells detect QSMs by MRGPRX2/B2 and induce degranulation, which releases mediators such as histamine and tumor necrosis factor–α (TNF-α). These mediators increase local vascular permeability to recruit neutrophils in the infection site, which in turn produces reactive oxygen species (ROS). ROS act as effector molecules by eliminating biofilm formation and inhibiting bacterial growth. Moreover, during infection, epithelial cells secrete host defense peptides (HDPs), which can also activate mast cells by MRGPRX2/B2 and induce degranulation to release histamine. Histamine further activates epithelial cells to produce more HDPs in a feedback loop. Small-molecule host defense peptide mimetics (smHDPMs), which have direct antimicrobial activity, also activate mast cells by MRGPRX2/B2.

Although the QSMs investigated by Pundir et al. (2019) are of nonoral organisms, oral bacteria also produce a number of QSMs to enable communication between mixed bacterial communities of the oral biofilm (Miller and Lamont 2019). Biofilm formation is one of the crucial virulence factors of oral pathogens for nutrient acquisition and processing, protection from immune responses, and increased tolerance to antimicrobial agents (Mira et al. 2017; Jiao et al. 2019). Periodontal pathogens, such as P. gingivalis, Fusobacterium nucleatum, and Aggregatibacter actinomycetemcomitans, as well as oral streptococci have been shown to form biofilms and possess the autoinducer 2 quorum-sensing system for signal exchange (Frias et al. 2001; Basavaraju et al. 2016; Miller and Lamont 2019). Compared to commensal bacteria, periodontal pathogens produce higher levels of autoinducer 2, which facilitate the development and maturation of biofilm, subsequently leading to periodontitis (Kolenbrander et al. 2010). For instance, autoinducer 2 produced by F. nucleatum stimulates the coaggregation and expression of adhesion molecules of P. gingivalis (Jang et al. 2013). Autoinducer 2 inhibition by quorum-sensing inhibitors, BMK-Q101 (a synthesized furanone analogue) and D-ribose, reduces alveolar bone loss induced by P. gingivalis/F. nucleatum coinfection in vivo (Ben Amara et al. 2018). It is therefore possible that oral bacterial QSMs activate mast cells via MRGPRX2/B2 to reduce biofilm mass, thus promoting gingival homeostasis.

Host Defense Peptides Contribute to Mast Cell–Mediated Bacterial Killing

Host defense peptides (HDPs) are positively charged amphipathic peptides that act as broad-spectrum antibiotics against bacteria, fungi, and viruses by interacting with their negatively charged phospholipid moieties and disrupting their membranes (Hazlett and Wu 2011). Mast cells display direct bactericidal activity through the expression and production of HDPs, such as cathelicidin LL-37, human β-defensins (hBDs), and a recently described lipocalin 2 (Di Nardo et al. 2003; Di Nardo et al. 2008; Chang et al. 2019). Secretion of cathelicidins by mast cells leads to protective antibacterial effects to prevent and control group A Streptococcus skin infection (Di Nardo et al. 2003; Di Nardo et al. 2008). A study by Di Nardo et al. (2008) demonstrated that mast cell–deficient mice developed larger skin lesions with higher numbers of bacteria and delayed recovery after group A Streptococcus infection compared to the WT mice. Reconstitution of those mice with WT mast cells, but not cathelicidin-deficient mast cells, significantly reduced edema formation and wound extension, indicating the importance of cathelicidin in controlling bacterial infection (Di Nardo et al. 2008). Furthermore, Chang et al. (2019) recently identified a novel HDP, lipocalin 2, expressed in human and mouse mast cells. Lipocalin 2 released from mast cells inhibits Escherichia coli growth, suggesting that it plays an important role in protection against E. coli infection (Chang et al. 2019).

Apart from the direct bactericidal effect, HDPs also demonstrate indirect antimicrobial activities through mast cell–dependent recruitment and activation of other immune cells (Hancock et al. 2016). These immunomodulatory features likely contribute to their effectiveness as antimicrobial agents. HDPs such as LL-37 are potent chemoattractants for mast cells and can induce mast cell degranulation (Niyonsaba et al. 2007; Babolewska and Brzezinska-Blaszczyk 2015). They also increase the expression of Toll-like receptor 4 on the mast cell surface, which may enhance the ability of mast cells to detect invading pathogens (Yoshioka et al. 2008).

MRGPRX2 Is a Novel Mast Cell Receptor for Host Defense Peptides

Using transfected cell lines and short hairpin RNA (shRNA)–mediated silencing in primary human mast cells, our lab was the first to demonstrate that LL-37 induces degranulation, chemokine generation, and chemotaxis via the activation of MRGPRX2 (Subramanian et al. 2011). hBDs are produced from skin and gingival epithelial cells upon microbial assault and consist of 4 members, hBD1–4 (Weinberg et al. 2012; Greer et al. 2013). Of these, hBD1–3 are expressed and secreted in the human gingival tissue. In vitro studies have demonstrated that P. gingivalis induces the expression of hBD1 (Vankeerberghen et al. 2005) and hBD2 (Taguchi and Imai 2006) in cultured human gingival epithelial cells, while F. nucleatum and A. actinomycetemcomitans stimulate the production of hBD2 and hBD3 (Feucht et al. 2003; Vankeerberghen et al. 2005). We demonstrated that hBD2 and hBD3 induce mast cell degranulation via MRGPRX2 (Subramanian et al. 2011; Subramanian et al. 2013). HDP-induced mast cell degranulation during bacterial infection likely provides critical antimicrobial host defense by enhancing vascular permeability and initiating the recruitment of neutrophils at the site of infection (Chen et al. 2007; Subramanian et al. 2016). Moreover, mast cell–derived mediators such as histamine can, in turn, induce hBD2 and hBD3 secretion from human epithelial cells (Ishikawa et al. 2009), which contributes to further mast cell degranulation, indicating the biphasic innate function of mast cells. Thus, QSMs and HDPs, which activate MC_TC-type mast cells via MRGPRX2/B2, may provide a general mechanism for host defense in the skin and gingiva (Table and Fig. 2).

Small-Molecule Host Defense Peptide Mimetic Antibacterial Agents Activate Mast Cells via MRGPRX2/B2

Although HDPs have an important therapeutic potential against microbial infection, their metabolic instability and cellular cytotoxicity have limited their utility. To overcome these limitations, a series of small-molecule HDP mimetics (smHDPMs) have been developed (Choi et al. 2009; Scott and Tew 2017). These compounds are relatively inexpensive to synthesize and have distinct advantages over HDPs in terms of stability, bioavailability, and low toxicity. Furthermore, these compounds exhibit potent antibacterial activity (Beckloff et al. 2007). Recently, we tested the effects of 5 smHDPMs on cytotoxicity, antimicrobial activity, and mast cell degranulation. None of these smHDPMs displayed cytotoxicity against mouse 3T3 fibroblasts or human-transformed liver HepG2 cells, but all compounds induced degranulation in human and murine mast cells via MRGPRX2 and MrgprB2, respectively (Alkanfari et al. 2019). These findings suggest that smHDPMs could serve as novel targets for the treatment of bacterial infection because of their ability to harness mast cells’ host defense functions (Fig. 2).

Mast Cell Activation via MrgprB2 Leads to Neutrophil Recruitment and Accelerated Bacterial Clearance

Mastoparan, a peptide toxin from wasp venom, displays antibacterial activity and induces mast cell degranulation via MRGPRX2/B2 (McNeil et al. 2015). In a Staphylococcus aureus–induced skin infection mouse model, topical application of mastoparan promotes neutrophil recruitment, and this action is highly dependent on the presence of MrgprB2-expressing mast cells (Arifuzzaman et al. 2019). A mastoparan derivative (MP-6I), which does not activate mast cells, failed to clear S. aureus infection despite having potent antibacterial property. By contrast, Duke Mast F, a mastoparan derivative with mast cell–degranulating capacity without any antibacterial activity, reduced the lesion size significantly compared to vehicle (Arifuzzaman et al. 2019). These results suggest that the therapeutic benefit of mastoparan depends on its mast cell–activating capacity rather than antibacterial property. Given that mastoparan activates mast cells via MRGPRX2/B2, it is likely that its host defense function is mediated via the activation of these receptors.

Mast Cell Activation via MrgprB2 Promotes Regenerative Healing

Wound healing is a coordinated, multicomponent, progressive series of events intended to restore the barrier function and mechanical integrity of the skin breached by mechanical or microbial assault (Schultz et al. 2011). It is well documented that persistence of bacteria at wound sites delays natural wound healing and increases scarring (Singer and McClain 2002). To determine if mastoparan-mediated acceleration in bacterial clearance, as described above, results in faster skin regeneration and reduced scarring, Arifuzzaman et al. (2019) allowed infected lesions to close completely and proliferative processes in the skin to reach steady-state levels. Analysis of skin sections with trichrome staining demonstrated reduced scarring in mastoparan-treated mice when compared with untreated mice (Arifuzzaman et al. 2019). Dermal CD301b⁺ dendritic cells play critical roles in promoting reepithelialization of sterile wounds (Yang et al. 2017), and this population is significantly decreased in S. aureus–infected skin but restored in later stages of infection. Furthermore, depletion of MrgprB2-expressing mast cells reduces CD301b⁺ cells, indicating that mast cells contribute to restoring the skin CD301b⁺ cell population to homeostatic levels (Arifuzzaman et al. 2019). Based on these findings, Arifuzzaman et al. (2019) proposed that mast cells contribute to the healing of infectious wounds not only by clearing bacteria via neutrophil recruitment but also by restoring the CD301b⁺ cell population, which promotes reepithelialization (Fig. 3). In addition, in a Pseudomonas aeruginosa–infected murine skin wound model, mast cell–deficient mice exhibited significantly delayed wound closure, which is associated with impaired bacterial clearance. However, reconstitution of mast cells restored both bacterial clearance and wound closure (Zimmermann et al. 2019). Thus, mast cell activation via MRGPRX2/B2 may contribute to the accelerated clearance of bacterial infection and regenerative healing.

Figure 3.

Mastoparan activates mast cells through MRGPRX2/B2 and generates innate and adaptive immunity against Staphylococcus aureus infection. Application of mastoparan in olive oil/DMSO mixture activates mast cells via MRGPRX2/B2 during S. aureus–mediated skin infection and results in mast cell degranulation and release of mediators. This increases vascular permeability and recruits neutrophils. Mastoparan-mediated neutrophil recruitment eliminates bacterial infection by reactive oxygen species (ROS) generation. Mast cells also release cytokines such as CCL2, CCL3, CCL4, and tumor necrosis factor–α (TNF-α), which facilitate the recruitment of CD301b⁺ dermal dendritic cells (DDCs) in the infection and mediate regenerative healing. Moreover, mastoparan treatment also produces S. aureus–specific IgG. These altogether boost antibacterial adaptive immunity and controls reinfection.

Mast Cell Activation via MrgprB2 Promotes Adaptive Immunity and Controls Bacterial Reinfection

In addition to neutrophil and dendritic CD301b⁺ cell recruitment, mast cells are responsible for recruiting antigen-presenting dendritic cells to the site of infection and promoting their migration from the inflamed site to the draining lymph nodes (Shelburne and Abraham 2011). Mastoparan induces the generation of chemokines such as CCL2, CCL3, and CCL4, which contribute to the mobilization of dendritic cells. Topical application of mastoparan into S. aureus–infected wound results in a significantly higher level of S. aureus–specific total IgG compared to vehicle control group. Furthermore, this increased IgG is associated with reduced lesion size following reinfection (Arifuzzaman et al. 2019). Based on these findings, it has been proposed that mastoparan not only accelerates bacterial clearance and promotes regenerative healing but is also capable of enhancing adaptive immunity, which provides protection against reinfection (Fig. 3).

Potential Roles of MRGPRX2 in Periodontal Tissue Homeostasis and Periodontitis

Mast cells of the MC_TC type, which express MRGPRX2, are found in healthy gingiva and have been implicated in periodontal tissue homeostasis (Steinsvoll et al. 2004; Gupta et al. 2017). Mast cells are activated by HDPs generated from gingival epithelium (hBD3) and infiltrating neutrophils (LL-37) via the activation of MRGPRX2 (Subramanian et al. 2011; Subramanian et al. 2013). Furthermore, mast cells may sense QSMs produced from periodontal pathogens in bacterial plaque via MRGPRX2, as discussed earlier. Upon activation, mast cells release several mediators, including cytokines and chemokines, that promote host defense by facilitating neutrophil recruitment through the periodontal tissue, resulting in maintenance of tissue homeostasis (Steinsvoll et al. 2004) (Fig. 4).

Figure 4.

MRGPRX2 contributes to both periodontal tissue homeostasis and periodontitis. During infection, host defense peptides (HDPs) produced from gingival epithelium activate mast cells through MRGPRX2, which undergo degranulation to release preformed mediators and tumor necrosis factor–α (TNF-α). This causes increased vascular permeability and increased migration of neutrophils in the infected tissue. Neutrophils clear microbial infection through effector molecule (reactive oxygen species [ROS]) secretion and maintain tissue homeostasis. In contrast, during chronic infections such as periodontitis, overproduction of HDPs from gingival epithelium leads to recruitment of more mast cells, and its activation by MRGPRX2 releases more proinflammatory cytokines such as interleukin (IL)–6, IL-1β, and TNF-α. This results in periodontal tissue breakdown, a major characteristic feature of periodontitis.

While MRGPRX2-mediated low-level inflammation may contribute to periodontal homeostasis, sustained inflammation due to MRGPRX2 dysregulation may, however, promote periodontal disease. The number of MRGPRX2-expressing mast cells is increased in gingiva obtained from chronic periodontitis patients compared with those with normal gingiva (Gupta et al. 2017). Of note, an increased number and expression of MRGPRX2 have been reported in skin mast cells from patients with chronic urticaria and have been linked to the pathogenesis of the disease (Fujisawa et al. 2014). It is noteworthy that expression of HDPs is upregulated in patients with periodontitis when compared to healthy individuals (Turkoglu et al. 2009; Pereira et al. 2013; Wang et al. 2015). Following activation by HDPs, mast cells produce and release an array of proinflammatory mediators, including TNF-α and interleukin (IL)–1, IL-6, and IL-31, which have been shown to induce gingival epithelial barrier dysfunction and alveolar bone loss (Malcolm et al. 2016; Tada et al. 2019). Thus, enhanced HDP/MRGPRX2-mediated mast cell degranulation and cytokine generation may lead to periodontal tissue breakdown, which is the characteristic feature of periodontitis (Fig. 4).

Novel Roles of MRGPRX2 on Neurogenic Inflammation and Pain

Mast cells are often found in close proximity to sensory nerve endings in most tissues and appear to serve as a functional homeostatic regulatory unit (Subramanian et al. 2016). However, dysregulation of mast cell–nerve interaction has been suggested as a major contributor in the pathogenesis of neurogenic inflammation and pain (Ren and Dubner 2010; Forsythe 2019). Mast cell–derived mediators such as histamine, leukotrienes, and tryptase interact with their specific receptors on sensory nerve endings, resulting in the release of neuropeptides, including SP (Steinhoff et al. 2000; Shim and Oh 2008; Taylor-Clark et al. 2008). In turn, SP released from nerve endings can evoke further mast cell degranulation, subsequently resulting in the release of a wide array of proinflammatory cytokines and chemokines, as well as the recruitment of innate immune cells (Kulka et al. 2008; Green et al. 2019).

SP has been well recognized for its roles in inflammation and pain. SP-induced neurogenic inflammation has been implicated in the pathogenesis of many chronic inflammatory conditions, including orofacial pain and inflammatory processes in pulpitis and periodontal diseases (Sacerdote and Levrini 2012; de Avila et al. 2014). Neurokinin-1 receptor (NK-1R) was previously thought to be responsible for SP-mediated pain and neurogenic inflammation due to its close anatomical distribution and functional role in pain pathways (Garcia-Recio and Gascon 2015). However, the failure of NK-1R antagonists as potential anti-inflammatory and analgesic therapeutics in human clinical studies raises the possibility that nociceptive and proinflammatory effects of SP may be mediated via alternative mechanisms rather than the interaction with NK-1R (Navratilova and Porreca 2019).

Recently, a landmark study by Green et al. (2019) provided strong support that the inflammatory effects of SP are mediated via the activation of mast cells through MRGPRX2/B2, independent of NK-1R. Activation of MRGPRX2/B2 is required for inflammatory and thermal hyperalgesia; release of proinflammatory cytokines and chemokines such as TNF-α, CCL2, and CCL3; and recruitment of immune cells at the injury site (Green et al. 2019). This immune cell recruitment reinforces the pathophysiology of neurogenic inflammation and inflammatory pain as it further facilitates inflammatory responses and peripheral nerve sensitization (Navratilova and Porreca 2019) (Fig. 5). Preventing the activation of mast cells from released SP either by blocking the MrgprB2 receptor or by ablation of mast cells greatly diminishes pain responses (Green et al. 2019). Thus, targeting MRGPRX2/B2 may lead to novel effective treatment modalities for inflammatory pain.

Figure 5.

The neuropeptide substance P (SP) contributes to neurogenic inflammation and pain through the activation of mast cells via MRGPRX2. Mast cells are found in close proximity to peripheral nerve endings. During tissue damage, SP released from peripheral nerve endings activates mast cells via MRGPRX2. Activated mast cells release several proinflammatory cytokines and chemokines such as CCL2 and CCL3, and they recruit immune cells in the injury site. These lead to the pathophysiology of neurogenic inflammation and peripheral neuron sensitization, resulting in pain sensation.

Therapeutic Prospects and Future Directions

The emergence of antibiotic resistance poses a tremendous public health concern globally and requires an urgent need to develop a novel therapy for the treatment of infections caused by drug-resistant microbes (Laxminarayan et al. 2013). Brilacidin, a defensin mimetic antibiotic, is being developed as a novel treatment for drug-resistant bacterial infections (Kowalski et al. 2016). It is also being investigated for possible application in the prevention of mucositis in patients undergoing chemoradiation for treatment of head and neck cancer (Sonis and Villa 2018). Based on the recent findings on mastoparan (Arifuzzaman et al. 2019) and smHDPMs (Alkanfari et al. 2019), it is possible that the potential clinical utility of brilacidin may be mediated by harnessing immunomodulatory properties through the activation of MRGPRX2, in addition to its direct antimicrobial activity. Taken together, these findings raise the interesting possibility that exploiting MRGPRX2-mediated mast cell activation by host defense peptidomimetics could provide a novel approach for the treatment of drug-resistant bacterial infections.

Although MRGPRX2-mediated mast cell activation could contribute to periodontal homeostasis, its inappropriate activation may lead to chronic inflammatory diseases, including periodontitis and inflammatory pain. The expression of MRGPRX2 and its ligands is increased in patients with chronic periodontitis (Gupta et al. 2017). Enhanced activation of MRGPRX2 by HDPs and neuropeptides may contribute to the inflammatory processes responsible for periodontal breakdown and inflammatory pain, respectively. Thus, development of MRGPRX2 antagonists targeting the receptor might therefore represents a novel and effective therapeutics approach for the prevention/treatment of periodontitis and inflammatory pain.

Author Contributions

C. Chompunud Na Ayudhya, S. Roy, M. Thapaliya, contributed to data acquisition and analysis, drafted and critically revised the manuscript; H. Ali, contributed to conception and design, drafted and critically revised the manuscript. All authors gave final approval and agree to be accountable for all aspects of the work.