Mast Cells: Sentinels of Innate Skin Immunity

TLR

Ligand [102, 103]

TLR1

Lipopeptide (Pam3csk4)

TLR2

Peptidoglycan, Zymosan, LTA, Lipopeptide (Pam3csk4)

TLR3

PolyI:C (dsRNA)

TLR4

LPS, RSV protein F, Mycobacterium tuberculosis

TLR6

Peptidoglycan, Zymosan

TLR9

Bacterial DNA (CpG motifs)

For example, lipopeptides from various gram-positive bacteria can bind to both TLR1 and TLR2 inducing heterodimer TLR1/2 formation and subsequent downstream signaling. Similarly, peptidoglycans (PGNs) and zymosan, components of microbial cell walls, cause heterodimerization between TLR 2 and 6. Despite the wide array of stimulatory ligands and TLRs expressed on the mast cell, a common downstream signaling pathway leads to activation of the transcription factor NFkB and the MAPKs, p38 and JNK [102–104].

Mast Cell Response to Bacteria

Due to the mast cells’ strategic location in the skin, they play an important role in innate immunity against bacteria through their ability to phagocytose pathogens, present bacterial antigens to T cells, recruit other phagocytic cells, release mediators and cytokines, and produce cathelicidin antimicrobial peptides [105–107].

Some of the earliest studies in mast cell responses to bacteria show that rodent mast cells are capable of phagocytosing bacteria via complement receptors [108]. Additional studies in complement C3-deficient mice demonstrate a decrease in peritoneal mast cell degranulation, production of TNFα, neutrophil infiltration and clearance of bacteria. Treating these mice with purified C3 protein reversed these defects, confirming that complement activation aids the mast cell’s full functionality in innate immune defense [109]. Additional preliminary research shows that certain bacteria are able to induce mast cell degranulation. Formalin-killed bacteria such as Escherichia coli, Enterobacter cloacae, Staphylococcus epidermidis, Proteus vulgaris and Klebsiella oxytoca in addition to bacterial antigens like hemolysin all have the ability to induce histamine release in mast cells [110–112]. However, some bacteria have been shown to have the opposite effect. For example, Helicobacter pylori, a major cause of gastritis, peptic ulcers and gastric cancer, is able to directly inhibit histamine release, perhaps contributing to the bacteria’s persistence in the gastric mucosa [113]. High doses of non-pathogenic, commensal E. coli have also been shown to be a direct inhibitor of degranulation in intestinal mast cells, suggesting a potential mechanism for this commensal bacteria’s survival in the intestine, which could potentially be applied to other commensal bacteria in the skin [114].

Despite the varying effects that different strains of bacteria can have on mast cell degranulation, it is considered a commonality that TLR-2 and TLR-4 play important roles in the mast cell’s innate immune response to most bacteria. More specifically, these receptors are required for mast cell release of cytokines and chemokines. For example, upon stimulation with LPS derived from E. coli, TLR-4 was shown to be necessary for release of the common inflammatory cytokines TNF-α, IL-1β, IL-6, and IL-13 from BMMCs [115]. Furthermore, mast cell-deficient mice that were reconstituted with TLR-4-mutated BMMCs showed defective neutrophil recruitment and production of inflammatory cytokines in the peritoneum, demonstrating the key role the TLR-4 receptor plays in mediating recruitment of other immune cells [115]. While LPS stimulation is mediated primarily through TLR-4, PGN from S. aureus stimulates mast cells in a TLR-2 dependent manner to produce TNF-α, IL-4, IL-5, IL-6, and IL-13 but not IL-1β. Additionally, intradermal injection of PGN promoted mast cell-mediated vasodilation and inflammation via TLR-2 [116]. Further evidence exists suggesting that skin-derived mast cells produce the pro-inflammatory cytokines TNF-α and IL-6, but not degranulation, in response to poly (I:C) stimulation via TLR-3, TLR-7 and TLR-9 [117]. Clearly, the mast cell response and production of cytokines depends greatly on the bacterial stimulus and TLR-mediated signaling.

In addition to cytokine production and release, mast cells are capable of releasing antimicrobial peptides, particularly cathelicidin, that can evoke direct bactericidal and anti-viral effects. Cathelicidin antimicrobial peptide was originally thought to be found only in neutrophils, but it has been identified in other cell types, including macrophages and epithelial cells in response to various microbes and 1,25-vitamin D [118]. Initial studies show that LL-37, the human cathelicidin peptide, is able to bind to high and low affinity receptors on the surface of mast cells. This binding cannot only induce degranulation but also mast cell chemotaxis, suggesting that mast cell recruitment to the site of infection is critical for innate immune responses [119]. However, more recent research shows that LL-37 is also synthesized by cultured murine and human mast cells and is necessary for efficient bacterial killing [120]. Furthermore, using mast cell- and cathelicidin-deficient mouse models, it has been demonstrated that mast cells help protect against invasive group A Streptococcus infection in the skin by producing cathelicidin. The LL-37 purified from mast cells has been identified as a unique 28-aa peptide by using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) analysis [121].

In conclusion, conserved bacterial elements not only have both stimulatory and inhibitory effects on mast cell degranulation, but they can also elicit recruitment of other immune cells via cytokine release through TLR signaling [122]. The mast cell’s ability to directly clear bacteria through cathelicidin release also demonstrates this cells critical role in the innate immune response.

Mast Cell Response to Viruses

While extensive research has been conducted on the mast cell’s role in bacterial infection, less has been studied about the mast cell response to viruses. Early work in this field shows that mast cells can be activated to both proliferate and release their granular contents [123, 124]. For example, Sendai virus can generate permeability lesions in the membranes of rat mast cells and induces release of histamine and various proteases by normal exocytotic mechanisms. [125] Although most research on mast cell interaction with viruses has been modeled in the airway connective tissue and epithelium, it is important to note that certain respiratory viruses, like respiratory syncytial virus (RSV) can induce degranulation as well as an increase in TNF-α production. However, this only occurs in co-culture with RSV-infected airway epithelial cells. Incubation of mast cells with medium from the RSV-infected epithelial cells fails to induce degranulation, suggesting a potential mechanism for crosstalk between infected cells and neighboring mast cells [126].

A similar occurrence has been noted between skin mast cells and epidermal keratinocytes infected with herpes simplex virus (HSV). Studies in mast cell-deficient mice showed increased severity of disease after transdermal injection with HSV2, and reconstitution of these mice with BMMCs reversed the high severity and mortality associated with HSV2 infection. Furthermore, TNF-α and IL-6 production in mast cells was stimulated by IL-33 derived from HSV2-infected keratinocytes [127]. Additionally, Wang et al. showed mast cell degranulation in the context of viral infection is dependent on the presence of a viral envelope, as found in vaccinia virus (VV). Lipid fusion of the VV envelope with the mast cell membrane was shown to be sufficient to induce cathelicidin release and TNF-α release, resulting in the recruitment of neutrophils and antimicrobial activity [128]. This further suggests a key role that mast cells may play in triggering inflammation after communication with virally infected cells.

While it is clear that mast cells assist in immune defense to help neighboring tissues, they have also been shown to be susceptible to direct infection by human immunodeficiency virus type 1 (HIV-1) due to their surface expression of CD4 and chemokine receptors CCR3, CCR5, and CXCR4 [129]. HIV-1 glycoprotein, gp120, can act as a viral superantigen inducing cytokine release from infected mast cells. The subsequent trafficking of infected mast cells to various tissues in combination with the mast cell’s long life span may contribute to the widespread and persistent viral reservoir in AIDS patients [130, 131]. However, some studies show immunohistochemical evidence of no active HIV replication in tissue mast cells, challenging the hypothesis of the mast cell HIV reservoir [132]. Additional research is warranted to elucidate the mast cell’s exact role in combating HIV.

It was previously mentioned that cathelicidin antimicrobial peptide from mast cells has the capacity to directly kill bacteria in the skin. Further research shows that release of cathelicidin by mast cells also has an effect on viral invasion by preventing vaccinia virus (VV) infection. VV is able to bind to the mast cell via its L1 viral membrane protein. Once bound, the virus is endocytosed leading to the activation of the phospholipid mediator S1P and expression of S1PR2. This S1PR2 receptor can function in an autocrine manner inducing degranulation of the mast cell. The granules released in response to VV are shown to contain cathelicidin, which is critical for maintaining low levels of infection [128]. Despite compelling evidence for the mast cell’s role in innate immunity against viral infection, the specific mechanisms of viral response in the mast cell are still under investigation.

Mast Cell Response to Fungi

Fungi are one of the four major groups of microorganisms that affect the skin. While knowledge of the mast cell’s role in defense against fungal infection remains extremely limited, some reports have highlighted the importance of the mast cell’s capacity to kill Candida albicans in vitro. Di Nardo et al. showed that when compared to wild type mice, mast cell-deficient mice exhibit persistently larger skin lesions when inoculated with C. albicans, suggesting that mast cells play a central role in protecting the skin during fungal infection. Additional in vitro studies using isolated rat peritoneal mast cells show potential for mast cell phagocytosis of opsonized C. albicans, while exerting fungicidal effects on non-opsonized fungus outside the cell membrane [133]. These fungicidal effects are carried out by mast cell degranulation, but further studies in the field are needed to further unveil interactions between mast cells and fungal microorganisms.

Mast Cell Response to Parasites

Throughout the past decades, multiple studies have been performed showing the activation of mast cells during parasitic infection. While the majority of this research has focused on peritoneal and mucosal mast cells in rodents, it is important to note the key role that the mast cell plays in regulating the inflammatory response to parasites.

For example, daily injections of phospholipid preparations from Ascaris suum or from Echinococcus granulosus cysts are able to induce blood eosinophilia and mast cell granule lysis along with mast cell hyperplasia [134]. Infection by the nematode parasite Trichinella spiralis has been shown to induce increased numbers of IgE containing mast cells in the intestinal mucosa of mice [135]. In addition to the recruitment of mast cells to the site of infection, parasites like Toxoplasma gondii have been shown to induce mast cell degranulation, followed by an observed increase in neutrophil count at the site of infection [136]. This suggests that mast cells are critically involved in parasite-mediated inflammation and recruitment of other immune cells. It has also been shown that mast cells play a role in direct elimination of the parasite. During infection with Schistosoma mansoni, a pronounced hepatic mastocytosis, or an abnormally high number of mast cells, is observed in rats. The majority of these recruited hepatic mast cells contain a highly soluble granular chymase that is released systemically into the blood during the period of parasite elimination. Thus, due to these chymases, infection is terminated in the liver before egg laying commences [137].

More relevant to the skin is the mast cell’s activity during Leishmania major infection. Cutaneous leishmaniasis is a quickly spreading, ulcerative skin disorder caused by protozoan parasitic infection [138]. While some evidence shows mast cells to be the cause of increased lesion size and intensity, this is likely due to the immediate release of mediators like beta-hexosaminidase and TNF-α upon mast cell contact with the parasite [139, 140]. Despite the mast cell’s apparent contribution to the appearance of the lesion, it has also been shown that mast cell degranulation may inhibit infection. When mast cells in mice were induced to degranulate before L. major challenge, these mice showed lower rates of infection along with high levels of IFN-γ and reduced levels of IL-4 [141].

Despite a large amount of evidence that parasites are capable of inducing mast cell activation and degranulation, it has been repeatedly shown that infection by certain parasitic roundworms leads to inhibitory signaling via TLR-4. In contrast to the binding of TLR-4 to LPS, when bound to the ES-62 protein from roundworms, a resultant inhibition of FcεRI activation occurs. Binding of ES-62 to TLR-4 on the surface of mast cells causes sequestration of PKC, which in turn inhibits downstream signaling of the high-affinity IgE receptor [142].

In summary, the presence of mast cells is strongly associated with parasitic infection, mediating inflammation and the recruitment of other immune cells. Although many mechanisms of mast cell defense against parasites remain unknown, it is now evident that mast cells can amplify protective responses against parasites in innate immunity as well as play a conflicting role in inflammation and pathology at sites of infection.

Mast Cell-Associated Skin Diseases

Hypersensitivity Reactions

Mast cells are the primary mediators of immediate anaphylactic hypersensitivity (type 1) reactions. Tissue mast cells that are sensitized with IgE antibodies bound to the high-affinity IgE receptor can also bind to antigen, inducing receptor cross-linking and subsequent mast cell activation and mediator release, as outlined in the section on mast cell activating receptors. Release of these mediators results in increased vascular permeability, edema and smooth muscle contraction. The most common manifestations include urticarial rash, erythema and pruritus, while extreme cases can lead to anaphylaxis. This type 1 hypersensitivity reaction is associated with both food and drug allergies [143].

Mast cells have also been shown to be considerably active in delayed-type hypersensitivity (type 4) reactions (DTHRs), and most relevant to the skin, contact hypersensitivity reactions. DTHRs depend on the presence of type 1 memory T cells and their ability to produce IFN-γ. Hapten binding is the initial step of these reactions. Low molecular weight contact allergens, called haptens, are able to penetrate the epidermal barrier and bind to various skin proteins [144]. During the initial sensitization phase of the reaction, the bound hapten can be recognized by Langerhans cells, leading to migration to the lymph nodes and clonal expansion of both CD4+ and CD8+ hapten-specific T cells [145, 146].

The following effector phase of the contact hypersensitivity reaction is less studied, but mast cells have been shown to play an important role. During the effector phase, Ig free light chains are produced by B lymphocytes, and they have been shown to be necessary for full development of contact hypersensitivity in mice [147]. Furthermore, these free light chains have been shown to be important in the sensitization of mast cells and for their activation when coming into contact with allergens [148]. While this evidence suggests that mast cells may be involved in the development of DTHRs, various other studies involving reconstitution of mast cell-deficient mice with mast cells cultured in vitro show that in various DTHRs, like allergic encephalitis, contact hypersensitivity and cutaneous responses to microbes, mast cells are needed for full manifestation of symptoms. [149–151] Specifically in the DTHR context, mast cells produce TNF-α for induction of dendritic cell migration, IL-3 for proliferation and activation of T cells, and they may play a role in direct antigen presentation [152]. While mast cells may not be the main mediators of contact hypersensitivity reactions, their presence proves necessary for a coordinated immune response, recruitment of other cell types, and full development of symptoms.

Rosacea

Recent studies show new evidence that demonstrates the mast cell’s central role in the pathogenesis of rosacea. With approximately 16 million Americans affected, this chronic inflammatory disease can flare up due to increased temperature, spicy foods, or various other environmental triggers. The resulting, often painful, inflammation can take weeks to subside without treatment [153]. It has also been shown that rosacea is associated with elevated levels of an aberrant form of antimicrobial peptide, cathelicidin LL-37, due to overactivity of kallikrein-related peptidases (KLKs), which can generate LL-37 from its precursor peptide [154]. Studies in mast cell-deficient Kit ^W-sh/W-shmouse strains show that upon intradermal injection with LL-37, Kit ^W-sh/W-shmice fail to develop rosacea-related inflammation, whereas wild type mice exhibit the phenotypically characteristic inflammation of rosacea. Furthermore, it is now known that upon stimulation with LL-37, mast cells respond by releasing metalloproteinase 9 (MMP9), the enzyme responsible for activating KLKs, and proinflammatory IL-6, suggesting a critical role for the mast cell in rosacea inflammation. Ultimately, treatment in human rosacea subjects with 4 % cromolyn sodium, a known mast cell stabilizer, significantly decreased rosacea-associated inflammation, and in mouse models, it has also been shown to reduce levels of MMP9 [155]. This data shows that the presence of mast cells in the skin is necessary for development of the rosacea phenotype. Furthermore, mast cell stabilizers may prove to be the treatment of choice for rosacea patients.

Mastocytosis

Mastocytosis is characterized by abnormally high numbers of mast cells localizing in one or multiple tissues. This rare disorder is accompanied by chronic or episodic degranulation and release of mast cell mediators into the tissue. Cutaneous mastocytosis involves only the skin, while systemic mastocytosis affects internal organs with or without involvement of the skin [156]. While pathogenesis of the disease is not fully understood, mastocytosis has been shown to be associated with mutations in the gene encoding the c-kit receptor or with increased expression of SCF. The most common c-kit mutation is a D816V mutation of exon 17. This activating mutation induces SCF-independent activation of c-kit, leading to both clonal expansion and apoptotic defects in the mast cells [157, 158]. Increased levels of free SCF have also been identified in the dermis and extracellular spaces of the epidermis in patients with cutaneous mastocytosis; however, this elevation of SCF may be variable in different patients [159]. Patients with cutaneous mastocytosis typically present with a yellow-tan to reddish-brown maculopapular skin lesion known as urticaria pigmentosa, a fixed accumulation of mast cells in the skin, which may also manifest itself in a plaque or nodular form. More rare presentation involves diffuse mastocytosis, which may present in a bullous form and involve the whole skin, or mastocytomas, which usually present in childhood [160, 161]. Mediator release from the mast cells has a presentation that mimics allergic reactions with the typical symptoms of pruritus and flushing [162]. While children with cutaneous mastocytosis typically clear the disease by adolescence, avoidance of triggers that lead to mediator release like temperature change, friction, and physical exertion is important for the disease to remain asymptomatic. Antihistamines and PUVA have proven to be effective therapies; however, treatment with corticosteroids has not been shown to diminish symptoms of the disease [162].

Conclusions

Mast cells prove to be an extremely important part of the innate immune system in the skin. From their ability to initiate recruitment of critical immune cells to their central role in the allergic response, mast cells truly are the sentinels and directors of many coordinated immune responses. Furthermore, key therapies like imatinib and similar biologics act by blocking the effect of c-kit, and thus mast cell development, in the context of various types of malignancies. The development of medications that can directly target the mast cells further demonstrates this cell’s key role in the innate immune response. While much has been discovered about the mast cell’s role in the skin, there remains great potential for further study and development of therapeutics centered on mast cell function.

Questions

1.
Which cytokine is critical for proper mast cell differentiation?
A.

IL1

B.

SCF

C.

IL6

D.

TNFα
2.
Binding of which of the following molecules to the mast cell surface can induce degranulation?
A.

IgE

B.

IgG

C.

Complement peptides

D.

All of the above
3.
Release of which of the following mast cell mediators has been shown to aid defense against microbes?
A.

Cathelicidin

B.

Histamine

C.

Leukotrienes

D.

Prostaglandins

Answers

1.

B
2.

D
3.

A

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10.

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11.

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12.

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13.

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14.

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