Primary Mast Cells Research Articles

Targeting of the IL-5 pathway in severe asthma reduces mast cell progenitors

BackgroundTherapies targeting interleukin-5 (IL-5) or its receptor (IL-5Rα) are currently used to treat patients with severe eosinophilic asthma. ObjectiveTo investigate the impact of anti-IL-5 and anti-IL-5Rα biological therapies on mast cells (MCs) and their progenitors. MethodsSurface IL-5Rα expression was investigated on MCs and their progenitors in mouse lungs and bone marrow, and in human lungs and blood. Isolated human MC progenitors cultured in the presence or absence of IL-5 were analyzed in vitro. Circulating MC progenitors were quantified in patients with severe asthma, before and after anti-IL-5 (mepolizumab) or anti-IL-5Rα (benralizumab) therapy. Gene expression analysis of MC progenitors was performed before and after anti-IL-5Rα therapy. ResultsApproximately 50% of the human primary lung MCs, and 30% of the human MC progenitors from individuals with allergic asthma expressed IL-5Rα. In patients with mild-to-moderate allergic asthma and mice with acute allergic airway inflammation, the fraction of IL-5Rα + MC progenitors was elevated. Additionally, IL-5 promoted the proliferation and/or survival of isolated human MC progenitors. Further, patients with severe asthma from two independent cohorts demonstrated a reduction of blood MC progenitors after anti-IL-5 or anti-IL-5Rα treatment. This was associated with improved asthma control, as well as a decline in both blood eosinophils and Th2 cells. Finally, the blood MC progenitors remaining after anti-IL-5Rα (benralizumab) treatment exhibited a downregulated expression of genes involved in growth and proliferation. ConclusionThis study introduces the possibility that the clinical effects of targeting IL-5/IL-5Rα in severe asthma also may involve reduction of MC populations.

Regulation of BCR-dependent germinal center B-cell formation by HGAL and insight into its emerging myeloid ortholog, C1ORF150.

The specificity of cytokine and immunoreceptor signaling frequently depends upon receptor recruitment of select adaptor proteins and specifically engaged effectors. This review focuses on the orthologous adaptor proteins, HGAL and C1ORF150, and aims to provide insight into their respective modulation of lymphoid and myeloid cell signaling, formation, and function. HGAL acts predominantly within germinal center B cells as an important BCR signal transducer. Effects on BCR signalosome assembly involve HGAL's localization to the plasma membrane via its lipidation, initial interactions with SYK, the pY-phosphorylation of HGAL including its recruitment of GRB2, and HGAL engagement of PDZ-RhoGEF and RhoA signaling. At ligated BCRs, this includes HGAL(-GRB2) stimulation of SYK kinase, attenuation of calcium flux-dependent and NF-κB expression, promotion of cSMAC formation, and cytoskeletal remodeling associated with HGAL-attenuated cell migration. HGAL and partnered effectors also impact on DLBCL pathogenesis, and studies are summarized on HGAL's actions (using DLBCL and Burkitt lymphoma B cells) including cell migration effects, HGAL modulation of cytoskeletal components, and insightful HGAL transgenic mouse and xenograft models. For C1ORF150, its HGAL-homologous subdomains are considered, together with studies that demonstrate C1OR150's FcϵRI- and KIT-mediated expression and phosphorylation in primary human mast cells. Intriguingly, recent GWAS studies have identified a C1ORF150 in-frame splice variant that is strongly associated with urticaria. Candidate mechanisms via which the encoded "C1ORF150-Δexon2" isoform affects mast cell degranulation are considered, including FcϵR1 and/or KIT receptor connections, and candidate "myristoylation switch" mechanisms.

Combinatorial Genomic Biomarkers Associated with High Response in IgE-Dependent Degranulation in Human Mast Cells.

Mast cells are the major effector cells that mediate IgE-dependent allergic reactions. We sought to use integrated network analysis to identify genomic biomarkers associated with high response in IgE-mediated activation of primary human mast cells. Primary human mast cell cultures derived from 262 normal donors were categorized into High, Average and Low responder groups according to their activation response profiles. Transcriptome analysis was used to identify genes that were differentially expressed in different responder cultures in their baseline conditions, and the data were analyzed by constructing a personalized perturbed profile (PEEP). For upregulated genes, the construction of PEEP for each individual sample of all three responder groups revealed that High responders exhibited a higher percentage of "perturbed" samples whose PEEP values lay outside the normal range of expression. Moreover, the integration of PEEP of four selected upregulated genes into distinct sets of combinatorial profiles demonstrated that the specific pattern of upregulated expression of these four genes, in a tandem combination, was observed exclusively among the High responders. In conclusion, this combinatorial approach was useful in identifying a set of genomic biomarkers that are associated with high degranulation response in human mast cell cultures derived from the blood of a cohort of normal donors.

Transcriptional profiling of primary human-skin derived mast cell activation by various MRGPRX2 agonists and inhibition with EVO756

Abstract Mas-related G-protein coupled receptor X2 (MRGPRX2) is a GPCR expressed predominantly by mast cells and has been implicated to play an important role in diseases such as chronic spontaneous urticaria, atopic dermatitis, and asthma. Cationic ligands, either endogenous or exogenous, can activate the receptor and induce IgE-independent mast cell activation, resulting in allergic responses and non-histaminergic itch. We have previously shown that EVO756, a novel small molecule antagonist of MRGPRX2, demonstrates potent, concentration dependent inhibition of MRGPRX2 mediated activation of transfected CHO-cells, ROSA and LAD2 mast cell lines, and primary human skin-derived mast cells. Furthermore, EVO756 does not affect mast cell viability. Here, we describe, for the first time, the transcriptional landscape of primary human skin-derived mast cells cultured in the presence of several MRGPRX2 agonists and analyze how the mast cell phenotype is altered in response to MRGPRX2 inhibition by EVO756.

Sialic acids from the Group B Streptococcus (GBS) capsule dampen mast cell activation through Siglec-9

Abstract GBS is a leading cause of bacterial infection in newborns, adult diabetics, pregnant women, and the elderly. A key virulence factor of GBS is its surface capsular polysaccharide, which displays terminal sialic acids that suppress host innate immune responses by engaging host inhibitory receptors like Sialic acid-binding immunoglobin-like lectin- 9 (Siglec-9). We reported that human mast cells (MC) express Siglec-9 and that engaging it with native ligands or antibodies results in reduced mast cell activation in vitro. We hypothesized that capsular sialic acids engage Siglec-9 to impair MC ability to initiate the host response. Mice expressing human Siglec -7 and -9 were infected with WT GBS or a sialic acid deficient strain, ΔcpsK, and the ΔcpsK infected mice showed reduced bacterial burden and less dissemination. Using bone marrow-cultured MC from a transgenic mouse expressing Siglec-9, we confirmed that GBS interacts with MC Siglec-9, which inhibited the release of the pro-inflammatory cytokine IL-6. Challenge of primary human MC with WT, ΔcpsK, or ΔneuA, another sialic acid deficient GBS strain, showed that MC interact more with ΔcpsK and ΔneuA GBS, and that MC kill sialic acid deficient GBS. MC challenged with ΔcpsK or ΔneuA GBS also produce more IL-8 and undergo cell death more than WT infected MC. Together, this data shows that sialylated GBS capsule engages MC Siglec-9 to dampen activation, which reduces the immune response and benefits the pathogen.

Mast cell versus basophil activation test in allergy: Current status.

In the past two decades, we witnessed the evolution of the basophil activation test (BAT) from mainly research applications to a potential complementary diagnostic tool to document IgE-dependent allergies. However, BAT presents some technical weaknesses. Around 10%-15% of tested patients are non-responders, BAT can be negative immediately post-reaction and the use of fresh basophils, ideally analysed within 4 h of collection, restricts the number of tests that can be performed per sample. The need for fresh basophils is especially limiting when conducting batch analyses and interlaboratory comparisons to harmonize BAT methodology. These limitations significantly hinder the wider application of BAT and urge the development of alternative testing, such as the mast cell activation test (MAT). The essential difference between BAT and MAT is the heterogeneity of the starting material used to perform the assays. Mast cells are tissue-resident, so cannot be easily accessed. Current alternative sources for functional studies are generating primary human mast cells, differentiated from donor progenitor cells, or using immortalized mast cell lines. Hence, the methodological approaches for MAT are not only vastly different from BAT, but also different among MAT protocols. This review summarizes the advantages and disadvantages of BAT and MAT assays, dedicating special attention to elucidating the key differences between the cellular sources used and provides an overview of studies hitherto performed comparing BAT and MAT in the diagnosis of IgE-mediated food and drug allergies.

A highly selective KIT inhibitor MOD000001 suppresses IgE-mediated mast cell activation

BackgroundThe KIT receptor tyrosine kinase and its ligand, stem cell factor (SCF) control proliferation and survival of mast cells. Thus, targeting KIT signaling may show promise for the treatment of allergic diseases involving mast cells. Recently, we discovered a new compound MOD000001 as a potential small-molecule KIT kinase inhibitor by using an in silico approach. ObjectiveThis study determined whether MOD000001 is highly-selective to KIT, inhibits KIT signaling in mast cells, and affects IgE-mediated mast cell activation. MethodsThe interaction of MOD000001 with 468 human kinases and its inhibitory activity against KIT were profiled and evaluated by using KinomeScan and cell-free kinase assays, respectively. The effects of MOD000001 on SCF-dependent signaling were examined by using primary mouse and human mast cells. The effects of MOD000001 on SCF-induced degranulation and passive cutaneous anaphylactic (PCA) reaction were examined in mice. ResultsMOD000001 interacted with KIT and inhibited KIT kinase activity with high selectivity. MOD000001 suppressed SCF-induced KIT signaling in mouse and human mast cells and in mice. PCA reaction was suppressed in mice treated with MOD000001 both for a short-term (1 week) and a long-term (7 weeks). Mice treated with MOD000001 for a long-term, but not for a short-term, showed skin mast cell reduction. ConclusionsMOD000001 is a highly selective KIT inhibitor that can suppress IgE-mediated mast cell activation in vivo. MOD000001 may do so by reducing tissue mast cell numbers or by other unknown mechanisms. The findings suggest potential benefits of MOD000001 for allergic diseases involving IgE-mediated mast cell activation.

Coronavirus Receptor Expression Profiles in Human Mast Cells, Basophils, and Eosinophils.

A major problem in SARS-CoV-2-infected patients is the massive tissue inflammation in certain target organs, including the lungs. Mast cells (MC), basophils (BA), and eosinophils (EO) are key effector cells in inflammatory processes. These cells have recently been implicated in the pathogenesis of SARS-CoV-2 infections. We explored coronavirus receptor (CoV-R) expression profiles in primary human MC, BA, and EO, and in related cell lines (HMC-1, ROSA, MCPV-1, KU812, and EOL-1). As determined using flow cytometry, primary MC, BA, and EO, and their corresponding cell lines, displayed the CoV-R CD13 and CD147. Primary skin MC and BA, as well as EOL-1 cells, also displayed CD26, whereas primary EO and the MC and BA cell lines failed to express CD26. As assessed using qPCR, most cell lines expressed transcripts for CD13, CD147, and ABL2, whereas ACE2 mRNA was not detectable, and CD26 mRNA was only identified in EOL-1 cells. We also screened for drug effects on CoV-R expression. However, dexamethasone, vitamin D, and hydroxychloroquine did not exert substantial effects on the expression of CD13, CD26, or CD147 in the cells. Together, MC, BA, and EO express distinct CoV-R profiles. Whether these receptors mediate virus-cell interactions and thereby virus-induced inflammation remains unknown at present.

Cytotoxicity assessment of exfoliated MoS2 using primary human mast cells and the progenitor cell-derived mast cell line LAD2.

Molybdenum disulfide is an emerging 2D material with several potential applications in medicine. Therefore, it is crucial to ascertain its biocompatibility. Mast cells are immune cells that are found in many organs and tissues in contact with the extracellular environment, and can be cultured from progenitor cells present in the bone marrow. Given the long period required for differentiation and proliferation of primary mast cells, human mast cell lines have emerged as a tractable model for biological and toxicological studies. Here, we compare two types of industrial MoS2 using CD34+-derived primary human mast cells and the LAD2 cell line. Minimal effects were observed on early-stage activation endpoints such as β-hexosaminidase release and expression of surface markers of mast cell activation. Transmission electron microscopy revealed limited uptake of the tested materials. Overall, MoS2 was found to be biocompatible, and the LAD2 cell line was validated as a useful in vitro model of mast cells.

Cannabidiol Inhibits IgE-Mediated Mast Cell Degranulation and Anaphylaxis in Mice.

Cannabidiol (CBD), the most abundant non-psychoactive constituent of the plant Cannabis sativa, is known to possess immune modulatory properties. This study investigates the effects of CBD on mast cell degranulation in human and mouse primary mast cells and passive cutaneous anaphylaxis in mice. Mouse bone marrow-derived mast cells and human cord-blood derived mast cellsare generated. CBD suppressed antigen-stimulated mast cell degranulation in a concentration-dependent manner. Mechanistically, CBD inhibited both the phosphorylation of FcεRI downstream signaling molecules and calcium mobilization in mast cells, while exerting no effect on FcεRI expression and IgE binding to FcεRI. These suppressive effectsare preserved in the mast cells thatare depleted of type 1 (CB1) and type 2 (CB2) cannabinoid receptors, as well as in the presence of CB1 agonist, CB2 agonist, CB1 inverse agonist, and CB2 inverse agonist. CBD also inhibited the development of mast cells in a long-term culture. The intraperitoneal administration of CBD suppressed passive cutaneous anaphylaxis in mice as evidenced by a reduction in ear swelling and decrease in the number of degranulated mast cells. Based on these results, the administration of CBD is a new therapeutic intervention in mast cell-associated anaphylactic diseases.

SCF and IL-33 regulate mouse mast cell phenotypic and functional plasticity supporting a pro-inflammatory microenvironment

Mast cells (MCs) are multifaceted innate immune cells often present in the tumor microenvironment (TME). Several recent findings support their contribution to the transition from chronic inflammation to cancer. However, MC-derived mediators can either favor tumor progression, inducing the spread of the tumor, or exert anti-tumorigenic functions, limiting tumor growth. This apparent controversial role likely depends on the plastic nature of MCs that under different microenvironmental stimuli can rapidly change their phenotype and functions. Thus, the exact effect of unique MC subset(s) during tumor progression is far from being understood. Using a murine model of colitis-associated colorectal cancer, we initially characterized the MC population within the TME and in non-lesional colonic areas, by multicolor flow cytometry and confocal microscopy. Our results demonstrated that tumor-associated MCs harbor a main connective tissue phenotype and release high amounts of Interleukin (IL)-6 and Tumor Necrosis Factor (TNF)-α. This MC phenotype correlates with the presence of high levels of Stem Cell Factor (SCF) and IL-33 inside the tumor. Thus, we investigated the effect of SCF and IL-33 on primary MC cultures and underscored their ability to shape MC phenotype eliciting the production of pro-inflammatory cytokines. Our findings support the conclusion that during colonic transformation a sustained stimulation by SCF and IL-33 promotes the accumulation of a prevalent connective tissue-like MC subset that through the secretion of IL-6 and TNF-α maintains a pro-inflammatory microenvironment.

A Technique to Isolate Primary Mast Cells from a Murine Peritoneal Cavity

A Technique to Isolate Primary Mast Cells from a Murine Peritoneal Cavity

A Technique to Isolate Primary Mast Cells from a Murine Peritoneal Cavity

A Technique to Isolate Primary Mast Cells from a Murine Peritoneal Cavity

The mRNA methyltransferase Mettl3 modulates cytokine mRNA stability and limits functional responses in mast cells

Mast cells are central players in allergy and asthma, and their dysregulated responses lead to reduced quality of life and life-threatening conditions such as anaphylaxis. The RNA modification N6-methyladenosine (m6A) has a prominent impact on immune cell functions, but its role in mast cells remains unexplored. Here, by optimizing tools to genetically manipulate primary mast cells, we reveal that the m6A mRNA methyltransferase complex modulates mast cell proliferation and survival. Depletion of the catalytic component Mettl3 exacerbates effector functions in response to IgE and antigen complexes, both in vitro and in vivo. Mechanistically, deletion of Mettl3 or Mettl14, another component of the methyltransferase complex, lead to the enhanced expression of inflammatory cytokines. By focusing on one of the most affected mRNAs, namely the one encoding the cytokine IL-13, we find that it is methylated in activated mast cells, and that Mettl3 affects its transcript stability in an enzymatic activity-dependent manner, requiring consensus m6A sites in the Il13 3’-untranslated region. Overall, we reveal that the m6A machinery is essential in mast cells to sustain growth and to restrain inflammatory responses.

Simultaneous reduction of all ORMDL proteins decreases the threshold of mast cell activation

In mammals, the ORMDL family of evolutionarily conserved sphingolipid regulators consists of three highly homologous members, ORMDL1, ORMDL2 and ORMDL3. ORMDL3 gene has been associated with childhood-onset asthma and other inflammatory diseases in which mast cells play an important role. We previously described increased IgE-mediated activation of mast cells with simultaneous deletions of ORMDL2 and ORMDL3 proteins. In this study, we prepared mice with Ormdl1 knockout and thereafter, produced primary mast cells with reduced expression of one, two or all three ORMDL proteins. The lone deletion of ORMDL1, or in combination with ORMDL2, had no effect on sphingolipid metabolism nor IgE-antigen dependent responses in mast cells. Double ORMDL1 and ORMDL3 knockout mast cells displayed enhanced IgE-mediated calcium responses and cytokine production. Silencing of ORMDL3 in mast cells after maturation increased their sensitivity to antigen. Mast cells with reduced levels of all three ORMDL proteins demonstrated pro-inflammatory responses even in the absence of antigen activation. Overall, our results show that reduced levels of ORMDL proteins shift mast cells towards a pro-inflammatory phenotype, which is predominantly dependent on the levels of ORMDL3 expression.

NS5806 reduces carrageenan-evoked inflammation by suppressing extracellular signal-regulated kinase activation in primary sensory neurons and immune cells.

The compound NS5806 attenuates neuropathic pain via inhibiting extracellular signal-regulated kinase (ERK) activation in neuronal somata located at the dorsal root ganglion (DRG) and superficial spinal dorsal horn. NS5806 also reduces the expansion of DRG macrophages and spinal microglia several days after peripheral nerve injury, implying an anti-inflammatory effect. To test whether NS5806 inhibits inflammation, as a model we intraplantarly injected carrageenan into a hind paw of the rat. To examine whether NS5806 reduces carrageenan-evoked mechanical allodynia, thermal hyperalgesia, and edema, as well as ERK activation in the nerve fibres, mast cells, and macrophages in the hind paw skin, we used behavioural, immunohistochemical, and cytological methods. NS5806 did not impair motor function, affect basal nociception, or cause edema in naive rats. Six hours after carrageenan injection, mechanical allodynia, thermal hyperalgesia, and edema appeared in the rat's ipsilateral hind paw, and all were reduced by intraplantar co-injection of NS5806. NS5806 suppressed carrageenan-evoked ERK activation in the peripheral axons and somata of L4 DRG neurons, as well as mast cells and macrophages in the paw skin. NS5806 also reduced carrageenan-evoked mast cell degranulation and macrophage proliferation. NS5806 and the ERK pathway inhibitor PD98059 had a similar effect in inhibiting the proliferation of cultured RAW264.7 macrophages. Furthermore, all the in vivo anti-inflammatory effects of NS5806 were similar to those of PD98059. Acting like an ERK pathway inhibitor, NS5806 reduces inflammation-evoked mechanical allodynia, thermal hyperalgesia, and edema by suppressing ERK activation in primary sensory neurons, mast cells, and macrophages. Previous studies show that NS5806 only acts on neurons. This report unveils that NS5806 also acts on immune cells in the skin to exert its anti-inflammatory effects. Since NS5806 is lipid soluble for skin penetration, it suggests that NS5806 could also be developed into an anti-inflammatory drug for external use.

Dual targeting of mast cells and TSLP with a bispecific antibody

Abstract Rationale: Reduction of mast cells (MCs) through stem cell factor (SCF) neutralization combined with inhibition of the alarmin TSLP in a single molecule may result in broader efficacy in inflammatory disorders. Methods: Monoclonal antibodies (mAb) separately targeting SCF and TSLP were generated from immunized mice. A potent neutralizing antibody against each target was selected and humanized. Bispecific tetravalent antibodies (bsAbs) with specificity towards SCF and TSLP were generated. Candidate bsAbs were tested in vitro for neutralizing activity and evaluated in pilot non-human primates (NHP) studies. Results: We discovered a novel anti-SCF neutralizing mAb (SCF-12) that potently inhibits SCF-dependent KIT phosphorylation and SCF-enhanced degranulation of primary human MCs. Separately, a novel anti-TSLP mAb (1D10) inhibits TSLP receptor binding and TSLP-dependent TARC release from human monocyte-derived DCs. bsAbs combining humanized SCF-12 and 1D10 with good expression, biophysical profile and robust neutralizing activity against each target were generated. Following demonstration of MC depletion in NHPs with SCF-12, we conducted a pilot NHP study with two lead candidate bsAbs. Pharmacokinetic, pharmacodynamic and tolerability data from this study will be presented. Conclusions: SCF x TSLP bsAbs were successfully generated from antibodies that neutralize soluble SCF, leading to MC reduction in NHP, and TSLP, a key alarmin implicated in inflammatory and autoimmune disorders. Combined inhibition of these two pathways using a single molecule may be of broad utility in multiple inflammatory disorders. Celldex Therapeutics

Siglec-9 is an inhibitory receptor on human mast cells in vitro

Mast cell activation is critical for the development of allergic diseases. Ligation of sialic acid-binding immunoglobin-like lectins (Siglecs), such as Siglec-6, -7, and -8 as well as CD33, have been shown to inhibit mast cell activation. Recent studies showed that human mast cells express Siglec-9, an inhibitory receptor also expressed by neutrophils, monocytes, macrophages, and dendritic cells. We aimed to characterize Siglec-9 expression and function in human mast cells invitro. We assessed the expression of Siglec-9 and Siglec-9 ligands on human mast cell lines and human primary mast cells by real-time quantitative PCR, flow cytometry, and confocal microscopy. We used a clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) gene editing approach to disrupt the SIGLEC9 gene. We evaluated Siglec-9 inhibitory activity on mast cell function by using native Siglec-9 ligands, glycophorin A(GlycA), and high-molecular-weight hyaluronic acid, a monoclonal antibody against Siglec-9, and coengagement of Siglec-9 with the high-affinity receptor for IgE (FcεRI). Human mast cells express Siglec-9 and Siglec-9 ligands. SIGLEC9 gene disruption resulted in increased expression of activation markers at baseline and increased responsiveness to IgE-dependent and IgE-independent stimulation. Pretreatment with GlycA or high-molecular-weight hyaluronic acid followed by IgE-dependent or -independent stimulation had an inhibitory effect on mast cell degranulation. Coengagement of Siglec-9 with FcεRI in human mast cells resulted in reduced degranulation, arachidonic acid production, and chemokine release. Siglec-9 and its ligands play an important role in limiting human mast cell activation invitro.

Transformation of primary murine peritoneal mast cells by constitutive KIT activation is accompanied by loss of Cdkn2a/Arf expression.

Mast cells (MCs) are immune cells of the myeloid lineage distributed in tissues throughout the body. Phenotypically, they are a heterogeneous group characterized by different protease repertoires stored in secretory granules and differential presence of receptors. To adequately address aspects of MC biology either primary MCs isolated from human or mouse tissue or different human MC lines, like HMC-1.1 and -1.2, or rodent MC lines like L138.8A or RBL-2H3 are frequently used. Nevertheless, cellular systems to study MC functions are very limited. We have generated a murine connective tissue-like MC line, termed PMC-306, derived from primary peritoneal MCs (PMCs), which spontaneously transformed. We analyzed PMC-306 cells regarding MC surface receptor expression, effector functions and respective signaling pathways, and found that the cells reacted very similar to primary wildtype (WT) PMCs. In this regard, stimulation with MAS-related G-protein-coupled receptor member B2 (MRGPRB2) ligands induced respective signaling and effector functions. Furthermore, PMC-306 cells revealed significantly accelerated cell cycle progression, which however was still dependent on interleukine 3 (IL-3) and stem cell factor (SCF). Phenotypically, PMC-306 cells adopted an immature connective tissue-like MCs appearance. The observation of cellular transformation was accompanied by the loss of Cdkn2a and Arf expression, which are both described as critical cell cycle regulators. The loss of Cdkn2a and Arf expression could be mimicked in primary bone marrow-derived mast cells (BMMCs) by sustained SCF supplementation strongly arguing for an involvement of KIT activation in the regulation of Cdkn2a/Arf expression. Hence, this new cell line might be a useful tool to study further aspects of PMC function and to address tumorigenic processes associated with MC leukemia.

Suprabasin-derived polypeptides: SBSN(50-63) induces inflammatory response via TLR4-mediated mast cell activation.

Psoriasis is a chronic, relapsing, immune-mediated, and papulosquamous skin disorder. Excessive mast cell activation, in psoriatic lesions, contributes to inflammation. Various endogenous peptides can participate in the pathogenesis of inflammatory diseases by activating mast cells. Suprabasin (SBSN) is expressed in multiple epithelial tissues and it regulates the normal epidermal barrier function. We have recently shown that suprabasin-derived polypeptides, SBSN(50-63), are significantly increased in psoriatic lesions, through differential peptide analysis. This study was conducted to clarify whether SBSN(50-63) plays a pivotal role in activating mast cells and mediating proinflammatory cytokines and chemokines production in psoriasis. The increased expression of SBSN in psoriatic lesions was confirmed by bioinformatics analysis, PCR and ELISA. Wild-type mice injected subcutaneously with SBSN(50-63) exhibited infiltration of inflammatory cells and the release of cytokines in vivo. SBSN(50-63) stimulated mouse primary mast cells (MPMC) and the laboratory of allergic disease 2 (LAD2) human mast cells to produce more inflammatory mediators than the control group, which were measured ex vivo and in vitro. Toll-like receptor 4 was identified as the receptor of SBSN on mast cells by molecular docking analysis, molecular dynamics simulation, and siRNA transfection. Collectively, SBSN(50-63) could activate mast cells through TLR4, which may increase the inflammatory response in psoriasis.