Mast Cell-mediated Inflammatory Responses Research Articles

Sambou Bamboo salt™ down-regulates the expression levels of angiotensin-converting enzyme 2 in activated human mast cells.

Mast cells have a detrimental impact on coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Sambou Bamboo salt™ (BS) suppresses mast cell-mediated inflammatory response and enhances immunity. In this study, we investigated the regulatory effects of BS on expression of angiotensin-converting enzyme 2 (ACE2) and transmembrane protease/serine subfamily member 2 (TMPRSS2) in human mast cell line (HMC)-1 cells. BS resulted in significant reductions in expression levels of ACE2 and TMPRSS2 in activated HMC-1 cells. Levels of tryptase were reduced by BS. In addition, BS blocked activation of activator protein 1 (AP-1), c-Jun NH2-terminal kinases (JNK), p38, and phosphatidylinositide-3-kinase (PI3K) in activated HMC-1 cells. Therefore, these results show that BS reduces levels of ACE2, TMPRSS2, and tryptase by inhibiting AP-1/JNK/p38/PI3K signaling pathways in mast cells. These findings can serve as valuable foundational data for the development of therapeutic agents aimed at preventing SARS-CoV-2 infection.

Roots of Lithospermum erythrorhizon Alleviated Ovalbumin-Induced Allergic Rhinitis and IgE-triggered Degranulation of RBL-2H3 Cells

Lithospermum erythrorhizon (L. erythrorhizon) root is used in traditional medicine for its anti-inflammatory, antibacterial, and antioxidant properties. However, no studies have examined its impact on allergic rhinitis (AR). Here, we explored the protective effects of L. erythrorhizon in immunoglobulin E (IgE)-stimulated RBL-2H3 cells and in an ovalbumin (OVA)-induced AR mouse model. In the latter, we examined nasal mucosal inflammation, allergen-specific cytokine production, and histological changes to the nasal mucosa. In the mouse model, oral administration of an ethanol extract of L. erythrorhizon (LE) led to a marked reduction in rubbing and sneeze frequency, a significant decrease in serum OVA-specific IgE and IgG1 levels, and a significant increase in the IgG2a/IgG1 ratio. LE also reduced expression of interleukin (IL)-4, IL-5, and IL-13 in nasal lavage fluid (NALF), and suppressed inflammatory cell infiltration and epithelial degradation in nasal tissues. In IgE-stimulated RBL-2H3 cells, LE suppressed release of degranulation markers such as β-hexosaminidase and histamine. Based on these findings, we suggest that LE may ameliorate OVA-induced AR by regulating mast cell-mediated inflammatory responses.

Perfluorooctane sulfonate and bisphenol A induce a similar level of mast cell activation via a common signaling pathway, Fyn-Lyn-Syk activation

Perfluoroalkyl compounds (PFCs) as food contaminants are widely distributed persistent organic pollutants (POPs) and have been suggested to induce immune dysfunction. However, their effects on immune function are not conclusive. Mast cells play a central role in allergic and non-allergic inflammatory responses. Therefore, we have examined the effects of PFCs (PFHxS, PFOA, PFOS) on mast cell-mediated inflammatory responses using in vitro mouse bone marrow-derived mast cells (BMMCs) and human mast cells (HMC-1) and in vivo mice model. The effects of PFCs were compared with those of bisphenol A (BPA), a well-studied environmental pollutant. Among PFCs tested, PFOS had the highest effects. Both PFOS and BPA increased degranulation and production of inflammatory eicosanoids in mast cells at a similar level, which subsequently led to increased skin edema and serum LTC4 and PGD2 in mice. Both PFOS and BPA increased not only downstream signaling (PLCγ1, AKT, ERK), but also upstream signaling (Fyn, Lyn, Syk/LAT) in mast cells. Taken together, PFOS and BPA induce mast cell-mediated inflammatory responses via a common signaling pathways. Our results may help establish the scientific basis for understanding the etiology of mast cell-mediated inflammatory responses and improve the immune dysfunction risk assessment for emerging POPs such as PFCs.

Inhibition of MDM2 expression by rosmarinic acid in TSLP-stimulated mast cell

Rosmarinic acid (RA) has an anti-inflammatory property while thymic stromal lymphopoietin (TSLP) has an important role in mast cell-mediated inflammatory responses. Thus, the aim of this study was to determine the regulatory effect of RA in TSLP-stimulated human mast cell line, HMC-1 cells, and short ragweed pollen-induced allergic conjunctivitis mouse model. As a result, we found that RA significantly decreased the TSLP-induced mast cell proliferation and murine double minute (MDM) 2 expression. RA significantly decreased the levels of interleukin (IL)-13 and phosphorylated the signal transducer and activation of transcription 6 in the TSLP-stimulated HMC-1 cells. RA induced the increment of p53 levels, caspase-3 activation, and poly-ADP-ribose polymerase cleavage and the reduction of the procaspase-3 and Bcl2. RA significantly reduced the production of tumor necrosis factor-α, IL-1β, and IL-6 on the TSLP-stimulated HMC-1 cells. In addition, RA significantly reduced the levels of IgE, IL-4, and TSLP in the short ragweed pollen-induced allergic conjunctivitis mouse model. In conclusion, the results of the study suggest that RA has a significant anti-inflammatory effect on TSLP-induced inflammatory reactions. These effects of RA are likely to be mediated through inhibiting the MDM2 increased by TSLP.

Effects of Hoesaeng-san Ethanol Extract on the Human Mast cell-mediated Inflammatory Responses

Hoesaeng-san is known to be effective for treating diarrhea and vomiting. However the therapeutic mechanism of Hoesaeng-san is still not well understood. The aim of the present study was to demonstrate the effects of Hoesaeng-san ethanol extract (HSSEE) on the expression of pro-inflammatory cytokines, as well as to elucidate its mechanism of action in the human mast cell line (HMC-1). Mast Cell were stimulated with phorbol 12-myristate 13-acetate (PMA) plus A23187 in the presence or absence of HSSEE. To study the possible effects of HSSEE, ELISA, RT-PCR, Western blot analysis were used in this study. HSSEE significantly inhibited the PMA plus A23187-induction of inflammatory cytokines such as tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-8. In activated HMC-1 cells, phosphorylation of extra-signal response kinase (ERK) 1/2 and c-jun n-terminal kinase (JNK)1/2 decreased after treatment with HSSEE. Moreover HSSEE inhibited PMA plus A23187-induced nuclear factor (NF)-κB activation and IκB degradation. HSSEE suppressed the expression of TNF-α, IL-6, IL-8 through a decrease in the ERK 1/2 and JNK 1/2, as well as activation of NF-κB. These results indicated that HSSEE exerted a regulatory effect on inflammatory reactions mediated by mast cells.

Suppressive effects of britanin, a sesquiterpene compound isolated from Inulae flos, on mast cell-mediated inflammatory responses.

Mast cells are central players in immediate-type hypersensitvity and inflammatory responses. In the present study, the effects of britanin on the passive cutaneous anaphylaxis (PCA) reaction in mice and on the phorbol 12-myristate 13-acetate and calcium ionophore A23187 (PMACI)-induced production of pro-inflammatory cytokines in human mast cell line (HMC-1) were evaluated. The oral administration of britanin (10-20 mg/kg) decreased the mast cell-mediated PCA reaction in IgE-sensitized mice. In the activity and mechanism of britanin in vitro assay, britanin suppressed the gene expression and secretion of pro-inflammatory cytokines in a dose-dependent manner in HMC-1. In addition, britanin attenuated PMACI-induced activation of NF-κB as indicated by the inhibition of the degradation of IκBα, nuclear translocation of NF-κB, NF-κB/DNA binding activity assay, and blocked the phosphorylation of p38 MAP kinase, in a dose-dependent manner. We conclude that britanin may have potential as a treatment for allergic-inflammatory diseases.

A Citrus Flavonoid, 6-Demethoxytangeretin, Suppresses Production and Gene Expression of Interleukin-6 in Human Mast Cell-1 <i>via</i> Anaplastic Lymphoma Kinase and Mitogen-Activated Protein Kinase Pathways

Citrus species has been traditionally used in Korea for the treatment of coughing, sputum and dyspepsia. Of the known citrus flavonoids, 6-demethoxytangeretin was reported to exert anti-inflammatory activity. In order to determine the anti-allergic activity of 6-demethoxytangeretin, we examined whether or not 6-demethoxytangeretin was able to suppress activation of the human mast cell line, HMC-1, induced by phorbol 12-myristate 13-acetate (PMA) plus A23187. Interleukin-6 production and relevant gene expression in activated HMC-1 cells were determined by enzyme-linked immunosorbent assay (ELISA) and quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis. Also, the involvement of the anaplastic lymphoma kinase (ALK) and mitogen-activated protein kinases (MAPKs) in activated HMC-1 cells were studied. 6-Demethoxytangeretin suppresses interleukin-6 production, tumor necrosis factor-alpha gene expression, ALK and MAPKs in HMC-1 cells stimulated by PMA plus A23187. Therefore, it was evident that 6-demethoxytangeretin suppressed activation of HMC-1 cells by PMA plus A23187 by inhibiting the activity of ALK and MAPKs and subsequently suppressing gene expression, which suggest that 6-demethoxytangeretin may be involved in the regulation of mast cell-mediated inflammatory responses.

Mouse mannose-binding lectin-A and ficolin-A inhibit lipopolysaccharide-mediated pro-inflammatory responses on mast cells

It is unknown how soluble pattern-recognition receptors in blood, such as mannose-binding lectin (MBL) and ficolins, modulate mast cell-mediated inflammatory responses. We investigate how mouse MBL-A or ficolin-A regulate mouse bone marrow-derived mast cells (mBMMCs)-derived inflammatory response against bacterial lipopolysaccharide (LPS) stimulation. LPS-mediated pro-inflammatory cytokine productions on mBMMCs obtained from Toll-like receptor4 (TLR4)-deficient mice, TLR2-defficient mice, and their wildtype, were specifically attenuated by the addition of either mouse MBL-A or ficolin-A in a dose-dependent manner. However, the inhibitory effects by mouse MBL-A or ficolin-A were restored by the addition of mannose or N-acetylglucosamine, respectively. These results suggest that mouse MBL-A and ficolin-A bind to LPS via its carbohydrate-recognition domain and fibrinogen-like domain, respectively, whereby cytokine production by LPSmediated TLR4 in mBMMCs appears to be down-regulated, indicating that mouse MBL and ficolin may have an inhibitory function toward mouse TLR4-mediated excessive inflammation on the mast cells. [BMB Reports 2013; 46(7): 376-381]

Saucerneol F, a New Lignan Isolated from Saururus chinensis, Attenuates Degranulation via Phospholipase Cγ1 Inhibition and Eicosanoid Generation by Suppressing MAP Kinases in Mast Cells

During our on-going studies to identify bioactive compounds in medicinal herbs, we found that saucerneol F (SF), a naturally occurring sesquilignan isolated from Saururus chinensis (S. chinensis), showed in vitro anti-inflammatory activity. In this study, we examined the effects of SF on the generation of 5-lipoxygenase (5-LO) dependent leukotriene C4 (LTC4), cyclooxygenase-2 (COX-2) dependent prostaglandin D2 (PGD2), and on phospholipase Cγ1 (PLCγ1)-mediated degranulation in SCF-induced mouse bone marrow-derived mast cells (BMMCs). SF inhibited eicosanoid (PGD2 and LTC4) generation and degranulation dose-dependently. To identify the molecular mechanisms underlying the inhibition of eicosanoid generation and degranulation by SF, we examined the effects of SF on the phosphorylation of PLCγ1, intracellular Ca2+ influx, the translocation of cytosolic phospholipase A2 (cPLA2) and 5-LO, and on the phosphorylation of MAP kinases (MAPKs). SF was found to reduce intracellular Ca2+ influx by inhibiting PLCγ1 phosphorylation and suppressing the nuclear translocations of cPLA2 and 5-LO via the phosphorylations of MAPKs, including extracellular signal-regulated protein kinase-1/2 (ERK1/2), c-Jun N-terminal kinase (JNK), and p38. Taken together, these results suggest that SF may be useful for regulating mast cell-mediated inflammatory responses by inhibiting degranulation and eicosanoid generation.

The border-land of epilepsy—Revisited

While probing host skin to search for blood vessels, the female <i>Anopheles</i> mosquito delivers <i>Plasmodium</i> parasites in the presence of saliva. Saliva from various blood-feeding vectors which contains several pharmacologically active components is believed to facilitate blood feeding as well as parasite transmission to the host. Recently, we found that mosquito saliva has the capacity to activate dermal mast cells and to induce local inflammatory cell influx. Our main objective in the present work is to investigate whether saliva, through mosquito bites, controls the magnitude of Ag-specific immune responses and whether this control is dependent on the mast cell-mediated inflammatory response. Using a mast cell knockin mouse model, we found that mosquito bites consistently induced MIP-2 in the skin and IL-10 in draining lymph nodes, and down-regulate Ag-specific T cell responses by a mechanism dependent on mast cells and mediated by IL-10. Our results provide evidence for new mechanisms which may operate during <i>Plasmodium</i> parasite transmission by mosquito bites.

Flavonoids in propolis acting on mast cell-mediated wound healing

Barroso et al. have shown, on the latest issue of Inflammopharmacology, that the topical application of propolis on surgical wounds affected the number of mast cells recruited in these sites, and suggested two well-known antiinflammatory components present in propolis, namely caffeic acid and artepillin C, as possible active molecules (Borrelli et al. 2002; Paulino et al. 2008). It is widely acknowledged that propolis down-regulates type I allergy and inflammation by affecting mast cells, but the effective components of propolis, which cause these effects, remain still unknown. Propolis components vary depending on the area from which they are collected, mainly because of the genetic variability among wild plants in different geographic regions: variability in phenolics composition may result in different biochemical property of the propolis, depending on the main component active in raw propolis or in its extracts. In Chinese propolis, chrysin, kaempferol and its derivative, pinocembrin and galangin were identified as the main flavonoids able to act on mast cell-mediated inflammatory response, while chrysin was shown to inhibit IL-4 and MCP-1 production from antigen-stimulated RBL2H3 basophil/mast cell lines (Nakamura et al. 2010). On the other hand, Brazilian propolis extract contains only small amounts of these flavonoids, which might suggest that variation in propolis components could affect antiallergic and anti-inflammatory properties (Nakamura et al. 2010). Brazilian propolis contains, therefore, major percentage of phenolic acids, such as caffeic acid phenethyl ester (CAPE) and artepillin C (Park et al. 2002). As the activity of propolis, like many natural products, may be due to the synergic effect of several bioactive components, it will be necessary to distinguish between different types of propolis and analyse its complex compositions to guarantee specific biological activities of propolis diffused worldwide (Frankland Sawaya et al. 2011). Furthermore, inflammation by mast cells can be inhibited by several flavonoids present in propolis. Recent evidence was reported showing that chrysin decreased gene expression of pro-inflammatory cytokines, such as TNF-a, IL-1b, IL-4 and IL-6 in mast cells by a nuclear factor-jB (NF-jB) and caspase-1 dependent mechanism (Bae et al. 2011). Genistein modulates NF-jB and TNF-a expression during the early stage of wound healing (Park et al. 2011). CAPE accelerates cutaneous wound healing and it is arguable that propolis with a significant amount of this phenolic acid may exert a wound repairing property (Serarslan et al. 2007). The antiinflammatory property attributed to flavonoids in propolis might suggest that these polyphenols should exert their action toward other newly discovered function of mast cells (Ng 2010), even in association with CAPE or other phenolic acids. Wound healing is a complex process of lysis and reconstitution controlled by a series of cell signaling proteins and involved tissue regeneration and angiogenesis (Hiromatsu and Toda 2003; Nienartowicz et al. 2006). Mast cells have been shown to play a significant role in the early inflammatory stage of wound healing and also influence proliferation and tissue remodeling in skin; according to some authors in wound healing, mast cells are able to modulate only the inflammatory but not the proliferative response in healing and reconstituting damaged tissue (Egozi et al. 2003), while other reports showed that mast cells are involved in tissue remodeling in the late phase of wound healing (Iba et al. 2004), thus expanding S. Chirumbolo (&) Unit of Geriatry, Department of Medicine, University Laboratory of Medical Research, University of Verona, Policlinico GB Rossi piazzale AL Scuro 10, 37134 Verona, Italy e-mail: salvatore.chirumbolo@univr.it Inflammopharmacol (2012) 20:99–101 DOI 10.1007/s10787-012-0125-9 Inflammopharmacology

Inhibitory effects of chelidonic acid on IL-6 production by blocking NF-κB and caspase-1 in HMC-1 cells

Chelidonic acid (CA) is a γ-pyrone which is contained in the rhizome of Chelidonium majus L. It has multiple pharmacological effects including those of a mild analgesic, an antimicrobial, an oncostatic and a central nervous system sedative, but the anti-inflammatory effect of CA and its molecular mechanisms are poorly understood. In this study, we investigated the regulatory mechanism of CA in mast cell-mediated inflammatory response by phorbol 12-myristate 13-acetate and calcium ionophore A23187. The results indicate that CA inhibits the production of interleukin-6 (IL-6) and the expression of IL-6 mRNA through the regulation of nuclear factor-κB. In addition, CA suppresses the activation and expression of caspase-1. These results provide new insights into the pharmacological actions of CA as a potential molecule for use in therapy in mast cell-mediated inflammatory diseases.

Panax Ginseng inhibited HIF-1a activation and inflammatory cytokine in HMC-1 cells activated by phorbol myristate acetate and A23187

This study investigated the role of Panax ginseng (PG) on the phorbol myristate acetate (PMA) + calcium ionophore A23187-induced hypoxia-inducible factor-<TEX>$1{\alpha}$</TEX> (HIF-<TEX>$1{\alpha}$</TEX>) activation, phosphorylation of the extracellular signal-regulated kinase (ERK), and inflammatory cytokine production from the human mast cell line, HMC-1. HIF-<TEX>$1{\alpha}$</TEX> and phosphorylation of ERK were observed by Western blotting. The inflammatory cytokine production was determined by enzyme-linked immunosorbent assay. PG inhibited the PMA+A23187-induced HIF-<TEX>$1{\alpha}$</TEX> expression and the subsequent production of vascular endothelial growth factor. In addition, PG suppressed PMA + A23187-induced phosphorylation of ERK. We also show that the increased cytokines interleukin (IL)-<TEX>$1{\beta}$</TEX>, IL-6, and tumour necrosis factor-<TEX>${\alpha}$</TEX> level was significantly inhibited by treatment of PG. In the present study, we report for the first time that PG is an inhibitor of HIF-<TEX>$1{\alpha}$</TEX> and cytokines on the mast cell-mediated inflammatory responses.

Mast cell mediated inflammatory response in chickens after infection with very virulent infectious bursal disease virus

The potential role of the mast cells in the invasion of very virulent infectious bursal disease virus (vvIBDV) is unknown. We evaluated mast cell activity and tryptase production after vvIBDV infection in special pathogen-free (SPF) chickens using cytochemistry and immunohistochemistry analyses. The results were as follows: (1) severe histologic lesions were observed in the thymus, spleen, cloacal bursa, liver, kidney and other tissues. vvIBDV viral antigens were detected and presented extensively in the parenchymatous organs, in particular, the cloacal bursa, liver, kidney, thymus, spleen and pancreas. (2) In the vvIBDV-infected group, the mast cell population increased markedly in the liver, kidney, thymus, glandular stomach, spleen and cloacal bursa on days 1, 2 and 3 after vvIBDV infection ( p < 0.05). However, very few mast cells were observed in those same tissues in the controls, especially in the bursa of Fabricius. (3) Tryptase, a marker for activated mast cells, has a positive correlation with mast cell distribution. The mast cells identified in the tissues were likely to be activated since they were associated with cell degranulation and the presence of tryptase. Furthermore, the co-localization of mast cells, and presence of vvIBDV antigens suggests that the mast cells were activated by vvIBDV infection. Our results also suggest that tryptase may contribute to the inflammation of acute IBD induced by vvIBDV infection. Our research contributes to the further understanding of inflammatory response mechanisms and the contribution of mast cell activity to this process.

FcγRI-Mediated Activation of Human Mast Cells Promotes Survival and Induction of the Pro-survival Gene Bfl-1

Activation of mast cells through either FcepsilonRI or FcgammaRI leads to release of mediators contributing to the inflammatory response. One of the biologic characteristics of mast cells in allergic pathology is that these cells have the capacity to recover and regranulate after aggregation of FcepsilonRI. We have previously demonstrated that the pro-survival protein A1/Bfl-1 is required for mast cells to survive IgE-mediated activation. In the present study, we have investigated whether human mast cells show similar induction of bfl-1 and activation-induced survival after aggregation of FcgammaRI. Human cord blood-derived mast cells were activated by aggregation of either FcepsilonRI or FcgammaRI, and activation-induced survival and induction of bfl-1 was measured. We found that aggregation of FcgammaRI-induced expression of Bfl-1 and caused a comparable activation-induced mast cell survival as FcepsilonRI does. These data suggests that activation through Fc-receptors contribute to mast cell survival during antibody-dependent mast cell mediated inflammatory responses.

Hesperidin inhibits expression of hypoxia inducible factor-1 alpha and inflammatory cytokine production from mast cells

The citrus unshiu peel has been used traditionally as a medicine to improve bronchial and asthmatic conditions or cardiac and blood circulation in Korea, China, and Japan. Here, we report the effects of citrus unshiu peel water extract (CPWE) on the phorbol myristate acetate (PMA)+calcium ionophore A23187-induced hypoxia-inducible factor-1alpha (HIF-1alpha) activation and inflammatory cytokine production from the human mast cell line, HMC-1 cells. We compared CPWE with hesperidin, a common constituent of citrus unshiu. CPWE and hesperidin inhibited the PMA+A23187-induced HIF-1alpha expression and the subsequent production of vascular endothelial growth factor (VEGF). In addition, CPWE suppressed PMA+A23187-induced phosphorylation of the extracellular signal-regulated kinase (ERK). We also show that the increased cytokines interleukin (IL)-1beta, IL-8, and tumor necrosis factor (TNF)-alpha level was significantly inhibited by treatment of CPWE or hesperidin. In the present study, we report that CPWE and hesperidin are inhibitors of HIF-1alpha and cytokines on the mast cell-mediated inflammatory responses.

Mast Cell-Dependent Down-Regulation of Antigen-Specific Immune Responses by Mosquito Bites

While probing host skin to search for blood vessels, the female Anopheles mosquito delivers Plasmodium parasites in the presence of saliva. Saliva from various blood-feeding vectors which contains several pharmacologically active components is believed to facilitate blood feeding as well as parasite transmission to the host. Recently, we found that mosquito saliva has the capacity to activate dermal mast cells and to induce local inflammatory cell influx. Our main objective in the present work is to investigate whether saliva, through mosquito bites, controls the magnitude of Ag-specific immune responses and whether this control is dependent on the mast cell-mediated inflammatory response. Using a mast cell knockin mouse model, we found that mosquito bites consistently induced MIP-2 in the skin and IL-10 in draining lymph nodes, and down-regulate Ag-specific T cell responses by a mechanism dependent on mast cells and mediated by IL-10. Our results provide evidence for new mechanisms which may operate during Plasmodium parasite transmission by mosquito bites.

TGF-β1 Inhibits Mast Cell FcεRI Expression

Mast cell activation through the high affinity IgE receptor (FcepsilonRI) is a critical component of atopic inflammation. The cytokine TGF-beta1 has been shown to inhibit IgE-dependent mast cell activation, possibly serving to dampen mast cell-mediated inflammatory responses. We present proof that TGF-beta1 inhibits mast cell FcepsilonRI expression through a reversible pathway that diminishes protein, but not mRNA, expression of the FcepsilonRI subunit proteins alpha, beta, and gamma. The stability of the expressed proteins and the assembled cell surface complex was unaltered by TGF-beta1 treatment. However, TGF-beta1 decreased the rate of FcepsilonRI beta-chain synthesis, arguing that this inhibitory cytokine exerts its effects at the level of mRNA translation. TGF-beta1 consistently diminished FcepsilonRI expression on cultured human or mouse mast cells as well as freshly isolated peritoneal mast cells. The related cytokines, TGF-beta2 and TGF-beta3, had similar effects. We propose that TGF-beta1 acts as a negative regulator of mast cell function, in part by decreasing FcepsilonRI expression.

Identification and characterization of a new pair of immunoglobulin-like receptors LMIR1 and 2 derived from murine bone marrow-derived mast cells

We have identified and characterized two mouse cDNAs in a mouse antigen-stimulated bone marrow-derived mast cell cDNA library, both of which encode type I transmembrane proteins. The genes were closely mapped in the distal region of mouse chromosome 11 and expressed not only in mast cells but also widely in leukocytes. The extracellular domains of their encoded proteins contain a single variable immunoglobulin (Ig) motif sharing about 90% identity with amino acids, showing that they comprise a pair of molecules and belong to the Ig superfamily. We named these molecules leukocyte mono-Ig-like receptor1 and 2 (LMIR1 and 2). The intracellular domain of LMIR1 contains several immunoreceptor tyrosine-based inhibition motifs (ITIMs). When cross-linked, the intracellular domain was tyrosine phosphorylated and capable of recruiting tyrosine phosphatases, SHP-1 and SHP-2 and inositol polyphosphate 5-phosphatase, SHIP. LMIR2, on the other hand, contains a short cytoplasmic tail and a characteristic transmembrane domain carrying two positively charged amino acids associated with three kinds of immunoreceptor tyrosine-based activation motif (ITAM)-bearing molecules, DAP10, DAP12, and FcRγ. These findings suggest that a new pair of ITIM/ITAM-bearing receptors, LMIR1 and 2, regulate mast cell-mediated inflammatory responses through yet to be defined ligand(s).

Inhibition of Kit Expression by IL-4 and IL-10 in Murine Mast Cells: Role of STAT6 and Phosphatidylinositol 3′-Kinase

The c-kit protooncogene encodes a receptor tyrosine kinase that is known to play a critical role in hemopoiesis and is essential for mast cell growth, differentiation, and cytokine production. Studies have shown that the Th2 cytokine IL-4 can down-regulate Kit expression on human and murine mast cells, but the mechanism of this down-regulation has remained unresolved. Using mouse bone marrow-derived mast cells, we demonstrate that IL-4-mediated Kit down-regulation requires STAT6 expression and phosphotidylinositide-3'-kinase activation. We also find that the Th2 cytokine IL-10 potently down-regulates Kit expression. IL-4 enhances IL-10-mediated inhibition in a manner that is STAT6 independent and phosphotidylinositide-3'-kinase dependent. Both IL-4- and IL-10-mediated Kit down-regulation were coupled with little or no change in c-kit mRNA levels, no significant change in Kit protein stability, but decreased total Kit protein expression. Inhibition of Kit expression by IL-4 and IL-10 resulted in a loss of Kit-mediated signaling, as evidenced by reduced IL-13 and TNF-alpha mRNA induction after stem cell factor stimulation. These data offer a role for STAT6 and phosphotidylinositide-3'-kinase in IL-4-mediated Kit down-regulation, coupled with the novel observation that IL-10 is a potent inhibitor of Kit expression and function. Regulating Kit expression and signaling may be essential to controlling mast cell-mediated inflammatory responses.