Evaluation on two types of paramyosin vaccines for the control of Haemaphysalis longicornis infestations in rabbits.

BACKGROUND: Haemaphysalis longicornis is an obligate hematophagous ectoparasite that transmits a variety of pathogens causing life-threatening diseases in humans and animals. Paramyosin (Pmy) is not only an invertebrate-specific myofibrillar protein but also an important immunomodulatory protein. Therefore, it is one of the ideal candidate antigens for vaccines. METHODS: We conducted two vaccine trials to evaluate the protective efficacy of Pmy recombinant protein (rPmy) and peptide vaccine (KLH-LEE). Each rabbit was immunized with three doses of rPmy or KLH-LEE adjuvanted with Freund's complete/incomplete at 500 µg/dose at 2-week intervals before challenge with 40 female H. longicornis/rabbit. PBS plus adjuvant, Trx or KLH was used as control group. The antibodies of rabbits were detected by ELISA. Then, female ticks were fed on the rabbits until detachment. RESULTS: ELISA results showed that both vaccines induced rabbits to produce antibodies. Compared with the Trx group, the engorgement weight, oviposition and hatchability of the rPmy group decreased by 8.87%, 26.83% and 38.86%, respectively. On the other hand, engorgement weight, oviposition and hatchability of female ticks in the KLH-LEE group correspondingly resulted in 27.03%, 53.15% and 38.40% reduction compared with that of the KLH group. Considering the cumulative effect of vaccination on the evaluated parameters, results showed 60.37% efficacy of the rPmy vaccine formulation and 70.86% efficacy in the KLH-LEE group. CONCLUSIONS: Pmy and particularly epitope LEE have potential for further development of an effective candidate vaccine to protect the host against tick infection. GRAPHIC ABSTARCT.

Evaluation of sensitization to the crude extract of Dermatophagoides farinae and its derived allergens, Der f 2 and Zen 1, in dogs with atopic dermatitis in Southern Brazil.

BACKGROUND: Atopic dermatitis is associated with the production of IgE antibodies against environmental allergens and allergens of the house dust miteDermatophagoides farinae are frequently implicated in the disease. OBJECTIVES: We aimed to observe the allergen-specific IgE against crudeD. farinae, Der f 2 and Zen 1 in dogs with atopic dermatitis and report if these dogs are in contact with material that could shelter mite allergens. METHODS: 100 dogs with clinical diagnosis of atopic dermatitis were included after exclusion of other forms of pruritic skin disease and dogs that already received specific or non-specific immunotherapy. These dogs were of different breeds and ages and they were presented at a veterinary teaching hospital and a private service of veterinary dermatology, both located in Curitiba, Southern Brazil. At the time of anamnesis, some questions were applied to know the possibility of these dogs having had contact with furniture and textile material which could shelter house dust mites. Sera samples were obtained and further analyzed by ELISA assay to measure serum IgE levels against these allergens with an established cut-off of 0.200 IgE optical density. RESULTS: The allergen-specific IgE positivity against crudeD. farinae (92 %) and Zen 1 (77 %) was higher than Der f 2 (56 %). There was a correlation in sensitization to crude D. farinae and Zen 1 that was not observed between crude D. farinae and Der f 2 and Der f 2 and Zen 1. The sensitization to D. farinae and its allergens was associated with an unrestricted exposition to furniture and textile material. CONCLUSION & CLINICAL RELEVANCE: dogs with atopic dermatitis are frequently sensitized to D. farinae and its allergens, Der f 2 and Zen 1, may be considered major allergens in these dogs. Zen 1 may be the main allergen responsible for the sensitization to crude D. farinae.

Platelets Independently Recruit into Asthmatic Lungs and Models of Allergic Inflammation via CCR3.

Platelet activation and pulmonary recruitment occur in patients with asthma and in animal models of allergic asthma, in which leukocyte infiltration, airway remodeling, and hyperresponsiveness are suppressed by experimental platelet depletion. These observations suggest the importance of platelets to various characteristics of allergic disease, but the mechanisms of platelet migration and location are not understood. The aim of this study was to assess the mechanism of platelet recruitment to extravascular compartments of lungs from patients with asthma and after allergen challenge in mice sensitized to house dust mite (HDM) extract (contains the DerP1 [Dermatophagoides pteronyssinus extract peptidase 1] allergen); in addition, we assessed the role of chemokines in this process. Lung sections were immunohistochemically stained for CD42b+ platelets. Intravital microscopy in allergic mice was used to visualize platelets tagged with an anti-mouse CD49b-PE (phycoerythrin) antibody. Platelet-endothelial interactions were measured in response to HDM (DerP1) exposure in the presence of antagonists to CCR3, CCR4, and CXCR4. Extravascular CD42b+ platelets were detected in the epithelium and submucosa in bronchial biopsy specimens taken from subjects with steroid-naive mild asthma. Platelets were significantly raised in the lung parenchyma from patients with fatal asthma compared with postmortem control-lung tissue. Furthermore, in DerP1-sensitized mice, subsequent HDM exposure induced endothelial rolling, endothelial adhesion, and recruitment of platelets into airway walls, compared with sham-sensitized mice, via a CCR3-dependent mechanism in the absence of aggregation or interactions with leukocytes. Localization of singular, nonaggregated platelets occurs in lungs of patients with asthma. In allergic mice, platelet recruitment occurs via recognized vascular adhesive and migratory events, independently of leukocytes via a CCR3-dependent mechanism.

Subolesin vaccination inhibits blood feeding and reproduction of Haemaphysalis longicornis in rabbits.

BACKGROUND: Ticks can transmit numerous tick-borne pathogens and cause a huge economic loss to the livestock industry. Tick vaccines can contribute to the prevention of tick-borne diseases by inhibiting tick infestation or reproduction. Subolesin is an antigenic molecule proven to be a potential tick vaccine against different tick species and even some tick-borne pathogens. However, its effectivity has not been verified in Haemaphysalis longicornis, which is a widely distributed tick species, especially in East Asian countries. Therefore, the purpose of this study was to evaluate the effectivity of subolesin vaccination against H. longicornis in a rabbit model. METHODS: Haemaphysalis longicornis (Okayama strain, female, adult, parthenogenetic strain) and Japanese white rabbits were used as the model tick and animal, respectively. The whole open reading frame of H. longicornis subolesin (HlSu) was identified and expressed as a recombinant protein using E. coli. The expression was verified using sodium dodecyl sulfate polyacrylamide gel electrophoresis, and the immunogenicity of rHlSu against anti-H. longicornis rabbit serum was confirmed using Western blotting. After vaccination of rHlSu in rabbits, experimental infestation of H. longicornis was performed. Variables related to blood-feeding periods, pre-oviposition periods, body weight at engorgement, egg mass, egg mass to body weight ratio, and egg-hatching periods were measured to evaluate the effectiveness of subolesin vaccination. RESULTS: The whole open reading frame of HlSu was 540 bp, and it was expressed as a recombinant protein. Vaccination with rHlSu stimulated an immune response in rabbits. In the rHlSu-vaccinated group, body weight at engorgement, egg mass, and egg mass to body weight ratio were statistically significantly lower than those in the control group. Besides, egg-hatching periods were extended significantly. Blood-feeding periods and pre-oviposition periods were not different between the two groups. In total, the calculated vaccine efficacy was 37.4%. CONCLUSIONS: Vaccination of rabbits with rHlSu significantly affected the blood-feeding and reproduction in H. longicornis. Combined with findings from previous studies, our findings suggest subolesin has the potential to be used as a universal tick vaccine.

Association between biomarkers and house dust mite sublingual immunotherapy in allergic asthma.

BACKGROUND: House dust mite (HDM) sublingual immunotherapy (SLIT) has demonstrated efficacy in clinical trials of patients with asthma. Airway inflammation is a characteristic of respiratory allergy, but its relationship to SLIT remains unclear. OBJECTIVE: We evaluate the association between clinical outcomes with pulmonary function and biomarkers in before and after HDM SLIT (UMIN Number 000022390). METHODS: One hundred twelve patients with asthma sensitized to HDM were randomized to add-on 6 standardized quality (SQ)-HDM SLIT to pharmacotherapy or pharmacotherapy alone for 48 weeks. At baseline and end of study, biomarkers, blood eosinophils, serum IgE, serum periostin, fractional exhaled nitric oxide (FeNO), and spirometry and clinical symptoms were measured. Association between biomarkers and an increase in FEV1 of 120 mL or greater were analysed. RESULTS: Sublingual immunotherapy (SLIT) demonstrated a significant reduction of serum periostin (P < .001), FeNO (P < .01), and increase in HDM-specific IgE (P < .05), FEV1 (P < .001) and improvement of clinical symptom scores, when compared to pharmacotherapy. The change in FEV1 correlated with the changes in serum periostin (r = .696, P < .001) and the changes in FeNO (r = .682, P < .001). The independent predictor of improvement in airflow limitation was changed in serum periostin (r2  = .753, P = .013) and FeNO (P = .038). Based on cut-off values derived by receiver operating characteristic analysis (periostin 30.9 ng/mL, FeNO 28.0 ppb), patients were distinguished responders from non-responders, but with no predictive value for blood eosinophils or total IgE. The proportion of patients with both high periostin and FeNO levels was significantly higher in responder than in non-responder (P = .026). CONCLUSIONS AND CLINICAL RELEVANCE: Adding HDM SLIT to pharmacotherapy resulted in reduced serum periostin and FeNO, and improved pulmonary function. Serum periostin and FeNO may be useful biomarkers for prediction of SLIT.

Tyrophagus putrescentiae group 4 allergen allergenicity and epitope prediction.

INTRODUCTION AND OBJECTIVES: Allergen-specific immunotherapy (ASIT) is the only allergic disease-modifying therapy available for children and adults, and recombinant allergens are an interesting approach to improve allergy diagnosis and ASIT. Tyrophagus putrescentiae is a common storage mite that produces potent allergens. The aim of this study was to express and characterize recombinant group 4 allergen protein of T. putrescentiae (Tyr p 4), and to further investigate allergenicity and potential epitopes of Tyr p 4. MATERIALS AND METHODS: The cDNA encoding Tyr p 4 was generated by RT-PCR and subcloned into pET-28a(+) plasmid. The plasmid was then transformed into E. coli cells for expression. After purification by nickel affinity chromatography and identification by SDS-PAGE, recombinant Tyr p 4 protein was used for a skin prick test and an ELISA to determine the allergic response. RESULTS: Study participants' allergic response rate to Tyr p 4 protein was 13.3% (16/120). Eight B-cell epitopes and three T-cell epitopes of Tyr p 4 were predicted. CONCLUSIONS: Similar to group 4 allergens of other species of mite, allergenicity of Tyr p 4 is weak. The expression, characterization and epitope prediction of recombinant Tyr p 4 protein provide a foundation for further study of this allergen in the diagnosis and ASIT of storage mite allergy.

Novel Mutant Phospholipase D from Hemiscorpius lepturus Acts as A Highly Immunogen in BALB/c Mice Against the Lethality of Scorpion Venom.

Hemiscorpius lepturus (H. lepturus) which belongs to the Scorpionidae family, is the deadliest scorpion in Iran. It causes pathological manifestations like dermonecrosis, hemolysis, renal failure, necrotic ulcers, and in some cases, even death. The venom of this scorpion is well-known for its cytotoxic effects in comparison with the other venomous scorpions which show significant neurotoxic effects. Due to the painless nature of the sting of this scorpion, the clinical symptoms occur in victims 24 to 72 h post-sting. In our previous studies during the last decade, we demonstrated that the medical complications are attributable to the presence of phospholipase D (PLD) as a major toxin in the venom. With the purpose of designing and constructing a vaccine against H. lepturus for humans, animal model experiments were performed. To achieve this goal, non-toxic PLD was developed by mutation of two critical catalytic residues-His12 and His48-into alanines and the product was then denominated mut-rPLD1. The in-vivo tests showed that the mice immunized with interval doses of 10 µg of mut-rPLD1, were completely protected against 10× the LD100 of the venom. In conclusion, this mutant may be an effective vaccine candidate against scorpion envenomation by H. lepturus in future clinical studies.

A role for early oral exposure to house dust mite allergens through breast milk in IgE-mediated food allergy susceptibility.

BACKGROUND: Successful prevention of food allergy requires the identification of the factors adversely affecting the capacity to develop oral tolerance to food antigen in early life. OBJECTIVES: This study sought to determine whether oral exposure to Dermatophagoides pteronyssinus through breast milk affects gut mucosal immunity with long-term effects on IgE-mediated food allergy susceptibility. METHODS: Gut immunity was explored in 2-week-old mice breast-fed by mothers exposed to D pteronyssinus, protease-inactivated D pteronyssinus, or to PBS during lactation. We further analyzed oral tolerance to a bystander food allergen, ovalbumin (OVA). In a proof-of-concept study, Der p 1 and OVA levels were determined in 100 human breast milk samples and the association with prevalence of IgE-mediated egg allergy at 1 year was assessed. RESULTS: Increased permeability, IL-33 levels, type 2 innate lymphoid cell activation, and Th2 cell differentiation were found in gut mucosa of mice nursed by mothers exposed to D pteronyssinus compared with PBS. This pro-Th2 gut mucosal environment inhibited the induction of antigen-specific FoxP3 regulatory T cells and the prevention of food allergy by OVA exposure through breast milk. In contrast, protease-inactivated D pteronyssinus had no effect on offspring gut mucosal immunity. Based on the presence of Der p 1 and/or OVA in human breast milk, we identified groups of lactating mothers, which mirror the ones found in mice to be responsible for different egg allergy risk. CONCLUSIONS: This study highlights an unpredicted potential risk factor for the development of food allergy, that is, D pteronyssinus allergens in breast milk, which disrupt gut immune homeostasis and prevents oral tolerance induction to bystander food antigen through their protease activity.

Amblyomma sculptum Salivary Protease Inhibitors as Potential Anti-Tick Vaccines.

Amblyomma sculptum is the main tick associated with human bites in Brazil and the main vector of Rickettsia rickettsii, the causative agent of the most severe form of Brazilian spotted fever. Molecules produced in the salivary glands are directly related to feeding success and vector competence. In the present study, we identified sequences of A. sculptum salivary proteins that may be involved in hematophagy and selected three proteins that underwent functional characterization and evaluation as vaccine antigens. Among the three proteins selected, one contained a Kunitz_bovine pancreatic trypsin inhibitor domain (named AsKunitz) and the other two belonged to the 8.9 kDa and basic tail families of tick salivary proteins (named As8.9kDa and AsBasicTail). Expression of the messenger RNA (mRNA) encoding all three proteins was detected in the larvae, nymphs, and females at basal levels in unfed ticks and the expression levels increased after the start of feeding. Recombinant proteins rAs8.9kDa and rAsBasicTail inhibited the enzymatic activity of factor Xa, thrombin, and trypsin, whereas rAsKunitz inhibited only thrombin activity. All three recombinant proteins inhibited the hemolysis of both the classical and alternative pathways; this is the first description of tick members of the Kunitz and 8.9kDa families being inhibitors of the classical complement pathway. Mice immunization with recombinant proteins caused efficacies against A. sculptum females from 59.4% with rAsBasicTail immunization to more than 85% by immunization with rAsKunitz and rAs8.9kDa. The mortality of nymphs fed on immunized mice reached 70-100%. Therefore, all three proteins are potential antigens with the possibility of becoming a new tool in the control of A. sculptum.

Heat shock protein 70 from Litopenaeus vannamei (LvHSP70) is involved in the innate immune response against white spot syndrome virus (WSSV) infection.

White spot syndrome (WSS) caused by white spot syndrome virus (WSSV) is a severe infectious disease in shrimp aquaculture. To find effective therapeutics to control WSSV, it is indispensable to understand the innate immune responses of shrimp to WSSV infection. Previous report demonstrated that the Litopenaeus vannamei heat shock protein 70 (LvHSP70) could induce shrimp innate immunity against bacterial infection. Herein, we further investigate the role of LvHSP70 in anti-WSSV infection. The temporal expression of LvHSP70 was significantly upregulated 2.5- and 1.5-fold at 6 and 24 h post systemic WSSV infection suggesting that the LvHSP70 was a WSSV responsive gene. The recombinant protein of LvHSP70 (rLvHSP70) was produced in an Escherichia coli system and its effect in protection against WSSV infection was investigated. Intramuscularly injection of juvenile shrimp with 1 nmol of rLvHSP70 could significantly prolong 50% mortality of WSSV-infected shrimp from 3 days to 5 days as compared to the control group injected with bovine serum albumin (BSA). Consistently, the injection of rLvHSP70 resulted in 24-fold, 20-fold and 100-fold decrease in the viral copy number after 6, 12 and 24 h post injection, respectively, compared to the control shrimp injected with BSA. Interestingly, it was found that the rLvHSP70 enhanced the expression of the key gene in the prophenoloxidase (proPO) activating system, LvproPO, but reduced the expression of Lvcaspase2 and LvIAP in WSSV-infected shrimp. These results suggested that the LvHSP70 is an important molecule involved in antiviral defense in shrimp presumably via modulating the proPO system and apoptosis.

The Immune Activity of PT-Peptide Derived from Anti-Lipopolysaccharide Factor of the Swimming Crab Portunus trituberculatus Is Enhanced when Encapsulated in Milk-Derived Extracellular Vesicles.

PT-peptide is derived from the anti-lipopolysaccharide factor of the swimming crab Portunus trituberculatus. The peptide, consisting of 34 amino acids, contains a lipopolysaccharide binding domain. In this study, we investigated the effect of PT-peptide encapsulated in raw milk-derived extracellular vesicles (EVs), designated as EVs-PT peptide, on immune regulation. The results showed that raw milk-derived EVs efficaciously delivered the PT-peptide into monocytes and elevated immune activity, including reactive oxygen species level, superoxide anion production, and phagocytosis. PT-peptide and EVs-PT peptide also elevated the secretion of cytokines, such as interferon-γ, interleukin-6, and tumor necrosis factor-α in human monocytic THP-1 cells. These results suggest that the PT-peptide could be developed as an immune stimulator.

A New Topical Eye Drop Containing LyeTxI-b, A Synthetic Peptide Designed from A Lycosa erithrognata Venom Toxin, Was Effective to Treat Resistant Bacterial Keratitis.

Bacterial keratitis is an ocular infection that can lead to severe visual disability. Staphylococcus aureus is a major pathogen of the eye. We recently demonstrated the strong antimicrobial activity of LyeTxI-b, a synthetic peptide derived from a Lycosa erithrognatha toxin. Herein, we evaluated a topical formulation (eye drops) containing LyeTxI-b to treat resistant bacterial keratitis. Keratitis was induced with intrastromal injection of 4 × 105 cells (4 µL) in New Zealand female white rabbits. Minimum inhibitory concentration (MIC) and biofilm viability were determined. LyeTxI-b ocular toxicity was evaluated through chorioallantoic membrane and Draize tests. One drop of the formulation (LyeTxI-b 28.9 µmol/L +0.5% CMC in 0.9% NaCl) was instilled into each eye four times a day, for a week. Slit-lamp biomicroscopy analysis, corneal histopathological studies and cellular infiltrate quantification through myeloperoxidase (MPO) and N-acetylglucosaminidase (NAG) detection were performed. LyeTxI-b was very effective in the treatment of keratitis, with no signs of ocular toxicity. Planktonic bacteria MIC was 3.6 µmol/L and LyeTxI-b treatment reduced biofilm viability in 90%. LyeTxI-b eliminated bacteria and reduced inflammatory cellular activity in the eyes. Healthy and treated animals showed similar NAG and MPO levels. LyeTxI-b is a potent new drug to treat resistant bacterial keratitis, showing effective antimicrobial and anti-inflammatory activity.

Oral Vaccination With a Formulation Combining Rhipicephalus microplus Subolesin With Heat Inactivated Mycobacterium bovis Reduces Tick Infestations in Cattle.

Vaccines are an environmentally friendly alternative to acaracides for the control of tick infestations, to reduce the risk for tick-borne diseases affecting human and animal health worldwide, and to improve animal welfare and production. Subolesin (SUB, also known as 4D8) is the functional homolog of Akirin2 involved in the regulation of development and innate immune response, and a proven protective antigen for the control of ectoparasite infestations and pathogen infection. Oral vaccination combining protein antigens with immunostimulants has proven efficacy with increased host welfare and safety, but has not been used for the control of tick infestations. Here we describe the efficacy of oral vaccination with a formulation combining Rhipicephalus microplus SUB and heat inactivated Mycobacterium bovis (IV) on cattle tick infestations and fertility. The levels of IgG antibody titers against SUB and M. bovis P22, and the expression of selected immune response genes were determined and analyzed as possible correlates of protection. We demonstrated that oral immunization with the SUB+IV formulation resulted in 51% reduction in the number of female ticks and 30% reduction in fertility with an overall efficacy of 65% in the control of R. microplus infestations by considering the cumulative effect on reducing tick survival and fertility in cattle. The akr2, IL-1ß, and C3 mRNA levels together with antibody levels against SUB correlated with vaccine efficacy. The effect of the oral immunization with SUB+IV in cattle on tick survival and fertility is essential to reduce tick infestations, and extended previous results on the effect of R. microplus SUB for the control of cattle tick infestations. These results support the development of oral vaccines formulations for the control of tick infestations and the incidence of tick-borne diseases.

Platelet-targeted dual pathway antithrombotic inhibits thrombosis with preserved hemostasis.

Despite advances in antithrombotic therapy, the risk of recurrent coronary/cerebrovascular ischemia or venous thromboembolism remains high. Dual pathway antithrombotic blockade, using both antiplatelet and anticoagulant therapy, offers the promise of improved thrombotic protection; however, widespread adoption remains tempered by substantial risk of major bleeding. Here, we report a dual pathway therapeutic capable of site-specific targeting to activated platelets and therapeutic enrichment at the site of thrombus growth to allow reduced dosing without compromised antithrombotic efficacy. We engineered a recombinant fusion protein, SCE5-TAP, which consists of a single-chain antibody (SCE5) that targets and blocks the activated GPIIb/IIIa complex, and tick anticoagulant peptide (TAP), a potent direct inhibitor of activated factor X (FXa). SCE5-TAP demonstrated selective platelet targeting and inhibition of thrombosis in murine models of both carotid artery and inferior vena cava thrombosis, without a significant impact on hemostasis. Selective targeting to activated platelets provides an attractive strategy to achieve high antithrombotic efficacy with reduced risk of bleeding complications.

Subcutaneous immunotherapy with purified Der p1 and 2 suppresses type 2 immunity in a murine asthma model.

BACKGROUND: Allergen-specific immunotherapy can induce long-term suppression of allergic symptoms, reduce medication use, and prevent exacerbations of allergic rhinitis and asthma. Current treatment is based on crude allergen extracts, which contain immunostimulatory components such as ß-glucans, chitins, and endotoxin. Use of purified or recombinant allergens might therefore increase efficacy of treatment. AIMS: Here, we test application of purified natural group 1 and 2 allergens from Dermatophagoides pteronyssinus (Der p) for subcutaneous immunotherapy (SCIT) treatment in a house dust mite (HDM)-driven mouse model of allergic asthma. MATERIALS AND METHODS: HDM-sensitized mice received SCIT with crude HDM extract, a mixture of purified Der p1 and 2 (DerP1/2), or placebo. Upon challenges, we measured specific immunoglobulin responses, allergen-induced ear swelling response (ESR), airway hyperresponsiveness (AHR), and inflammation in bronchoalveolar lavage fluid (BAL) and lung tissue. RESULTS: ESR measurement shows suppression of early allergic response in HDM-SCIT- and DerP1/2-SCIT-treated mice. Both HDM-SCIT and DerP1/2-SCIT are able to suppress AHR and eosinophilic inflammation. In contrast, only DerP1/2-SCIT is able to significantly suppress type 2 cytokines in lung tissue and BAL fluid. Moreover, DerP1/2-SCIT treatment is uniquely able suppress CCL20 and showed a trend toward suppression of IL-33, CCL17 and eotaxin levels in lung tissue. DISCUSSION: Taken together, these data show that purified DerP1/2-SCIT is able to not only suppress AHR and inflammation, but also has superior activity toward suppression of Th2 cells and HDM-induced activation of lung structural cells including airway epithelium. CONCLUSIONS: We postulate that treatment with purified natural major allergens derived from HDM will likely increase clinical efficacy of SCIT.

Der f1 induces pyroptosis in human bronchial epithelia via the NLRP3 inflammasome.

Damage to the bronchial epithelium leads to persistent inflammation and airway remodelling in various respiratory diseases, such as asthma and chronic obstructive pulmonary disease. To date, the mechanisms underlying bronchial epithelial cell damage and death by common allergens remain largely unknown. The aim of the present study was to investigate Der f1, an allergen of Dermatophagoides farinae, which may result in the death of human bronchial epithelial cells (HBECs). Der f1 induces BECs to undergo the inflammatory cell death referred to as pyroptosis, induced by increasing lactate dehydrogenase release and propidium iodide penetration. Stimulation by Der f1 enhances interleukin (IL)­1ß cleavage and release, which is associated with caspase­1 activation. In addition, the NOD­like receptor family pyrin domain­containing 3 (NLRP3), is required for the activation of caspase­1 through increasing the formation of the inflammasome complex. Consistent with these findings, pre­treatment of HBECs with a caspase­1 inhibitor, or silencing of NLRP3 by siRNA transfection, reduced Der f1­mediated IL­1ß and pyroptosis. Therefore, the common allergen Der f1 was not only found to induce allergy, but also led to pyroptosis and IL­1ß secretion via the NLRP3­caspase­1 inflammasome in HBECs. This newly identified connection of the Der f1 allergen with BEC damage and inflammation may play an important role in the pathogenesis of asthma.

Long-term effects of adenotonsillectomy on serum-specific immunoglobulin E.

BackgroundThe biased immune reactions of the adenotonsillar tissues are not always reflected by the serum immunoglobulin E (IgE); thus, we hypothesize that the systemic atopic status may not be changed after the adenotonsillectomy (AT) in children.MethodsTwenty-five children with AT and 23 age-matched healthy children were enrolled into this study, and followed up for ~4 years. Nasal Symptoms Scores (NSS), Quality of Life Scores (QOLS), specific IgE (sIgE), cytokines, and inflammatory cell were documented in all the subjects before and after study.ResultsFourteen patients and three healthy controls had positive serum sIgE levels (>0.35 kU/l) at the study-start that was not changed by the study-end. Two patients and two sIgE-negative healthy controls showed the Dermatophagoidespteronyssinus sensitization at the study-end. NSS and QOLS showed significant improvement after the surgery in the sIgE-positive patients (P<0.05), whereas no significant changes were found in the sIgE-negative patients (P=1.00). In addition, the serum sIgE-negative patients showed significant increases in interleukin (IL)-4, IL-5, and IL-10 levels in the serum (P<0.001), although no significant differences were found post surgery (P=0.667, 0.408, and 0.714, respectively).ConclusionsOur study showed that AT did not affect the pediatric atopic status. The systemic atopy may be independent of the tonsillar and adenoid tissues in children.

Proof of concept of the preventive efficacy of high-dose recombinant mono-allergen immunotherapy in atopic dogs sensitized to the Dermatophagoides farinae allergen Der f 2.

BACKGROUND: Allergen immunotherapy is currently the only intervention proposed to specifically prevent clinical flares after allergen challenges. The low molecular weight Der f 2 (Df2) is a major allergen in Japanese dogs sensitized to Dermatophagoides farinae house dust mites. OBJECTIVES: Pilot, blinded, placebo-controlled experiment testing the efficacy of subcutaneous immunotherapy (SCIT) with high doses of recombinant Df2 conjugated to the maltotriose pullulan (rDf2-P). METHODS: Eight Maltese beagle atopic dogs were sensitized to rDf2 then randomized to SCIT with rDf2-P (six dogs) or placebo (two). The immunotherapy consisted of six weekly injections of increasing doses (0.1-10.0 µg) of rDf2-P followed by four monthly injections of 10 µg of this allergen. Epicutaneous rDf2 challenges, rDf2-specific IgE serology and intradermal reactivity, as well as serum cytokine level measurements, were performed throughout the study. RESULTS: Subcutaneous injections of placebo did not alter the cutaneous reactivity after rDf2 challenge, while that of the dogs treated with rDf2-P SCIT disappeared in five of six dogs (83%) and was reduced in one of six (17%). During SCIT maintenance, skin lesion scores were significantly lower in dogs receiving SCIT compared to those treated with placebo. This clinical improvement was accompanied by a concurrent, yet not significant, decrease in rDf2-specific IgE serology and immediate intradermal reactivity. Cytokine serum levels were inconclusive. There were no adverse events seen with rDf2-P SCIT. CONCLUSIONS AND CLINICAL IMPORTANCE: The new mono-allergen SCIT appears safe and effective for reducing skin lesions after allergen challenges; it deserves further testing in dogs with spontaneous atopic dermatitis.