Activated brain mast cells release histamine that can cause phenotypic changes and activation of microglial cells [101
]. Exogenously added histamine triggered activation of cultured primary cortical microglia, murine N9 microglia and hippocampal organotypic slice cultures to secrete TNF-α and IL-6 and RNOS [102
]. Although histamine stimulated microglial cell motility in control microglia, histamine inhibited microglial migration and IL-1β release in LPS-stimulated microglia, suggesting a dual role of histamine in modulating microglia-induced inflammatory responses [102
]. Histamine exerts its functions by signaling through four types of G-protein coupled receptors, namely Histamine 1 receptor (H1R), H2R, H3R, and H4R, which are expressed on innate immune cells, neurons and endothelial cells [105
]. In these cultures, the anti-inflammatory effects of histamine were mediated by activation of H4R which involved α5β1 integrin, p38 and protein kinase B (AKT) signaling to restrain exacerbated microglial responses in neuroinflammation [102
]. All four types of histamine receptors are expressed by microglial cells and can modulate microglia-mediated neuroinflammation [104
]. Rocha SM et. al., directly injected histamine into the substantia nigra of mice and studied its effects on microglial activity and dopaminergic neuron survival [106
]. In accordance with other studies, histamine induced microglial activation and dopaminergic neuronal toxicity via H1R activation, probably through NADPH oxidase dependent oxidative stress signaling pathways and microglia phagocytosis. Altogether, these studies show that histamine per se triggers a pro-inflammatory response and under inflammatory conditions, histamine activates an anti-inflammatory response, dampens microglial-induced inflammation and is associated with neuroprotection.
Similar to peripheral mast cells, brain mast cells are also known to secrete proteases, such as tryptase. Exposure of primary microglia to mast cell-derived tryptase stimulated microglia to subsequently secrete TNF-α and IL-6 and RNOS. These effects were mediated by protease-activated receptor-2 (PAR-2) signaling via activation of mitogen-activated protein (MAP) kinase (Erk and p38) and NF-kappa B (NF-kB) pathways [68
]. Furthermore, PAR-2 activation induces the expression of ATP-sensitive ionotropic P2X4 receptors on microglia and exposure to ATP leads to secretion of BDNF, a potent trophic factor [107
]. The presence of functional P2X4 receptors are also expressed by human mast cell lines [108
]. Since mast cells play a pivotal role in neuroinflammation, it is important to determine the exact molecular mechanisms employed by activated mast cells and microglia and their role in disease progression.
Mast cell-derived pro-inflammatory cytokines such as CCL2, TNF-α and IL1β can also influence microglia activation. To recapitulate in vitro mast cell-glia-neuron crosstalk during neuroinflammation, mast cells were cocultured with mixed cultures of neuron and glia or enriched cultures of neurons or astroglia and challenged with MPP+
or GMF or mast cell proteases [109
]. Mast cells cocultured with glia had increased production of CCL2 and IL-33, highlighting the importance of mast cell-glia coupling and their role in neuroinflammation [110
]. Increased CCL2 levels have been demonstrated in AD patients which is associated with accelerated cognitive decline and AD progression [111
]. CCL2 expression altered β-amyloid phagocytosis, supporting the notion that microglial phagocytosis could be regulated by mast cells [112
]. Although there is no direct evidence of CCL2 expression in the brain of PD patients, increased serum levels of CCL2 have been reported [113
]. Recently, two polymorphisms have been reported in the promoter region of CCL2 which are associated with an increased risk of PD [114
]. The exact role of CCL2-CCR2 axis in regulating mast cells and microglia in neurodegenerative diseases is still not well understood.
Mast cell degranulation has been shown to activate microglia. Stereotaxic injection of a mast cell degranulation compound 48/80 (C48/80) and activator of the mas-related G protein-coupled receptor Mrgpr [115
] in the hypothalamus of rats induced mast cell degranulation, production of pro-inflammatory cytokines and microglia activation [103
]. These effects were mediated by activation of MAP kinase and AKT pathways and an increased protein expression of H1R, H4R, PAR-2 and TLR4 on microglial cells. Treatment with a mast cell stabilizer disodium cromoglycate (cromolyn), inhibited microglial activation and downstream signaling, suggesting mast cell involvement. Most importantly, C48/80 had no effect on microglial activation in mast cell-deficient Kitw-sh/w-sh
mice. These data support the notion that stabilization of brain mast cells during neuroinflammation could be a new therapeutic strategy to restrain microglial hyperactivity. Tranilast has been used to inactivate the NLR family pyrin domain containing 3 (NLRP3) inflammasome, yet is also used as an anti-allergy medication as a “mast cell stabilizer” [116
] suggesting that its effect in the brain may also modulate mast cell functions via the inflammasome.
NLRP3 Inflammasome: Common Sensor in Microglia and Mast Cells
In the sections above, the individual signals that activate either mast cells or microglia were described. However, it is possible that mast cells and microglia can be activated by the same signal and function concurrently to orchestrate inflammation in the brain. Both mast cells and microglia contain key signaling hubs in the cytoplasm, called inflammasomes [117
]. The main function of inflammasomes is to detect and eliminate a variety of stimuli including PAMPs, DAMPs, or NAMPs by subsequent proteolytic cleavage of pro-IL-1β and pro-IL-18 to generate their bioactive forms and secretion of IL-1β and IL-18, respectively (Figure 3
B). The NLRP3 is the best-studied inflammasome which consists of three major components: (i) the NLR protein as sensor, (ii) the Apoptosis-associated Speck-like protein containing a Caspase-activating and recruitment domain (ASC) as adaptor, and (iii) the protease caspase-1 as effector. The adaptor, ASC acts as a direct bridge between the sensor and the downstream effector caspase-1 and plays a central role in the assembly of the NLRP3 inflammasome machinery [119
The activation and assembly of NLRP3 inflammasome is under tight regulation and occurs by a two-signal process, consisting of a priming signal and an activation signal (Figure 3
]. The priming signal is delivered by stimulation of TLRs on host cells and is associated with NFĸB-dependent transcription of pro-IL-18, pro-IL-1β and NLRP3. The second signal is elicited by multiple stimuli, including NAMPs released from damaged or degenerating neurons, abnormally aggregated β-amyloid and α-synuclein, ionic flux (e.g., efflux of K+
ions or influx of Ca2+
ions), high glucose, exogenous ATP, reactive oxygen species from mitochondrial damage or lysosomal proteases [17
]. Upon activation, NLRP3 inflammasome assembles by recruiting ASC which rapidly forms dimers and polymerizes to assemble into large filamentous structures known as ASC-specks, a macromolecular form of the inflammasome which activates caspase-1 [122
]. The ASC-specks are released into the extracellular space, where they act as signaling platforms to activate free precursor IL-1β or are phagocytosed by myeloid cells leading to further activation of caspase-1 and release of IL-1β or IL-18 [123
]. The overactivated NLRP3 inflammasome can constitutively release cytokines resulting in pyroptosis, an inflammatory form of cell death, which can exacerbate chronic inflammatory responses.
Increasing number of preclinical and human clinical studies have shown that the NLRP3 inflammasome can be activated by several NAMPs in chronic neurodegenerative diseases, including AD and PD [22
]. The binding of β-amyloid to TLR2 on the microglia triggers NLRP3 activation, thereby releasing IL-1β. Accordingly, components of NLRP3 inflammasome such as increased levels of cleaved caspase-1 and IL-1β were detected in the serum of early AD patients or patients with mild cognitive impairment (MCI) as compared to non-demented and age-matched controls [125
]. More importantly, ASC bound β-amyloid was also found in the post-mortem AD brain tissue [123
]. These findings suggest that NLRP3 activation represents an early biomarker of AD. A large number of studies conducted in the APP/PS1 transgenic mice, which recapitulates most features of AD pathology, has consistently concluded that β-amyloid deposition, microglial NLRP3 activation and IL-1β release is an early feature of AD progression [124
]. The evidence for ASC-speck release by microglial pyroptosis and its spread by a ‘prion-like’ mechanism was obtained in the APP/PS1 mouse brains. ASC-specks released by microglia were able to cross-seed β-amyloid in the surrounding microglia, in turn, leading to propagation of neuroinflammatory responses in AD progression [122
]. Interestingly, APP/PS1 mice that lacked components of the NLRP3 apparatus showed a reduction in β-amyloid burden, possibly due to increased microglial phagocytosis and reduction in cognitive decline [127
]. Moreover, treatment with MCC950, a specific inhibitor of the NLRP3 that inhibits NLRP3-induced ASC oligomerization, promoted non-phlogistic clearance of β-amyloid and ameliorated cognitive impairment [128
]. These studies revealed NLRP3 as a potential therapeutic target for treating early-stage AD.
More recently, the NLRP3 inflammasome has also been shown to play an important role in PD pathogenesis. Hallmarks of inflammasome activation, i.e., cleaved caspase-1, IL-1β NLRP3 and ASC-specks were found in the postmortem PD brain tissues and in plasma of PD patients compared to age-matched controls, suggesting systemic increase in inflammasome activation [129
]. Furthermore, systemic increases in NLRP3 activation shows a positive correlation with motor function decline and PD progression [132
]. A strong association between α-synuclein levels and microglial NLRP3 inflammasome activation accompanied with dopaminergic neuron toxicity has been shown in several animal models of PD, including the MPTP mouse model of PD [129
]. Oral administration of MCC950 reduced pro-inflammatory cytokines and attenuated dopaminergic neuron loss [129
]. Furthermore, NLRP3-deficiency abolished MPTP-induced microglial activation, caspase-1 and subsequent IL-1β release, suggesting that NLRP3 pathway plays a pivotal role in MPTP-induced neurodegeneration [133
]. These results indicate that therapeutic targeting of the NLRP3 pathway has the potential to slow down or halt PD progression.
Although, to date, no reports have investigated the expression of the NLRP3 inflammasome in brain mast cells, peripheral tissue mast cells from cryopyrin-associated periodic syndrome (CAPS) patients were, for the first time, shown to express functional inflammasomes and secrete IL-1β [134
]. Notably, mast cells from the skin of CAPS patients expressing the disease-associated NLRP3 mutations constitutively produced IL-1β and mediated histamine-independent urticarial rash. Furthermore, these patients responded to IL-1 receptor antagonist therapy rather than antihistamines, suggesting that urticaria in CAPS patients is mediated by IL-1β [134
]. Seminal work from Melissa Brown’s research team has also shown that mast cell inflammasome is a critical mediator of inflammation in the meninges and regulates disease severity in a rodent model of multiple sclerosis [135
]. The meningeal mast cells, which lie outside the BBB, release IL-1β and licenses T-cell pathogenicity [135
]. These studies support the notion that mast cells are principal drivers of inflammatory responses. Although the direct role of NLRP3 inflammasomes in brain mast cell activation and function in AD and PD is lacking, it is not unreasonable to postulate that brain mast cells may play an important role in neuroinflammation and neurodegeneration.