Seung Taek Nam

Mesenteric IL-10-producing CD5+ regulatory B cells suppress cow’s milk casein-induced allergic responses in mice

Food allergy is a hypersensitive immune reaction to food proteins. We have previously demonstrated the presence of IL-10-producing CD5+ B cells and suggested their potential role in regulating cow’s milk casein allergy in humans and IgE-mediated anaphylaxis in mice. In this study, we determined whether IL-10-producing CD5+ regulatory B cells control casein-induced food allergic responses in mice and, if so, the underlying mechanisms. The induction of oral tolerance (OT) by casein suppressed casein-induced allergic responses including the decrease of body temperature, symptom score, diarrhea, recruitment of mast cells and eosinophils into jejunum, and other biological parameters in mice. Notably, the population of IL-10-producing CD5+ B cells was increased in mesenteric lymph node (MLN), but not in spleen or peritoneal cavity (PeC) in OT mice. The adoptive transfer of CD5+ B cells from MLN, but not those from spleen and PeC, suppressed the casein-induced allergic responses in an allergen-specific and IL-10-dependent manner. The inhibitory effect of IL-10-producing CD5+ B cells on casein-induced allergic response was dependent on Foxp3+ regulatory T cells. Taken together, mesenteric IL-10-producing regulatory B cells control food allergy via Foxp3+ regulatory T cells and could potentially act as a therapeutic regulator for food allergy.

Autocrine stimulation of IL-10 is critical to the enrichment of IL-10-producing CD40hiCD5+ regulatory B cells in vitro and in vivo

IL-10-producing B (Breg) cells regulate various immune responses. However, their phenotype remains unclear. CD40 expression was significantly increased in B cells by LPS, and the Breg cells were also enriched in CD40hiCD5+ B cells. Furthermore, CD40 expression on Breg cells was increased by IL-10, CD40 ligand, and B cell-activating factor, suggesting that CD40hi is a common phenotype of Breg cells. LPS-induced CD40 expression was largely suppressed by an anti-IL-10 receptor antibody and in IL-10−/−CD5+CD19+ B cells. The autocrine effect of IL-10 on the CD40 expression was largely suppressed by an inhibitor of JAK/STAT3. In vivo, the LPS treatment increased the population of CD40hiCD5+ Breg cells in mice. However, the population of CD40hiCD5+ B cells was minimal in IL-10−/− mice by LPS. Altogether, our findings show that Breg cells are largely enriched in CD40hiCD5+ B cells and the autocrine effect of IL-10 is critical to the formation of CD40hiCD5+ Breg cells. [BMB Reports 2015; 48(1): 54-59]

Role of NADH: quinone oxidoreductase-1 in the tight junctions of colonic epithelial cells.

NADH:quinone oxidoreductase 1 (NQO1) is known to be involved in the regulation of energy synthesis and metabolism, and the functional studies of NQO1 have largely focused on metabolic disorders. Here, we show for the first time that compared to NQO1-WT mice, NQO1-KO mice exhibited a marked increase of permeability and spontaneous inflammation in the gut. In the DSS-induced colitis model, NQO1-KO mice showed more severe inflammatory responses than NQO1-WT mice. Interestingly, the transcript levels of claudin and occludin, the major tight junction molecules of gut epithelial cells, were significantly decreased in NQO1-KO mice. The colons of NQO1-KO mice also showed high levels of reactive oxygen species (ROS) and histone deacetylase (HDAC) activity, which are known to affect transcriptional regulation. Taken together, these novel findings indicate that NQO1 contributes to the barrier function of gut epithelial cells by regulating the transcription of tight junction molecules. [BMB Reports 2014;47(9): 494-499]

Suppression of IgE-mediated mast cell activation and mouse anaphylaxis via inhibition of Syk activation by 8-formyl-7-hydroxy-4-methylcoumarin, 4μ8C

ABSTRACT Mast cells trigger IgE‐mediated allergic reactions by releasing various allergic mediators. 8‐Formyl‐7‐hydroxy‐4‐methylcoumarin, also called 4&mgr;8C, was originally known as an inositol‐requiring enzyme 1 (IRE1) suppressant, but no study has examined its relationship with mast cells and allergic diseases. Therefore, the purpose of this study was to determine whether 4&mgr;8C is effective in suppressing allergic reactions in mast cells and in IgE‐mediated allergic animal model. 4&mgr;8C suppressed the degranulation of IgE‐mediated mast cells (IC50 = 3.2 &mgr;M) and the production of cytokines such as tumor necrosis factor‐&agr; (TNF‐&agr;) and interleukin‐4 (IL‐4) in a dose‐dependent manner. 4&mgr;8C also suppressed passive cutaneous anaphylaxis (PCA) in mice (ED50 = 25.1 mg/kg). In an experiment on mast cell signaling pathways stimulated by antigen, the phosphorylation and activation of Syk was decreased by 4&mgr;8C, and phosphorylation of downstream signaling molecules, such as linker for activated T cells (LAT), Akt, and the three MAP kinases, ERK, p38, and JNK, were suppressed. Mechanistic studies showed that 4&mgr;8C inhibited the activity of Lyn and Fyn in vitro. Based on the results of those experiments, the suppressor mechanism of allergic reaction by 4&mgr;8C involved reduced activity of Lyn and Fyn, which is pivotal in an IgE‐mediated signaling pathway. In summary, for the first time, this study shows that 4&mgr;8C inhibits Lyn and Fyn, thus suppressing allergic reaction by reducing the degranulation and the production of inflammatory cytokines. This suggests that 4&mgr;8C can be used as a new medicinal candidate to control allergic diseases such as seasonal allergies and atopic dermatitis. HighlightsAnti‐allergic effect of 8‐formyl‐7‐hydroxy‐4‐methylcoumarin, 4&mgr;8C, was investigated in vitro and in vivo.4&mgr;8C reversibly inhibited the degranulation of mast cells by antigen.4&mgr;8C suppressed passive cutaneous anaphylaxis (PCA) in mice.The anti‐allergic effect of 4&mgr;8C was mediated by inhibition of Lyn and Fyn in mast cells.

An indoxyl compound 5-bromo-4-chloro-3-indolyl 1,3-diacetate, CAC-0982, suppresses activation of Fyn kinase in mast cells and IgE-mediated allergic responses in mice.

Mast cells, constituents of virtually all organs and tissues, are critical cells in IgE-mediated allergic responses. The aim of this study was to investigate the effect and mechanism of an indoxyl chromogenic compound, 5-bromo-4-chloro-3-indolyl 1,3-diacetate, CAC-0982, on IgE-mediated mast cell activation and allergic responses in mice. CAC-0982 reversibly suppressed antigen-stimulated degranulation in murine mast cells (IC50, ~3.8μM) and human mast cells (IC50, ~3.0μM). CAC-0982 also inhibited the expression and secretion of IL-4 and TNF-α in mast cells. Furthermore, CAC-0982 suppressed the mast cell-mediated allergic responses in mice in a dose-dependent manner (ED50 27.9mg/kg). As for the mechanism, CAC-0982 largely suppressed the phosphorylation of Syk and its downstream signaling molecules, including LAT, Akt, Erk1/2, p38, and JNK. Notably, the tyrosine kinase assay of antigen-stimulated mast cells showed that CAC-0982 inhibited Fyn kinase, one of the upstream tyrosine kinases for Syk activation in mast cells. Taken together, these results suggest that CAC-0982 may be used as a new treatment for regulating IgE-mediated allergic diseases through the inhibition of the Fyn/Syk pathway in mast cells.

The Extract of Chrysanthemum zawadskii var. latilobum Ameliorates Collagen-Induced Arthritis in Mice.

Chrysanthemum zawadskii var. latilobum (CZ) has been used for beverage or tea and also as folk medicine for the remedy of diverse inflammatory diseases. Nevertheless, the therapeutic effect of CZ on arthritis remains to be unknown. In this paper we aim to investigate the CZs antiarthritic effect and mechanism of action both in vitro and in vivo. To assess CZs antiarthritic effect, mouse models of type II collagen-induced arthritis (CIA) were used. Mice were used to gauge clinical arthritis index and histopathological changes. Reverse transcriptase-polymerase chain reaction (RT-PCR), western blotting, electrophoretic mobility shift assay (EMSA), and other biological methods were adopted to measure CZs effect on arthritis and to understand the veiled mechanism of action. CZ greatly suppressed CIA, histopathological score, bone erosion, and osteoclast differentiation. Mechanistically, CZ inhibited the production of various inflammatory and arthritic mediators like inflammatory cytokines, matrix metalloproteinases (MMPs), and chemokines. Of note, CZ significantly suppressed the activation of the NF-κB pathway in vivo. CZ exerted an antiarthritic effect in CIA mice by curbing the production of crucial inflammatory and arthritis mediators. This study warrants further investigation of CZ for the use in human rheumatoid arthritis (RA).

DJ-1 controls bone homeostasis through the regulation of osteoclast differentiation

Receptor activator of NF-kB ligand (RANKL) generates intracellular reactive oxygen species (ROS), which increase RANKL-mediated signaling in osteoclast (OC) precursor bone marrow macrophages (BMMs). Here we show that a ROS scavenging protein DJ-1 negatively regulates RANKL-driven OC differentiation, also called osteoclastogenesis. DJ-1 ablation in mice leads to a decreased bone volume and an increase in OC numbers. In vitro, the activation of RANK-dependent signals is enhanced in DJ-1-deficient BMMs as compared to wild-type BMMs. DJ-1 suppresses the activation of both RANK-TRAF6 and RANK-FcRγ/Syk signaling pathways because of activation of Src homology region 2 domain-containing phosphatase-1, which is inhibited by ROS. Ablation of DJ-1 in mouse models of arthritis and RANKL-induced bone disease leads to an increase in the number of OCs, and exacerbation of bone damage. Overall, our results suggest that DJ-1 plays a role in bone homeostasis in normal physiology and in bone-associated pathology by negatively regulating osteoclastogenesis.Osteoclasts are involved in arthritis, and their differentiation depends on RANKL signaling. The author show that the ROS-scavenging protein DJ-1 negatively regulates RANKL signaling and that its ablation increases osteoclast numbers and exacerbates bone damage in mouse models of arthritis.

JQ1, a BET inhibitor, controls TLR4-induced IL-10 production in regulatory B cells by BRD4-NF-κB axis

Regulatory B cells, also well-known as IL-10-producing B cells, play a role in the suppression of inflammatory responses. However, the epigenetic modulation of regulatory B cells is largely unknown. Recent studies showed that the bromodomain and extra-terminal domain (BET) protein inhibitor JQ1 controls the expression of various genes involving cell proliferation and cell cycle. However, the role of BET proteins on development of regulatory B cells is not reported. In this study, JQ1 potently suppressed IL-10 expression and secretion in murine splenic and peritoneal B cells. While bromodomain-containing protein 4 (BRD4) was associated with NF-κB on IL-10 promoter region by LPS stimulation, JQ1 interfered the interaction of BRD4 with NF-κB on IL-10 promoter. In summary, BRD4 is essential for toll like receptor 4 (TLR4)-mediated IL-10 expression, suggesting JQ1 could be a potential candidate in regulating IL-10-producing regulatory B cells in cancer.

Furaltadone suppresses IgE-mediated allergic response through the inhibition of Lyn/Syk pathway in mast cells

ABSTRACT Mast cells are critical cells that prompt various allergic response‐inducing factors, contributing to allergic diseases. While used as an antibiotic for livestock, there is no study on the effect of furaltadone on allergic response. This study investigated the effect of furaltadone on mast cells and passive cutaneous anaphylaxis (PCA). Furaltadone inhibited the degranulation of mast cells stimulated by antigen (IC50, ˜ 3.9&mgr;M), and also suppressed the production of tumor necrosis factor (TNF)‐&agr; and interleukin (IL)‐4 in a concentration dependent manner. In addition, furaltadone inhibited allergic responses in an acute allergy animal model, PCA. Further investigation on the mechanism for these inhibitory effects of furaltadone found that the activities of Lyn/Syk and Syk‐dependent downstream proteins such as mitogen‐activated protein (MAP) kinases were inhibited by furaltadone in mast cells. Taken together, this study demonstrates that furaltadone inhibits the activation of mast cells by antigen via the suppression of the Lyn/Syk pathway and ameliorates allergic responses in vivo.

An Anti-Cancer Drug Candidate CYC116 Suppresses Type I Hypersensitive Immune Responses through the Inhibition of Fyn Kinase in Mast Cells

Mast cells are the most prominent effector cells of Type 1 hypersensitivity immune responses. CYC116 [4-(2-amino-4-methyl-1,3-thiazol-5-yl)-N-[4-(morpholin-4-yl)phenyl] pyrimidin-2-amine] is under development to be used as an anti-cancer drug, but the inhibitory effects of CYC116 on the activation of mast cells and related allergy diseases have not reported as of yet. In this study, we demonstrated, for the first time, that CYC116 inhibited the degranulation of mast cells by antigen stimulation (IC50, ∼1.42 µM). CYC116 also inhibited the secretion of pro-inflammatory cytokines including TNF-α (IC50, ∼1.10 µM), and IL-6 (IC50, ∼1.24 µM). CYC116 inhibited the mast cell-mediated allergic responses, passive cutaneous anaphylaxis (ED50, ∼22.5 mg/kg), and passive systemic anaphylaxis in a dose-dependent manner in laboratory experiments performed on mice. Specifically, CYC116 inhibited the activity of Fyn in mast cells and inhibited the activation of Syk and Syk-dependent signaling proteins including LAT, PLCγ, Akt, and MAP kinases. Our results suggest that CYC116 could be used as an alternative therapeutic medication for mast cell-mediated allergic disorders, such as atopic dermatitis and allergic rhinitis.