A novel natural Syk inhibitor suppresses IgE-mediated mast cell activation and passive cutaneous anaphylaxis.

Spleen tyrosine kinase (Syk) plays a crucial role as a target for allergy treatment due to its involvement in immunoreceptor signaling. The purpose of this study was to identify natural inhibitors of Syk and assess their effects on the IgE-mediated allergic response in mast cells and ICR mice. A list of eight compounds was selected based on pharmacophore and molecular docking, showing potential inhibitory effects through virtual screening. Among these compounds, sophoraflavanone G (SFG) was found to inhibit Syk activity in an enzymatic assay, with an IC50 value of 2.2 µM. To investigate the conformational dynamics of the SYK-SFG system, we performed molecular dynamics simulations. The stability of the binding between SFG and Syk was evaluated using root mean square deviation (RMSD) and root mean square fluctuation (RMSF). In RBL-2H3 cells, SFG demonstrated a dose-dependent suppression of IgE/BSA-induced mast cell degranulation, with no significant cytotoxicity observed at concentrations below 10.0 µM within 24 h. Furthermore, SFG reduced the production of TNF-α and IL-4 in RBL-2H3 cells. Mechanistic investigations revealed that SFG inhibited downstream signaling proteins, including phospholipase Cγ1 (PLCγ1), as well as mitogen-activated protein kinases (AKT, Erk1/2, p38, and JNK), in mast cells in a dose-dependent manner. Passive cutaneous anaphylaxis (PCA) experiments demonstrated that SFG could reduce ear swelling, mast cell degranulation, and the expression of COX-2 and IL-4. Overall, our findings identify naturally occurring SFG as a direct inhibitor of Syk that effectively suppresses mast cell degranulation both in vitro and in vivo.

Suppression of Mast Cell Activation by GPR35: GPR35 Is a Primary Target of Disodium Cromoglycate.

Mast cell stabilizers, including disodium cromoglycate (DSCG), were found to have potential as the agonists of an orphan G protein-coupled receptor, GPR35, although it remains to be determined whether GPR35 is expressed in mast cells and involved in suppression of mast cell degranulation. Our purpose in this study is to verify the expression of GPR35 in mast cells and to clarify how GPR35 modulates the degranulation. We explored the roles of GPR35 using an expression system, a mast cell line constitutively expressing rat GPR35, peritoneal mast cells, and bone marrow-derived cultured mast cells. Immediate allergic responses were assessed using the IgE-mediated passive cutaneous anaphylaxis (PCA) model. Various known GPR35 agonists, including DSCG and newly designed compounds, suppressed IgE-mediated degranulation. GPR35 was expressed in mature mast cells but not in immature bone marrow-derived cultured mast cells and the rat mast cell line. Degranulation induced by antigens was significantly downmodulated in the mast cell line stably expressing GPR35. A GPR35 agonist, zaprinast, induced a transient activation of RhoA and a transient decrease in the amount of filamentous actin. GPR35 agonists suppressed the PCA responses in the wild-type mice but not in the GPR35-/- mice. These findings suggest that GPR35 should prevent mast cells from undergoing degranulation induced by IgE-mediated antigen stimulation and be the primary target of mast cell stabilizers. SIGNIFICANCE STATEMENT: The agonists of an orphan G protein-coupled receptor, GPR35, including disodium cromoglycate, were found to suppress degranulation of rat and mouse mature mast cells, and their antiallergic effects were abrogated in the GPR35-/- mice, indicating that the primary target of mast cell stabilizers should be GPR35.

Kechuanning Gel Plaster Exerts Anti-inflammatory and Immunomodulatory Effects on Ovalbumin-induced Asthma Model Rats via ERK Pathway.

BACKGROUND: We aimed to evaluate the therapeutic effects of Kechuanning gel plaster on ovalbumin (OVA)-induced rat model of asthma. METHODS: Rats were injected with OVA to induce asthma, and Kechuanning gel plaster was administered after the OVA challenge. The immune cell counts in the bronchial alveolar lavage fluid (BALF) were calculated after Kechuanning gel plaster administration. The levels of immune factors in BALF and serum OVA-specific IgE levels were analyzed. Western blot analysis and immunohistochemistry were carried out to analyze the following proteins: C-FOS, C-JUN, RAS p21 protein activator 1 (RASA1), matrix metalloproteinase 9 (MMP9), RAF1, p-MEK1, tissue inhibitor of metalloproteinase-1 (TIMP1), and p-extracellular signal-regulated kinase 1 (ERK1). RESULTS: Administration of Kechuanning gel plaster led to decreased immune cell counts, inflammatory cytokines (interleukin (IL)-1ß, IL13, and IL17), and OVA-specific IgE expression. Compared to the normal group, the C-FOS, C-JUN, RASA1, MMP9, RAF1, MEK1, TIMP1, and p- ERK1 expressions in the model group were significantly increased, whereas Kechuanning gel plaster administration decreased C-JUN, MMP9, TIMP1, RAF1, MEK1, p-ERK1, C-FOS, and RASA1 protein levels. CONCLUSION: Kechuanning gel plaster exerted its therapeutic effects on OVA-induced asthma model rats through the ERK signaling pathway. Kechuanning gel plaster could be considered as a potential alternative therapeutic agent for the management of asthma.

Development and Evaluation of Tacrolimus Loaded Nano-Transferosomes for Skin Targeting and Dermatitis Treatment.

Tacrolimus (TRL) is used for the treatment of atopic dermatitis (AD) due to its T-cell stimulation effect. However, its significantly poor water solubility, low penetration and cytotoxicity have reduced its topical applications. Herein, tacrolimus loaded nano transfersomes (TRL-NTs) were prepared, followed by their incorporation into chitosan gel to prepare tacrolimus loaded nano transfersomal gel (TRL-NTsG). TEM analysis of the TRL-NTs was performed to check their morphology. DSC, XRD and FTIR analysis of the TRL-NTs were executed after lyophilization. Similarly, rheology, spreadability and deformability of the TRL-NTsG were investigated. In vitro release, ex vivo permeation and in vitro interaction of TRL-NTsG with keratinocytes and fibroblasts as well as their co-cultures were investigated along with their in vitro cell viability analysis. Moreover, in vivo skin deposition, ear thickness, histopathology and IgE level were also determined. Besides, 6 months stability study was also performed. Results demonstrated the uniformly distributed negatively charged nanovesicles with a mean particle size distribution of 163 nm and zeta potential of -27 mV. DSC and XRD exhibited the thermal stability and amorphous form of the drug, respectively. The TRL-NTsG showed excellent deformability, spreadability and rheological behavior. In vitro release studies exhibited an 8-fold better release of TRL from the TRL-NTsG. Similarly, 6-fold better permeation and stability of the TRL-NTsG with keratinocytes and fibroblasts as well as their co-cultures was observed. Furthermore, the ear thickness (0.6 mm) of the TRL-NTsG was found significantly reduced when compared with the untreated (1.7 mm) and TRL conventional gel treated mice (1.3 mm). The H&E staining showed no toxicity of the TRL-NTsG with significantly reduced IgE levels (120 ng/mL). The formulation was found stable for at least 6 months. These results suggested the efficacy of TRL in AD-induced animal models most importantly when incorporated in NTsG.

Fenofibrate attenuates asthma features in an ovalbumin-induced mouse model via suppressing NF-κB binding activity.

BACKGROUND/AIM: Asthma is a chronic inflammatory disease of the airways with a high prevalence. Asthma has a complex pathophysiology and about 5-10% of patients are not fully responsive to the currently available treatments. The aim of this study is to investigate the involvement of NF-κB in the effects of fenofibrate on a mouse model of allergic asthma. MATERIALS AND METHODS: A total of 49 BALB/c mice were randomly distributed into 7 groups (n = 7). Allergic asthma model was created by administering i.p. injections of ovalbumin on days 0, 14 and 21, followed by provocation with inhaled ovalbumin on days 28, 29 and 30. Fenofibrate was orally given in 3 different doses; 1, 10 and 30 mg/kg through days 21-30 of the experiment. On day 31, pulmonary function test using whole body plethysmography was performed. The mice were sacrificed 24 h later. Blood samples were obtained, and serum of each sample was separated for IgE determination. Bronchoalveolar lavage fluid (BALF) and lung tissues were collected to measure IL-5 and IL-13 levels. Nuclear extracts of lung tissues were employed to assess nuclear factor kappa B (NF-κB) p65 binding activity. RESULTS: Enhanced Pause (Penh) values were significantly increased in ovalbumin-sensitized and challenged mice (p < 0.01). Administration of fenofibrate (10 and 30 mg/kg) resulted in improved pulmonary function as shown by significantly lower Penh values (p < 0.01). Interleukin (IL) - 5 and IL-13 levels in BALF and lung tissues and immunoglobulin E (IgE) levels in serum were significantly elevated in the allergic mice. IL-5 levels in the lung tissues of mice treated with 1 mg/kg fenofibrate (FEN1) group were significantly reduced (p < 0.01). BALF and lung tissue IL-5 and IL-13 levels in mice treated with 10 and 30 mg/kg fenofibrate, FEN10 and FEN30, respectively, were significantly diminished when compared to the ovalbumin-treated (OVA) group, whereas treatment with 1 mg/kg fenofibrate resulted in insignificant changes. IgE levels in the serum of FEN30 group mice have shown a prominent reduction (p < 0.01). NF-κB p65 binding activity was higher in mice sensitized and challenged with ovalbumin (p < 0.01). NF-κB p65 binding activity was significantly reduced in allergic mice treated with 30 mg/kg (p < 0.01) fenofibrate. CONCLUSIONS: In this study, we showed that administration of 10 and 30 mg/kg fenofibrate effectively attenuated airway hyperresponsiveness and inflammation in a mouse model of allergic asthma, possibly through inhibition of NF-κB binding activity.

Endocannabinoid modulation of allergic responses: Focus on the control of FcεRI-mediated mast cell activation.

Allergic reactions are highly prevalent pathologies initiated by the production of IgE antibodies against harmless antigens (allergens) and the activation of the high-affinity IgE receptor (FcεRI) expressed in the surface of basophils and mast cells (MCs). Research on the mechanisms of negative control of those exacerbated inflammatory reactions has been intense in recent years. Endocannabinoids (eCBs) show important regulatory effects on MC-mediated immune responses, mainly inhibiting the production of pro-inflammatory mediators. However, the description of the molecular mechanisms involved in eCB control of MC activation is far from complete. In this review, we aim to summarize the available information regarding the role of eCBs in the modulation of FcεRI-dependent activation of that cell type, emphasizing the description of the eCB system and the existence of some of its elements in MCs. Unique characteristics of the eCB system and cannabinoid receptors (CBRs) localization and signaling in MCs are mentioned. The described and putative points of cross-talk between CBRs and FcεRI signaling cascades are also presented. Finally, we discuss some important considerations in the study of the effects of eCBs in MCs and the perspectives in the field.

Bufotalin ameliorates ovalbumin-induced allergic rhinitis by restoring the Tregs.

BACKGROUND: Allergic rhinitis (AR) is one of the most common inflammatory diseases. IgE, inflammatory cytokine production and Th17/Tregs imbalance have been implicated in AR pathogenesis. Bufotalin, a component extracted from toad venom skin secretions and auricular glands, has anti-inflammatory activity and regulates Th17/Tregs balance. Here, the effects of bufotalin on AR were explored. METHODS: The AR mice model was established using ovalbumin (OVA). AR mice were treated with bufotalin started on Day 22 with various doses (1, 10, 100 µg or 1 mg per mouse) every day to Day 30. The sneezing and rubbing frequencies were counted. Serum levels of IL-1ß, IL-10 and OVA-specific IgE were measured. The superficial cervical lymph nodes were harvested and the percentage of Tregs in lymph node was determined using CD4 and Foxp3 markers. RESULTS: OVA treatment successfully induced AR model in mice with significantly increased sneezing and rubbing frequency, elevated levels of serum histamine, IL-1ß, IL-10 and OVA-specific IgE. Bufotalin treatment significantly ameliorated AR symptoms, with reduced histamine, IgE and IL-1ß levels, as well as sneezing and rubbing frequency. Moreover, bufotalin treatment decreased the serum levels of IL-1ß, IL-10 and OVA-specific IgE in AR mice. CONLCUSION: Bufotalin ameliorated allergic rhinitis symptoms in AR mice by restoring Tregs in lymph node.

The role of calcium-sensitive receptor in ovalbumin-induced airway inflammation and hyperresponsiveness in juvenile mice with asthma.

The role of the calcium-sensitive receptor (CaSR) was assessed in a juvenile mouse model of asthma induced by ovalbumin (OVA). The experiment was divided into normal control, OVA, and OVA +2.5/5 mg/kg NPS2143 (a CaSR antagonist) groups. OVA induction was performed in all groups except the normal control, followed by assessing airway hyperresponsiveness (AHR) and lung pathological changes. Serum OVA-specific IgE and IgG1 were detected with an enzyme-linked immunosorbent assay (ELISA), and inflammatory cells were counted in bronchoalveolar lavage fluid (BALF). Real-time quantitative polymerase chain reaction, ELISA, and western blotting were performed to detect gene and protein expression. NPS2143 improved the OVA-induced AHR in mice, and AHR was higher in the OVA +2.5 mg/kg NPS2143 group than in the OVA +5 mg/kg NPS2143 group. Furthermore, NPS2143 reduced the production of OVA-specific IgE and IgG1 in serum and the number of eosinophils and lymphocytes in BALF in OVA mice with reduced CaSR expression in lung tissues. Besides, OVA-induced mice exhibited peribronchial and perivascular inflammatory cell infiltration, which was accompanied by severe goblet cell hyperplasia/hyperplasia and airway mucus hypersecretion. Furthermore, these mice exhibited increased levels of Interleukin (IL)-5, IL-13, MCP-1, and eotaxin, which were alleviated by NPS2143. The 5 mg/kg NPS2143 showed more effective than the 2.5 mg/kg treatment. CaSR expression was elevated in the lung tissues of OVA-induced asthmatic juvenile mice, whereas the CaSR antagonist NPS2143 reduced AHR and attenuated the inflammatory response in OVA-induced juvenile mice, possibly exerting therapeutic effects on childhood asthma.

Pulmonary Delivery of Levamisole Nanoparticles as an Immunomodulator Affecting Th and a Potential ADAM10 Inhibitor to Ameliorate Severe Allergic Asthma.

Asthma is a common chronic lung disease without absolute treatment, and hypersensitivity reactions and type 2 immune responses are responsible for asthma pathophysiology. ADAM10 as a metalloproteinase transmembrane protein is critical for development of Th2 responses, and levamisole as an anthelmintic drug has immunomodulatory effects, which not only regulates ADAM10 activity but also can suppress the bone marrow and neutrophil production. Therefore, in the present study, nanoparticles were used as a levamisole delivery system to reduce bone marrow suppression, and the immunomodulatory and ADAM10 inhibitory effects of levamisole were studied in allergic asthma. Asthmatic mice were treated with PLGA-levamisole nanoparticles. Then, AHR, BALF, and blood cell counts, levels of the IgG1 subclass, total and OVA-specific IgE, IL2, IL-4, IL-5, IL-10, IL-13, IL-17, IL-25, IL-33, INF-γ, and TNF-α, gene expression of FoxP3, T-bet, RORγt, PU.1, GATA3, FcεRII, CysLT1R, eotaxin, and ADAM10, and lung histopathology were evaluated. PLGA-LMHCl with considered characteristics could control airway hyper-responsiveness, eosinophils in the BALF, levels of immunoglobulins, Th2-, Th9-, and Th17-derived cytokines and pivotal genes, eosinophilic inflammation, hyperplasia of the goblet cell, and hyperproduction of mucus and could increase Th1- and Treg-derived cytokines and also pivotal genes. It could also modulate the ADAM10 activity and had no effect on the number of neutrophils in the bloodstream. The novel safe nanodrug had no side effect on the bone marrow to produce neutrophils and could control the allegro-immuno-inflammatory response of asthma.

Regulatory effects of miRNA-126 on Th cell differentiation and cytokine expression in allergic rhinitis.

BACKGROUND: Allergic rhinitis (AR) is a common disease worldwide. Imbalances in T helper (Th) cell differentiation and the dysregulation of related cytokines form the immunological basis of AR. miR-126 may play an important regulatory role in AR as a new marker and predictor of the disease. Therefore, the aim of this study was to explore the regulatory effects of miR-126 on Th cell differentiation and cytokine expression in AR. METHODS: T lymphocytes and rat models were transfected with a miR-126 mimic and an inhibitor. The expression of miR-126 and Th cell-related cytokines was detected by RT-qPCR and western blotting. The serum IgE levels were detected using ELISA. In the nasal mucosa, pathological changes were observed by HE staining, protein expression was detected by immunohistochemistry, and the differentiation ratio of Th cell subsets was detected by flow cytometry. RESULTS: During the occurrence and development of AR, the expression of miR-126 and the IgE levels were increased in the AR group. The number of Treg cell subsets decreased in the AR rats, increased after the miR-126 agomir intervention and decreased after miR-126 antagomir intervention. The number of Th1 and Th2 cell subsets increased in the AR rats, decreased after miR-126 agomir intervention and increased after the miR-126 antagomir intervention. CONCLUSION: We propose that miR-126 may be involved in the pathogenesis of AR by positively regulating the expression of Treg cytokines and negatively regulating the expression of the Th1 and Th2 cytokines.

A human IgE bispecific antibody shows potent cytotoxic capacity mediated by monocytes.

The generation of bispecific antibodies (bsAbs) targeting two different antigens opens a new level of specificity and, compared to mAbs, improved clinical efficacy in cancer therapy. Currently, the different strategies for development of bsAbs primarily focus on IgG isotypes. Nevertheless, in comparison to IgG isotypes, IgE has been shown to offer superior tumor control in preclinical models. Therefore, in order to combine the promising potential of IgE molecules with increased target selectivity of bsAbs, we developed dual tumor-associated antigen-targeting bispecific human IgE antibodies. As proof of principle, we used two different pairing approaches - knobs-into-holes and leucine zipper-mediated pairing. Our data show that both strategies were highly efficient in driving bispecific IgE formation, with no undesired pairings observed. Bispecific IgE antibodies also showed a dose-dependent binding to their target antigens, and cell bridging experiments demonstrated simultaneous binding of two different antigens. As antibodies mediate a major part of their effector functions through interaction with Fc receptors (FcRs) expressed on immune cells, we confirmed FcεR binding by inducing in vitro mast cell degranulation and demonstrating in vitro and in vivo monocyte-mediated cytotoxicity against target antigen-expressing Chinese hamster ovary cells. Moreover, we demonstrated that the IgE bsAb construct was significantly more efficient in mediating antibody-dependent cell toxicity than its IgG1 counterpart. In conclusion, we describe the successful development of first bispecific IgE antibodies with superior antibody-dependent cell toxicity-mediated cell killing in comparison to IgG bispecific antibodies. These findings highlight the relevance of IgE-based bispecific antibodies for clinical application.

Dietary scallop oil prepared from the internal organs of Japanese giant scallop (Patinopecten yessoensis) improved mite allergen-induced atopic dermatitis-like symptoms on the dorsal skin of NC/Nga mice.

This study aimed to investigate the effects of scallop oil (SCO) on atopic dermatitis (AD)-like symptoms induced by mite allergens in the dorsal and ear skins of NC/Nga mice compared to those of refined corn oil and krill oil (KO). SCO, rich in n-3 polyunsaturated fatty acids and phospholipids, was prepared from the internal organs of Japanese giant scallop, an underutilized fishery resource in Japan. Results showed that SCO intake improved AD-like symptoms, including ear edema, ear thickness, and transepidermal water loss of dorsal skin, and tended to decrease the scratching behavior, whereas KO intake did not. Further, SCO intake decreased the degranulated mast cell count and increased the tight junction protein claudin-1 expression, which is important for the barrier function, in the dorsal skin compared to refined corn oil intake. SCO improved the AD-like symptoms by suppressing mast cell degranulation and strengthening the barrier function of dorsal skin in NC/Nga mice.

Bronchodilatory effect of higenamine as antiallergic asthma treatment.

Asthma is a complex airway disease that affects more than 350 million humans worldwide. Allergic asthma symptoms are induced by Th2 immune response with the release of cytokines and allegro-inflammatory mediators that amplify the inflammatory response, airway hyper-responsiveness (AHR) and hyperproduction of mucus. Higenamine, as a chemical compound, is a ß2 adrenoreceptor agonist and can be used as bronchodilator in allergic asthma.BALB/c mice were allocated in four groups and then allergic asthma was induced in three groups. One of the asthmatic groups was treated with albuterol and other one was treated with higenamine. At least, methacholine challenge to determine the AHR, measurement of cytokines, total immunoglobulin E (IgE), LTB4 and LTC4 levels, evaluation of gene expression of Muc5ac, Muc5b, Agr2 and Arg1, and histopathological study were done.Higenamine treatment reduced AHR, interleukin (IL)-4, IL-13 levels, mRNA expression of MUC5ac, MUC5b, Arg1 and Agr2, goblet cell hyperplasia and mucus hypersecretion. Higenamine had no significant effect on IL-5, interferon-γ (INF-γ), IgE, LTB4, LTC4 levels and eosinophilic inflammation in lung tissue.Higenamine treatment controls asthma acute attack and breathlessness and can be used as asthma treatment with control of AHR and decrease of airflow obstruction and mucus hypersecretion and had allegro-immune-regulatory effect. But higenamine treatment had no notable effect on the inflammation and inflammatory factors.

IgE-Based Therapeutic Combination Enhances Antitumor Response in Preclinical Models of Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) represents 3% of all cancer cases and 7% of all cancer deaths in the United States. Late diagnosis and inadequate response to standard chemotherapies contribute to an unfavorable prognosis and an overall 5-year survival rate of less than 10% in PDAC. Despite recent advances in tumor immunology, tumor-induced immunosuppression attenuates the immunotherapy response in PDAC. To date, studies have focused on IgG-based therapeutic strategies in PDAC. With the recent interest in IgE-based therapies in multiple solid tumors, we explored the MUC1-targeted IgE potential against pancreatic cancer. Our study demonstrates the notable expression of FceRI (receptor for IgE antibody) in tumors from PDAC patients. Our study showed that administration of MUC1 targeted-IgE (mouse/human chimeric anti-MUC1.IgE) antibody at intermittent levels in combination with checkpoint inhibitor (anti-PD-L1) and TLR3 agonist (PolyICLC) induces a robust antitumor response that is dependent on NK and CD8 T cells in pancreatic tumor-bearing mice. Subsequently, our study showed that the antigen specificity of the IgE antibody plays a vital role in executing the antitumor response as nonspecific IgE, induced by ovalbumin (OVA), failed to restrict tumor growth in pancreatic tumor-bearing mice. Utilizing the OVA-induced allergic asthma-PDAC model, we demonstrate that allergic phenotype induced by OVA cannot restrain pancreatic tumor growth in orthotopic tumor-bearing mice. Together, our data demonstrate the novel tumor protective benefits of tumor antigen-specific IgE-based therapeutics in a preclinical model of pancreatic cancer, which can open new avenues for future clinical interventions.

COX-1 dependent biosynthesis of 15-hydroxyeicosatetraenoic acid in human mast cells.

15-hydroxyeicosatetraenoic acid (15-HETE) is an arachidonic acid derived lipid mediator which can originate both from 15-lipoxygenase (15-LOX) activity and cyclooxygenase (COX) activity. The enzymatic source determines the enantiomeric profile of the 15-HETE formed. 15-HETE is the most abundant arachidonic acid metabolite in the human lung and has been suggested to influence the pathophysiology of asthma. Mast cells are central effectors in asthma, but there are contradictory reports on whether 15-HETE originates from 15-LOX or COX in human mast cells. This prompted the current study where the pathway of 15-HETE biosynthesis was examined in three human mast cell models; the cell line LAD2, cord blood derived mast cells (CBMC) and tissue isolated human lung mast cells (HLMC). Levels and enantiomeric profiles of 15-HETE and levels of the downstream metabolite 15-KETE, were analyzed by UPLC-MS/MS after stimulation with anti-IgE or calcium ionophore A23187 in the presence and absence of inhibitors of COX isoenzymes. We found that 15-HETE was produced by COX-1 in human mast cells under these experimental conditions. Unexpectedly, chiral analysis showed that the 15(R) isomer was predominant and gradually accumulated, whereas the 15(S) isomer was metabolized by the 15-hydroxyprostaglandin dehydrogenase. We conclude that during physiological conditions, i.e., without addition of exogenous arachidonic acid, both enantiomers of 15-HETE are produced by COX-1 in human mast cells but that the 15(S) isomer is selectively depleted by undergoing further metabolism. The study highlights that 15-HETE cannot be used as an indicator of 15-LOX activity for cellular studies, unless chirality and sensitivity to pharmacologic inhibition is determined.

Effect of sex and genotype of the host on the anthelmintic efficacy of albendazole microcrystals, in the CBi-IGE Trichinella infection murine model.

Albendazole (ABZ) is an anthelmintic pharmaceutical commonly used in the treatment of nematode infections. It is a Class II drug poorly water-soluble, with very low bioavailability, a feature particularly limiting to treat the trichinellosis chronic phase. Microcrystals obtained by controlled precipitation using hydroxyethyl cellulose and chitosan have previously been shown to improve ABZ biopharmaceutical properties. This investigation aimed to test the systems' in vivo efficacy in the CBi-IGE murine model of Trichinella spiralis infection in the infection's different phases and parasite' stages. Treatment in the enteral phase led to a 90% decrease in the larval muscle load, probably due to its effect on T. spiralis female fecundity. Both microcrystal systems given in the migratory phase halved muscle load in males, a response not observed in females. The chitosan-based microcrystals proved to be the best when administered in the chronic phase of the infection ­ an increased proportion of L1 dead larvae was found compared to controls, except in CBi+-treated females. Males and females from the highly susceptible CBi+ line presented a significantly different treatment response in this phase. In vivo efficacy depended on the host genotype and sex and was related to the parasite cycle stage in which the formulations were administered.

Generation of a Human Allergic Mast Cell Phenotype from CD133+ Stem Cells.

Cultured human mast cells are a useful tool for research into innate immune responses as well as allergic mechanisms. Mast cells cultured from peripheral blood can provide information on immune mechanisms of known, selected individuals. With the method presented here, eight million mast cells can be cultured from ca. one million stem cells purified from one unit (450 mL) of human peripheral blood. Culture with IgE and IL4 optimizes an allergic phenotype of the mast cells.

5-Bromo-3,4-dihydroxybenzaldehyde from Polysiphonia morrowii attenuate IgE/BSA-stimulated mast cell activation and passive cutaneous anaphylaxis in mice.

The present study investigates the anti-allergic activity of the marine algal bromophenol, 3-bromo-4,5-dihydroxybenzaldehyde (BDB), isolated from Polysiphonia morrowii Harvey in immunoglobulin (Ig)E/bovine serum albumin (BSA)-stimulated mouse bone marrow-derived cultured mast cells (BMCMCs) and a passive cutaneous anaphylaxis (PCA) mice ear model. BDB effectively inhibited ß-hexosaminidase release (IC50 = 80.12 µM), in IgE/BSA-stimulated BMCMCs without a cytotoxic response. Also, BDB down-regulated the expression or secretion of cytokines, interleukin (IL)-1ß, IL-4, IL-5, IL-6, IL-10, IL-13, interferon (IFN)-γ, and tumor necrosis factor (TNF)-α and the chemokine (thymus and activation-regulated chemokine (TARC). The above effects could be attributed to the dose-dependent decrease of FcεRI expression on the surface of BMCMCs and its stable IgE binding. Moreover, BDB suppressed the nuclear factor (NF)-κB and spleen tyrosine kinase (SYK)-linker for T-cell activation (LAT)-GRB2 associated binding protein 2 (Gab2) signaling axis activated by IgE/BSA stimulation. Furthermore, oral administration of BDB to IgE-sensitized mice effectively attenuated IgE-triggered PCA reaction. Collectively, the anti-allergic effects of BDB suggest its potential applicability as a candidate for in-depth test trials.

Sialylation of immunoglobulin E is a determinant of allergic pathogenicity.

Approximately one-third of the world's population suffers from allergies1. Exposure to allergens crosslinks immunoglobulin E (IgE) antibodies that are bound to mast cells and basophils, triggering the release of inflammatory mediators, including histamine2. Although IgE is absolutely required for allergies, it is not understood why total and allergen-specific IgE concentrations do not reproducibly correlate with allergic disease3-5. It is well-established that glycosylation of IgG dictates its effector function and has disease-specific patterns. However, whether IgE glycans differ in disease states or affect biological activity is completely unknown6. Here we perform an unbiased examination of glycosylation patterns of total IgE from individuals with a peanut allergy and from non-atopic individuals without allergies. Our analysis reveals an increase in sialic acid content on total IgE from individuals with a peanut allergy compared with non-atopic individuals. Removal of sialic acid from IgE attenuates effector-cell degranulation and anaphylaxis in several functional models of allergic disease. Therapeutic interventions-including removing sialic acid from cell-bound IgE with a neuraminidase enzyme targeted towards the IgE receptor FcεRI, and administering asialylated IgE-markedly reduce anaphylaxis. Together, these results establish IgE glycosylation, and specifically sialylation, as an important regulator of allergic disease.

TGF-ß secreted by human umbilical cord blood-derived mesenchymal stem cells ameliorates atopic dermatitis by inhibiting secretion of TNF-α and IgE.

Human mesenchymal stem cells (MSCs) are promising therapeutics for autoimmune diseases due to their immunomodulatory effects. In particular, human umbilical cord blood-derived MSCs (hUCB-MSCs) have a prominent therapeutic effect on atopic dermatitis (AD). However, the underlying mechanism is unclear. This study investigated the role of transforming growth factor-beta (TGF-ß) in the therapeutic effect of hUCB-MSCs on AD. Small interfering RNA (siRNA)-mediated depletion of TGF-ß disrupted the therapeutic effect of hUCB-MSCs in a mouse model of AD by attenuating the beneficial changes in histopathology, mast cell infiltration, tumor necrosis factor-alpha (TNF-α) expression, and the serum IgE level. To confirm that hUCB-MSCs regulate secretion of TNF-α, we investigated whether they inhibit TNF-α secretion by activated LAD2 cells. Coculture with hUCB-MSCs significantly inhibited secretion of TNF-α by LAD2 cells. However, this effect was abolished by siRNA-mediated depletion of TGF-ß in hUCB-MSCs. TNF-α expression in activated LAD2 cells was regulated by the extracellular signal-related kinase signaling pathway and was suppressed by TGF-ß secreted from hUCB-MSCs. In addition, TGF-ß secreted by hUCB-MSCs inhibited maturation of B cells. Taken together, our findings suggest that TGF-ß plays a key role in the therapeutic effect of hUCB-MSCs on AD by regulating TNF-α in mast cells and maturation of B cells.