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In this study, the effect of upsetting-extrusion-upsetting (UEU) on the microstructure of an Mg-9Gd-4Y-2Zn-0.5Zr alloy and on dynamic recrystallization (DRX) was investigated. The results showed that after UEU, DRX occurred in the Mg-9Gd-4Y-2Zn-0.5Zr alloy. Namely, the grain refinement at the cross-sectional edges of UEU specimens was significant, with the grain size being refined from 8.27 µm in the necking region to 6.57 µm. Grain refinement is mainly caused by DRX where the long-period stacking order (LPSO) phase stimulates DRX around the grain boundaries through a particle-stimulated nucleation mechanism. DRX grains play an important role in weakening the texture by offsetting the severely deformed structure of deformed grains.

People evaluated their own voices as sounding more attractive than others rated their voices (i.e., self-enhancement effect from the perspective of the rater, termed "SE_rater"), and people also rated their own voices as more attractive than the voices of others (i.e., self-enhancement effect from the perspective of the voice, termed "SE_voice"). The aim of the present study is to explore whether the gender context (i.e., same-sex and opposite-sex rating context) could influence the SE effect of voice attractiveness evaluation. Male and female participants were asked to rate the attractiveness of their own voices and other participants' voices, either in a same-sex session or an opposite-sex session. The results demonstrated both the SE_rater and SE_voice effect in the same-sex and opposite-sex contexts, for both male and female. More importantly, we found that the SE_rater for the male voices was significantly greater than that for the female voices in the same-sex context whereas no such difference was found in the opposite-sex context. In addition, the SE_voice effect in men was larger in the same-sex context than that in the opposite-sex context whereas the SE_voice in women was smaller in the same-sex context than that in the opposite-sex context. These findings indicated that the self-enhancement effect of vocal attractiveness was modulated by the gender context.

The aim of the present study was to determine whether the self-enhancement effect of voice attractiveness evaluation is due to general self-positivity bias and/or the familiarity effect. The participants were asked to rate the attractiveness of their own voice, a friend's voice and strangers' voices. In addition, a self-reference valence (SR-valence) task was adopted in the experiment. Significant self-enhancement effects in voice attractiveness ratings were demonstrated, regardless of whether the participants recognized their self-voice or not. However, the friend-enhancement effect was found in only those participants who successfully recognized their friend's voice. Moreover, a significant correlation was found between self-positivity bias in the SR-valence task and the self-enhancement effect (but not the friend-enhancement effect). Our findings suggest that both the familiarity effect and self-positivity bias account for the vocal self-enhancement effect, and the influence of self-positivity bias could be implicit. The present study thus provides empirical evidence to clarify the potential explanations for the self-enhancement of voice attractiveness assessment.

Aim: To develop IL-18 peptide-based virus-like particle vaccines that elicit autoantibodies against IL-18 and to evaluate the in vivo effects of the vaccines in murine colitis. Methods: Recombinant IL-18 vaccines were constructed, and the effects of the vaccines were evaluated in trinitrobenzene sulfonic acid-induced acute and chronic colitis in mice. Results: Two murine IL-18 peptide-based vaccines (A and D) were developed, which induced relative long-lasting specific antibodies against IL-18. Vaccine-immunized mouse antisera could partially block IL-18-induced IFN-γ production in vitro. Mice receiving vaccine D, not vaccine A, had a significant decrease in intestinal inflammation, collagen deposition and pro-inflammatory cytokine levels in colon tissue. Conclusion: IL-18 vaccine may provide a potential therapeutic approach in the treatment of Crohn’s disease., Lay abstract Many proinflammatory cytokines, including IL-18, play important roles in exaggerating the disease progression of inflammatory bowel disease (IBD). Inflammatory bowel disease is a chronic autoimmune disease, which usually requires long-term treatment. Blocking these proinflammatory cytokines by using monoclonal antibodies has shown certain clinical efficacy, but it requires repeated injection of these antibodies. To induce relative long-lasting antibodies, we developed IL-18 peptide-based virus-like particle vaccines and evaluated their therapeutic effects in a murine colitis model. Our results showed that immunization of mouse with IL-18 peptide-based vaccine could improve murine colitis, which indicated this vaccine strategy might be a potential treatment approach.

Background Allergic reactions to mosquito bites are an increasing clinical concern. Due to the lack of availability of mosquito salivary allergens, they are underdiagnosed. Here, we reported a newly cloned mosquito Aedes (Ae.) aegypti salivary allergen. Methods A cDNA encoding a 30-kDa Ae. aegypti salivary protein, designated Aed a 3, was isolated from an expression library. The full-length cDNA was cloned into a baculovirus expression vector, and recombinant Aed a 3 (rAed a 3) was expressed, purified, and characterized. Skin prick tests with purified rAed a 3 and Ae. aegypti bite tests were performed in 43 volunteers. Serum rAed a 3-specific IgE levels were measured in 28 volunteers. Results The primary nucleotide sequence, deduced amino acid sequence, and IgE-binding sites of Aed a 3 were identified. rAed a 3-selected antibodies recognized a 30-kDa Ae. aegypti saliva protein. rAed a 3 bound IgE in mosquito-allergic volunteers and the binding could be inhibited by the addition of natural mosquito extract dose dependently. Immediate skin test reactions to rAed a 3 correlated significantly with mosquito bite-induced reactions. Of the bite test-positive volunteers, 32% had a positive rAed a 3 skin test and 46% had specific IgE. No bite test-negative volunteers reacted to rAed a 3 in either the skin tests or the IgE assays, confirming the specificity of the assay. Conclusions Aed a 3 that corresponds to the Aegyptin protein is a major mosquito salivary allergen. Its recombinant form has biological activity and is suitable for use in skin tests and specific IgE assays in mosquito-allergic individuals.

Interleukin (IL)-12 and IL-23 that share subunit p40 are important cytokines in the pathogenesis of inflammatory bowel disease. We reported that mouse p40 peptide-based vaccines ameliorated intestinal inflammation in the prevention of trinitrobenzene sulfonic acid (TNBS)–induced murine colitis model. Here, we evaluated whether administration of the vaccine after establishment of colitis would be effective in modifying both TNBS-induced and dextran sulfate sodium (DSS)–induced chronic colitis and the underlying immune mechanisms. We further examined whether vaccination could exacerbate infections. Chronic colitis was developed by either intrarectally administrating TNBS or drinking 4% DSS water. Vaccination started after two TNBS administrations or 7 days of DSS treatment. Results showed that administrating p40 vaccine induced high tittered antibodies to IL-12 and IL-23, improved clinical scores, reduced intestinal inflammation and fibrosis, and down-regulated proinflammatory cytokine productions in colon tissue, compared with control mice. Furthermore, in lamina propria mononuclear cells and/or mesenteric lymph nodes, mice immunized with p40 peptide vaccine exhibited high ratios of Treg/Th1 and Treg/Th17 cells and increased IL-10 expression in CD11c(+)IL-10(+)cells. In mice infected with lung chlamydia, in which the protective role of Th1/Th17 is well documented, vaccine immunization did not increase lung bacterial burden. We conclude that p40 vaccine may provide a potential and safe approach for treatment of IBD.

Background: Accurate diagnosis of mosquito allergy has been hampered by the laborious task of obtaining mosquito salivary allergens. We have previously studied 3 recombinant (r) Aedes aegypti mosquito salivary allergens: rAed a 1, rAed a 2 and rAed a 3. Here, we report the expression, purification, identification and evaluation of rAed a 4, a 67-kDa α-glucosidase. Methods: rAed a 4 was expressed using a baculovirus/insect cell system, purified by a combination of anion- and cation-exchange chromatography, and identified by immunoblotting. A. aegypti saliva extract was prepared in our laboratory. An indirect enzyme-linked immunosorbent assay (ELISA) was developed to measure rAed a 4-specific immunoglobulin E (IgE) and IgG antibodies in sera from 13 individuals with a positive mosquito-bite test from a laboratory-reared mosquito. Sera from 18 individuals with a negative bite test served as controls. Results: Purified rAed a 4 bound to the IgE in mosquito-allergic sera, as detected by ELISA and immunoblotting. The binding of rAed a 4 to IgE could be inhibited in a dose-dependent manner by the addition of an A. aegypti extract. Mosquito-allergic individuals had significantly higher mean levels of rAed a 4-specific IgE and IgG than controls. Using the mean of the controls ± 2 SD as a cut-off level, 46% of the 13 allergic individuals had a positive IgE, while none of the controls was positive (p < 0.001). Conclusions: Aed a 4 is a major allergen in mosquito saliva. Its recombinant form has the hydrolase function and can be used for the diagnosis of mosquito allergy.

Myeloid-derived suppressor cells (MDSCs) are regulatory cell populations that have the ability to suppress effector T cell responses and promote the development of regulatory T cells (Tregs). They are a heterogeneous population of immature myeloid progenitors that include monocytic and granulocytic subsets. We postulated that given the rapid expansion of myeloid cells post-transplant, these members of the innate immune system may be important contributors to the early immune environment post-transplant. To evaluate the kinetics of recovery and function of MDSCs after allogeneic hematopoietic stem cell transplant (HSCT), 26 patients undergoing allogeneic HSCT were studied at 6 time points in the first 3 months after HSCT. Both MDSC subsets recovered between 2 and 4 weeks, well before the recovery of T and B lymphocytes. MDSC subset recovery positively correlated with T, B, and/or double-negative T cell numbers after HSCT. MDSCs isolated from patients post-transplant were functional in that they suppressed third-party CD4+ T cell proliferation and Th1 differentiation and promoted Treg development. In conclusion, functional MDSC are present early after HSCT and likely contribute to the regulatory cell population post-transplant.

Transforming growth factor beta1 (TGF-β1) plays an important role in hepatic fibrogenesis. In this study, we documented the effects of active immunization against TGF-β1 on hepatic fibrosis in an animal model of chronic liver disease. BALB/c mice were immunized against 3 different peptides of TGF-β1 ligated into hepatitis B virus core protein (HBVc). Titers of TGF-β1 antibodies were documented by enzyme linked immunoassays and antibody activity by cell membrane receptor binding and proliferation assays. The most immunogenic recombinant HBVc + TGF-β1 peptide (HBVc + C) then served as a vaccine in Sprague-Dawley rats with dimethylnitrosamine-induced chronic liver disease. Hepatic fibrosis was documented by serum hyaluronic acid levels, liver histology, and reverse transcriptase polymerase chain reaction for hepatic collagen I (α1) and smooth muscle alpha actin mRNA expression. Relative to control rats vaccinated with HBVc alone, recombinant HBVc + C vaccinated animals had significantly lower serum hyaluronic acid levels, less histologic evidence of hepatic fibrosis, and reduced expression of collagen I (α1) and smooth muscle alpha actin mRNA in the liver. The results of this proof-of-concept study suggest that active immunization against TGF-β1 is a worthwhile strategy to pursue in efforts to prevent hepatic fibrosis associated with chronic liver disease.

We sought to develop an IL-33 vaccine and evaluate its efficacy in a mouse model of asthma. The full-length molecules of putative mature IL-33 were inserted into the immunodominant epitope region of hepatitis B core antigen using gene recombination techniques. The expressed chimeric protein presented as virus-like particles (VLPs) under observation using an electron microscopy. To investigate immunization characteristics of the VLPs, mice were immunized by using different doses, adjuvants, and routes. The VLPs induced sustained and high titers of IL-33-specific IgG and IgA even without the use of a conventional adjuvant, and the lowered ratio of IgG1/IgG2a in vaccinated mice indicated a shift from Th2 to Th1-like responses. To assess the vaccine effects on blocking the signaling of IL-33/ST2 pathway, mice receiving 3 vaccinations subjected to intraperitoneal sensitization and intranasal challenge with ovalbumin (OVA). Control animals received carrier or PBS in place of the vaccine. Immunization with the VLPs significantly suppressed inflammatory cell number and IL-33 level in BALF. OVA -induced goblet cell hyperplasia and lung tissue inflammatory cell infiltration were significantly suppressed in vaccinated mice. Our data indicate that IL-33 molecule-based vaccine, which may block IL-33/ST2 signaling pathway on a persistent basis, holds potential for treatment of asthma and, by extension, other diseases where overexpressed IL-33 plays a pivotal role in pathogenesis.

Background: Overexpression of IL-23 has been implicated in the pathogenesis of Crohn’s disease. Using vaccines to block overexpressed endogenous cytokines has emerged as a new therapeutic strategy for the long-term treatment of the disease. Aim: We sought to develop peptide-based vaccines specific to IL-23 and evaluate their effects in colitis mice. Materials & methods: The vaccine was developed by inserting a peptide derived from mouse IL-23 p19 into the carrier protein, hepatitis B core antigen, using molecular engineering methods. One vaccine against IL-23 p19 was obtained that induced high-titered and long-lasting antibodies to IL-23 without the use of adjuvants. The inhibitory effect of vaccine-immunized serum was subsequently evaluated in vitro. To evaluate the in vivo effects, mice were subcutaneously injected with the vaccine, carrier or saline three times. Two weeks after the last injection, chronic colitis was induced in mice by seven weekly administrations with 2,4,6-trinitrobenzene sulfonic acid. Results:In vitro studies revealed that serum IL-23 p19-specific IgG significantly suppressed IL-23-induced IL-17 production by splenocytes. In vivo evaluation of the effect of the vaccine in mice with chronic colitis indicated that vaccine-immunized mice exhibited a decrease in colon inflammation, collagen deposition and levels of IL-23 and IL-12 cytokines, compared with control groups. Conclusion: IL-23 p19 vaccine is capable of downregulating inflammatory responses in chronic murine colitis, providing a novel therapeutic approach in Crohn’s disease.

We previously reported that a recombinant IL-13 peptide-based virus-like particle vaccine significantly suppressed murine acute airway allergic inflammatory responses. The impact of this strategy on the development of chronic airway inflammation and remodeling has not been investigated. We evaluated whether the vaccine-mediated sustained suppression of IL-13 attenuates features of chronic airway inflammation and remodeling in mice repeatedly challenged with allergen. BALB/c mice received two intraperitoneal sensitizing injections of ovalbumin (OVA) and alum, followed by six consecutive 2-day intranasal OVA challenges at 12-day intervals and then a 4-week recovery period. Anti-IL-13 antibodies were induced with a vaccine before (preventive experiments) or after (interventional experiments) the OVA challenge commenced. Respiratory mechanics were assessed using low-frequency forced oscillation with a small animal ventilator. Cytokine concentrations in bronchoalveolar lavage fluid (BALF) and lung histology were also assessed. In the preventive experiments, vaccination significantly suppressed IL-13 concentrations, the accumulation of inflammatory cells in BALF, lung mucus production, and collagen deposition. Furthermore, vaccination significantly attenuated OVA challenge-induced increases in airway resistance, tissue resistance, and tissue elastance, both acutely and after a 4-week recovery from allergen challenges. In the interventional experiments, vaccination decreased IL-13, TGF-β1, and IL-12p40 concentrations in BALF, as well as mucus production and collagen deposition. Chronic inflammation and sustained airway hyperresponsiveness were not significantly reversed. The persistent suppression of IL-13 with a vaccine inhibits chronic airway inflammation and the development of several key components of airway remodeling, and this intervention is more effective at early stages than later during chronic inflammation.

Accurate diagnosis of mosquito allergy has been hampered by the laborious task of obtaining mosquito salivary allergens. We have previously studied 3 recombinant (r) Aedes aegypti mosquito salivary allergens: rAed a 1, rAed a 2 and rAed a 3. Here, we report the expression, purification, identification and evaluation of rAed a 4, a 67-kDa α-glucosidase.rAed a 4 was expressed using a baculovirus/insect cell system, purified by a combination of anion- and cation-exchange chromatography, and identified by immunoblotting. A. aegypti saliva extract was prepared in our laboratory. An indirect enzyme-linked immunosorbent assay (ELISA) was developed to measure rAed a 4-specific immunoglobulin E (IgE) and IgG antibodies in sera from 13 individuals with a positive mosquito-bite test from a laboratory-reared mosquito. Sera from 18 individuals with a negative bite test served as controls.Purified rAed a 4 bound to the IgE in mosquito-allergic sera, as detected by ELISA and immunoblotting. The binding of rAed a 4 to IgE could be inhibited in a dose-dependent manner by the addition of an A. aegypti extract. Mosquito-allergic individuals had significantly higher mean levels of rAed a 4-specific IgE and IgG than controls. Using the mean of the controls ± 2 SD as a cut-off level, 46% of the 13 allergic individuals had a positive IgE, while none of the controls was positive (p0.001).Aed a 4 is a major allergen in mosquito saliva. Its recombinant form has the hydrolase function and can be used for the diagnosis of mosquito allergy.

Transforming growth factor (TGF)-β1 is involved in the processes of airway inflammation and remodeling; however, its reported roles in asthma pathogenesis are controversial. We sought both to investigate the effects of active immunization targeting TGF-β1 on allergen-induced airway inflammatory responses and to evaluate its possible application for asthma treatment. BALB/c mice were immunized with a virus-like-particle (VLP) vaccine presenting a TGF-β1 peptide. For the preventive intervention of acute allergic airway inflammation, immunization was conducted before sensitization and challenges with ovalbumin (OVA), and for the therapeutic treatment of chronic inflammatory responses, immunization was initiated after inflammatory responses were established. Preventive immunization with VLPs led to increased proinflammatory IL-4, IL-13, and IL-33 levels in the bronchoalveolar lavage fluids (BALF) with no significant effects on lung tissue inflammation and airway goblet cell hyperplasia. Therapeutic treatment showed that at 24 h after the fourth 2-day challenge with OVA following 2 intraperitoneal sensitizations, airway subepithelial collagen deposition was significantly ameliorated in vaccinated mice, whereas the lung histology and cytokine profile in the BALF were not changed. In contrast, after a 4-week recovery from the last OVA challenge, the vaccinated mice's collagen deposition remained reduced, but they sustained lung-tissue inflammation and goblet-cell hyperplasia; elevated IL-13, TNF, and IFN-γ levels in the BALF; and increased airway resistance, tissue resistance, and tissue elastance. In a conclusion, the role of TGF-β1 is complicated in allergic airway inflammatory responses. It is important to make a careful assessment in accordance with specific disease conditions when targeting TGF-β1 for a therapeutic purpose.

Interleukin (IL)-12 and IL-23 both share the p40 subunit and are key cytokines in the pathogenesis of Crohn’s disease. Previously, we have developed and identified three mouse p40 peptide-based and virus-like particle vaccines. Here, we evaluated the effects and immune mechanisms of the optimal vaccine in downregulating intestinal inflammation in murine acute and chronic colitis, induced by intrarectal administrations of trinitrobenzene sulfonic acid (TNBS). Mice were injected subcutaneously with vaccine, vaccine carrier or saline three times, and then intrarectally administered TNBS weekly for 2 wks (acute colitis) or 7 wks (chronic colitis). The severity of colitis was evaluated by body weight, histology and collagen and cytokine levels in colon tissue. Th1 and Th17 cells in mesenteric lymph nodes (MLN) were determined. Our results showed the vaccine induced high level and long-lasting specific IgG antibodies to p40, IL-12 and IL-23. After administrations of TNBS, vaccinated mice had significantly less body weight loss and a significant decrease of inflammatory scores, collagen deposition and expression of p40, IL-12, IL-23, IL-17, TNF, iNOS and Bcl-2 in colon tissues, compared with carrier and saline groups. Moreover, vaccinated mice exhibited a trend to lower percentages of Th1 cells in acute colitis and of Th17 cells in chronic colitis in MLN than in controls. In summary, administration of the vaccine induced specific antibodies to IL-12 and IL-23, which was associated with improvement of intestinal inflammation and fibrosis. This suggests that the vaccine may provide a potential approach for the long-term treatment of Crohn’s disease.

IL-10 is a regulatory cytokine that plays important roles in promoting disease progression in cutaneous leishmaniasis and suppressing allergic responses in asthma. We sought to develop an IL-10 peptide-based vaccine for the control of IL-10-related diseases. To break self-tolerance, hepatitis B core antigen (HBcAg) was used as a carrier. The vaccine was prepared by inserting a peptide derived from mouse IL-10 into the carrier using gene recombination methods. This vaccine presented as virus-like particles, bound to polyclonal anti-IL-10 antibodies, and induced high titers of IL-10-specific IgG. The in vivo effects of the vaccine were investigated in a murine model of cutaneous leishmaniasis. Unexpectedly, vaccinated mice developed larger cutaneous lesions, harbored significantly more parasites, and cells from lymph nodes produced higher amounts of parasite-specific IL-4, IL-10 and IFN-γ in cultures. Further in vitro studies showed that serum IL-10-specific IgG from vaccinated mice significantly enhanced IL-10 bioactivity dose-dependently. This enhancing effect was confirmed in OVA-induced asthmatic mice. Vaccinated mice exhibited a significant decrease in airway eosinophils, lung inflammation, goblet hyperplasia, and levels of serum OVA-specific IgE, compared to control mice. We concluded that the IL-10 vaccine enhances the bioactivity of IL-10 in vitro and in vivo , providing a potential therapeutic approach in diseases associated with insufficient IL-10 production.

Overexpression of IL-12 and IL-23, which share the p40 subunit, has been implicated in the pathogenesis of Crohn's disease. Targeting these cytokines with monoclonal antibodies has emerged as a new and effective therapy, but one with adverse reactions. In this study, we sought to develop p40 peptide-based virus-like particle vaccines that elicit autoantibodies to IL-12 and IL-23 for a long-term treatment of the disease. Three vaccines (named C, D and F) against the p40 were developed by inserting peptides derived from p40 into the carrier, hepatitis B core antigen, using molecular engineering methods. Immunization with the vaccines, without the use of adjuvants, induced high titered and long-lasting antibodies to IL-12, IL-23 and p40. The three vaccines were evaluated in vivo in 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced chronic murine colitis. Mice were immunized with a vaccine three times, followed by weekly intrarectal administrations of TNBS. Vaccine groups, especially groups C and F, showed reduced expression of IL-12/IL-23p40, less inflammation, and decreased collagen deposition in colon tissues when compared with controls. We concluded that IL-12/IL-23p40 vaccines may be a potential therapeutic approach in the treatment of Crohn's disease and other autoimmune diseases.

All-trans retinoic acid inhibits inflammatory responses of human ulcerative and murine colitis. IBD is characterized by uncontrolled immune responses in inflamed mucosa, with dominance of IL-17-producing cells and deficiency of Treg cells. The aim of this study was to explore the effect and mechanisms of RA, the ligand of RARα, on immune responses in human and murine colitis. Colonic biopsies from patients with UC were cultured and treated with RA as the agonist of RARα or LE135 as the antagonist of RARα. Expressions of IL-17 and FOXP3 were detected by immunohistochemistry. Murine colitis was induced by intrarectal administration with TNBS at Day 1. Mice were then i.p.-treated with RA or LE135 daily for 7 days. Cytokine levels in the cultures of mouse LPMCs were measured. Expressions of FOXP3 and IL-17 in colon tissues or MLN were detected by immunohistological analysis. Body weight and colon inflammation were evaluated. RA treatment up-regulated FOXP3 expression and down-regulated IL-17 expression in colon biopsies of patients and in colon tissues and MLN of mice with colitis compared with controls. LPMCs from RA-treated mice produced lower levels of proinflammatory cytokines (TNF-α, IL-1β, IL-17) but more regulatory cytokines (IL-10, TGF-β) compared with that of untreated mice. LE135 showed the opposite effect of RA. Furthermore, RA ameliorated TNBS-induced colitis in a dose-dependent manner, as seen by improved body weight and colon inflammation. RA down-regulates colon inflammatory responses in patients with IBD in vitro and in murine colitis in vivo, representing a potential therapeutic approach in IBD treatment.

Allergic diseases including asthma are characterized by an increase of serum IgE levels. Since IgE was discovered in 1966, it has been considered to be the most important biological target in the treatment of allergy and asthma. Indeed, recent studies reveal that IgE, through its high affinity IgE receptors (FcepsilonRI), is now considered a critical regulator of Th2 responses. This is supported by the great success of the anti-IgE monoclonal antibody (mAb) in the treatment of allergy and asthma. Nonetheless, adverse reactions such as anaphylaxis, urticaria and serum sickness have been reported with this therapy and repeated injections at extremely high costs are required to maintain effectiveness. To overcome these disadvantages, a new strategy using vaccines against IgE that may offer long-term efficacy with fewer adverse effects is being investigated. This article reviews IgE's role in allergy and asthma, currently used anti-IgE mAb omalizumab, and the advantages, types, effectiveness and development stages of vaccines against IgE. This review also discusses concerns with the vaccine strategy.

Allergic reactions to mosquito bites are caused by allergens in mosquito saliva. In this review, allergic reactions to mosquito salivary allergens, and characteristics of salivary allergens and their recombinant forms are described. The use of the recombinant allergens in the diagnosis of mosquito allergy is discussed.

Interleukin (IL)-13 plays a pivotal role in the pathogenesis of allergic asthma. Passive administration of its monoclonal antibody or soluble receptor to block overproduced IL-13 has been proven to be effective in controlling airway allergic responses in animal models, but these approaches have disadvantages of short half-lives, high costs, and possible adverse effects.We sought to develop a novel therapeutic strategy through constructing an IL-13 peptide-based vaccine for blocking IL-13 on a persistent effect basis and to evaluate its in vivo effects using a murine model.To break self-tolerance, truncated hepatitis B core antigen was used as a carrier. Vaccine was prepared by inserting a peptide derived from the receptor binding site of mouse IL-13 into the immunodominant epitope region of the carrier using gene recombination methods. Mice received vaccine subcutaneously three times, and then subjected to intraperitoneal sensitization and intranasal challenge with ovalbumin. Control animals received carrier or saline in place of vaccine.The vaccine presented as virus-like particles and induced sustained and high titered IL-13-specific IgG without the use of conventional adjuvant. Vaccination significantly suppressed ovalbumin-induced inflammatory cell number, and IL-13 and IL-5 levels in bronchoalveolar lavage fluids. Serum total and ovalbumin-specific IgE were also significantly inhibited. Moreover, allergen-induced goblet cell hyperplasia, lung tissue inflammatory cell infiltration, and pulmonary hyperresponsiveness to inhaled methacholine were significantly suppressed in vaccinated mice.Our data indicate that IL-13 peptide-based vaccines could be an effective therapeutic approach in the treatment of asthma.

Background: Mosquito salivary proteins cause allergic reactions in humans. The allergenicity, clinical relevance, and species cross-reactivity of a new 37-kDa recombinant mosquito (Aedes aegypti) salivary allergen, rAed a 2, were evaluated. Methods: rAed a 2 was expressed using a baculovirus/insect cell system and purified. Its allergenicity was examined using an enzyme-linked immunosorbent assay (ELISA), ELISA inhibition tests, immunoblots, and skin tests. Epicutaneous tests with the allergen, mosquito whole body extracts, and mosquito bite tests were performed on 48 volunteers. Serum rAed a 2-specific immunoglobulin E (IgE) was measured in individuals with positive mosquito saliva-specific IgE and negative controls. Results: Both immunoblots and ELISAs demonstrated that rAed a 2 bound to the IgE of mosquito-allergic individuals. The binding could be inhibited by the addition of a natural mosquito preparation. Furthermore, rAed a 2 induced immediate and delayed skin reactions. Ten per cent of 31 participants with a positive mosquito bite test had positive skin reactions to rAed a 2, compared with 32% for mosquito whole body extract. None of the participants with a negative bite test showed positive reactions to either of the two extracts. Forty-three per cent of individuals with positive saliva-IgE had positive rAed a 2-IgE. rAed a 2 was a species-shared allergen, being present in the saliva of the 11 species studied. Conclusions: rAed a 2 has identical antigenicity and biologic activity to its native form. It can be used in the diagnosis of mosquito allergy, and is an important species-shared antigen.

Mosquito bite-induced acute systemic allergic reactions are an increasing clinical concern and have not been optimally characterized immunologically.We wanted to study the immunologic basis of these reactions.Sera were received from 14 individuals with a history of acute systemic allergic reactions to mosquito bites, defined as the presence of one or more of the following: urticaria, angioedema, wheezing, dyspnea, hypotension, and decrease or loss of consciousness. Ten individuals were from the United States and one each was from Canada, Germany, Japan, and Switzerland. An indirect ELISA was developed to measure specific IgE and IgG antibodies to saliva from 5 common mosquito species with different geographic distributions: Aedes aegypti, Aedes vexans, Aedes albopictus, Anopheles sinensis, and Culex quinquefasciatus. Twenty-nine individuals with negative bite test results from laboratory-reared mosquitoes served as control subjects.Mosquito saliva-specific IgE levels to all 5 species were significantly increased in the individuals with systemic allergic reactions compared with the control subjects ( P.061 for Aedes vexans and P.008 for the remaining 4 species). By using the mean of the control subjects plus 1 SD as a cut-off level, 11 individuals had positive results to Aedes albopictus and up to 4 additional species; 3 individuals had positive results to only one species. Saliva-specific IgG levels were not significantly increased in the individuals with systemic allergic reactions compared with levels seen in the control subjects ( P.05).Acute systemic allergic reactions to mosquito bites involve mosquito saliva-specific IgE and can be characterized immunologically. Aedes albopictus is the most common species associated with systemic allergic reactions to mosquito bites.

Reactions to mosquito bites are immunological in nature, with the involvement of IgE-, IgG- and T lymphocyte-mediated hypersensitivities. Acquired desensitization to mosquito saliva may occur during childhood and adolescence or during long-term exposure to mosquito bites. Due to the lack of salivary preparations, allergic reactions to mosquito bites are underdiagnosed and undertreated. Recombinant mosquito saliva allergens with biological activity are being developed. These recombinant allergens will significantly improve diagnosis of mosquito allergy and eventually will also improve specific immunotherapy for patients with systemic reactions to mosquito bites.

Summary Background Topical bovine thrombin (TBT) is used as a topical haemostatic agent in haemodialysis (HD) units and for surgical procedures. It has caused allergic reactions during surgical procedures, but no previous cases have been reported with dialysis usage. Objective A case is reported of anaphylaxis due to TBT in an HD patient. This led to a determination of whether HD patients exposed to TBT develop specific IgE and IgG antibodies to TBT and an assessment of the frequency of allergic symptoms in HD patients as compared with peritoneal dialysis (PD) patients. Methods A cross-sectional study of 100 HD and 52 PD patients was performed using a questionnaire to determine allergic symptoms. An ELISA assay was developed to determine TBT-specific IgE and IgG serum antibodies among HD patients. HD patients without TBT exposure and age- and gender-matched blood donors served as controls. Results Allergic symptoms temporally related to dialysis reported by a questionnaire were significantly more frequent for HD patients than for PD patients: rhinoconjuctivitis 11% vs. 2%, asthma symptoms 20% vs. 6%, urticaria 23% vs. 8% (P values

Accurate diagnosis of mosquito allergy has been precluded by the difficulty of obtaining salivary allergens. In this study, we expressed, purified, characterized and investigated the clinical relevance of a recombinant Aedes aegypti salivary allergen, rAed a 1. Two cDNA segments were ligated together to form the full-length Aed a 1 gene. rAed a 1 was expressed using a baculovirus/insect cell system, and purified using a combination of anion-exchange and gel-filtration chromatography. The purified rAed a 1 bound to human IgE, as detected by ELISA, ELISA inhibition tests and immunoblot analyses. Epicutaneous tests with rAed a 1 and a commercial whole-body AE: aegypti extract, and AE: aegypti bite tests were performed in 48 subjects. Nine of 31 (29%) of the subjects with positive immediate bite tests also had a positive rAed a 1 immediate skin reaction and 32% had an positive immediate test to the commercial extract. Six of 33 (18%) of the subjects with positive delayed bite tests also had a positive rAed a 1 delayed skin reaction and 6% had a positive delayed test to the commercial extract. Furthermore, rAed a 1-induced flare sizes significantly correlated with mosquito bite-induced flare sizes. None of the subjects with negative bite tests had a positive skin test to rAed a 1 or to commercial extract. We conclude that the rAed a 1 has identical antigenicity and biological activity to native Aed a 1, can be used in the in vitro and in vivo diagnosis of mosquito allergy, and is more sensitive than mosquito whole-body extract for detecting delayed skin reactions.

Background: Most people develop skin reactions to mosquito bites, however, little is known about mosquito salivary allergens and the IgE responses to them. Objectives: We sought to identify these allergens and the specific IgE responses they elicit. Methods: Saliva or salivary gland extracts were prepared from 10 mosquito species, including seven species with worldwide distribution: Aedes (Ae.) aegypti, Ae. vexans, Ae. albopictus, Ae. togoi, Ae. triseriatus, Culex (Cx.) quinquefasciatus, Cx. pipiens, Cx. tarsalis, Anopheles (An.) sinensis , and Culiseta (Cs.) inornata . Proteins from these preparations were separated by sodium dodecylsulfate–polyacrylamide gel electrophoresis and transferred to nitrocellulose membranes, which were immunoblotted by sequential incubations with human serum, monoclonal anti-human IgE, and enzyme-conjugated goat anti-mouse IgG. Salivary allergens were analyzed by using a pooled serum from subjects allergic to mosquitos. Individual IgE responses to each allergen were evaluated in 12 subjects allergic to mosquitos living in Canada, the United States, and China, as well as in five subjects not allergic to mosquito bites. To study species-shared allergens, the membranes were immunoblotted with two rabbit antibodies specific to recombinant mosquito salivary proteins. Results: Three to sixteen salivary allergens with molecular masses ranging from 16 to 95 kd were found in each species. Both species-shared and species-specific allergens were identified by molecular masses, binding to the two rabbit antibodies, and individual IgE responses to species indigenous to and absent from the regions where the subjects lived. Salivary allergens, especially from Ae. aegypti, Ae. vexans, and Ae. albopictus , elicited higher IgE responses in subjects allergic to mosquitos than in nonallergic subjects. Conclusions: Species-shared and species-specific allergens that cause IgE responses in subjects allergic to mosquitos are immunologically identified. Species-shared allergens are the most important for potential use in diagnosis and immunotherapy. (J Allergy Clin Immunol 1998;101:498-505.)

To date, there are no ideal animal models for study of natural sensitization leading to IgE– and lymphocyte–mediated hypersensitivities. We established such a model in which four BALB/c mice were each sensitized by exposure to at least 16 mosquito Aedes aegypti bites, twice a week for 4 weeks. Four non–exposed control mice were also studied. Mosquito A. aegypti head and thorax extract, saliva, and two recombinant salivary allergens (rAed a 1 and rAed a 2) were used in vitro as antigens. Intradermal tests were performed. Serum mosquito antigen–specific IgG, IgG1, IgG2a, and total IgE were measured by ELISA; specific IgE was measured by passive cutaneous anaphylaxis (PCA). IgE responses to each antigen in the saliva were analyzed using Western blotting. Spleen lymphocyte proliferation assays were performed to determine the cell–mediated hypersensitivity response. Antigen–induced IL–4 and IFN–γ production in the spleen lymphocytes were evaluated using ELISA. After 4 weeks, all 4 mosquito–sensitized mice developed a positive immediate wheal 20 min after skin tests with mosquito antigens, and a positive delayed papule 24 h later, while control mice did not. Also, the sensitized mice had a positive PCA response, which correlated significantly with total IgE levels (r = 0.84, p

MDSCs, a heterogeneous population of cells that expand during many pathogenic conditions, have remarkable abilities to suppress T cell responses. Their role in murine colitis, induced by TNBS and therapeutic application, remains unclear. Murine colitis was induced through intrarectally administrating TNBS, twice. MDSCs in spleen and colonic LPMCs were identified using flow cytometric analysis. In adoptive transfer, MDSCs were isolated from spleen after TNBS challenges by using microbeads or generated in vitro by coculturing bone marrow cells with HSCs and then transferred into naïve mice. Two hours later, mice were then challenged with TNBS, once/week for 2 weeks. The mice were killed four days after the second TNBS delivery, and intestinal inflammation and cytokine levels and MDSC percentages were evaluated. The percentages of CD11b+Gr-1+MDSCs and subsets (CD11b+Ly6C+ and CD11b+Ly6G+MDSCs) were increased in spleen and/or colonic LPMCs in colitis mice and also correlated with the severity of intestinal inflammation. MDSCs isolated from colitis mice suppressed the proliferation of splenocytes in vitro. Adoptive transfer of MDSCs, isolated from colitis mice or generated in vitro, decreased intestinal inflammation, levels of IFN-γ, IL-17, and TNF, and percentages of spleen MDSCs when compared with controls. MDSCs that have inhibitory function in vitro and in vivo are increased and correlated with intestinal inflammation, suggesting that they may be used as a biomarker of disease activity and a cell-based biotherapy in IBD.

We report the cross-reactivities and comparative molecular masses of the IgE epsilon chains in humans, rats, mice, and dogs. Monoclonal human, rat, and mouse IgE, and our purified polyclonal dog IgE were used in the study. IgE of the 4 species, separated by SDS-PAGE, were analyzed by immunoblotting with polyclonal and monoclonal antihuman IgE, polyclonal and monoclonal antimouse IgE, monoclonal antirat IgE, and polyclonal antidog IgE antibodies. The polyclonal antihuman and polyclonal antimouse IgE cross-reacted with the IgE of the other 3 species, while their monoclonal forms cross-reacted with dog IgE only. Polyclonal antidog IgE cross-reacted with human and mouse IgE, while the monoclonal antirat IgE did not cross-react with any other species. "Reverse' passive cutaneous anaphylaxis in ragweed-sensitized dogs revealed that polyclonal antihuman and polyclonal antimouse IgE were able to elicit positive skin responses, and monoclonal antihuman, antirat, and antimouse IgE antibodies were not. The molecular masses of the epsilon chains were 77 kDa for mice, 75 kDa for rats and dogs, and 70 kDa for humans.

Extracts of one globally distributed mosquito species (Aedes vexans) and two North American species (Culex tarsalis and Culiseta inornata) were prepared from heads and thoraxes. Proteins of the three extracts were separated by 12% SDS-PAGE and transferred to nitrocellulose membranes for immunoblotting. Immunoblotting was completed by sequential incubations of the membranes with a pooled human serum from subjects allergic to mosquito bites, monoclonal antibodies to human IgE or IgG, and goat antimouse IgG conjugate. Twelve to sixteen antigens with molecular masses ranging from 18.5 to 160 kDa were found in each extract. Nine antigens were shared by 3 species and 6 were shared by 2 species. Only 3 were species-unique. Most antigens bound to both IgE and IgG antibodies. IgE and IgG antibodies against A. vexans were studied by immunoblotting using individual serum from subjects with or without skin reactions to A. vexans bites. All 3 subjects with severe skin reactions had strong IgE and IgG antibodies to 32.5, 40, and 50 kDa proteins. The patterns and magnitudes of IgE and IgG antibodies to the antigens varied among individuals. Very faint IgE antibodies to these antigens were found in the 2 subjects with no skin reactions, suggesting that IgE plays a role in the development of mosquito allergy.

Aims: To develop an IL-17 peptide-based virus-like particle vaccine that elicits autoantibodies to IL-17 and to evaluate the effects of the vaccine in mice with experimental colitis. Materials & methods: Recombinant IL-17 vaccines were constructed by inserting selected peptides derived from mouse IL-17 into the carrier protein, hepatitis B core antigen, using molecular engineering methods. To evaluate the in vivo effects of the vaccine, mice with 2,4,6-trinitrobenzene sulfonic acid-induced chronic colitis were injected three times with the vaccine, carrier or saline after the second delivery of 2,4,6-trinitrobenzene sulfonic acid. Colon inflammation and fibrosis were evaluated by histological examination. Serum IL-17-specific IgG and colon-tissue cytokine levels were measured by ELISA. In vitro inhibition tests of sera from vaccine-immunized mice were performed using IL-17-induced IL-6 production by NIH 3T3 cells and IL-17-induced TNF production by macrophages. Results: Immunization with the vaccine without the use of adjuvants induced high-titered and long-lasting antibodies to IL-17. Unexpectedly, vaccinated mice exhibited increases in colon inflammation, collagen deposition, levels of TNF and IL-17 cytokines compared with carrier and saline groups. Furthermore, in vitro study revealed that serum IL-17-specific IgG from vaccine-immunized mice significantly enhanced IL-17-induced IL-6 production and IL-17-induced TNF production dose-dependently. Conclusion: The IL-17 peptide-based vaccine enhances the bioactivity of IL-17 in vitro and in vivo, providing a potential immunotherapy for treatment of diseases associated with insufficient IL-17 production, such as hyper-IgE syndrome.

The binding of dog immunoglobulins G, A, M and E to concanavalin A (Con A) has been investigated. A passive cutaneous anaphylaxis test was used for measurement of dog IgE, and enzyme-linked immunosorbent assay was used for measurement of dog IgG, IgA and IgM. After the dog serum fraction was applied to a Con A-Sepharose column, sequential elution with different buffers was performed; 100% of IgE and IgM, 60% of IgG and 58% of IgA bound to the Con A-Sepharose. IgE was eluted by mannose, methylglucose, and methylmannoside. IgG was eluted by glucose, mannose, methylglucose, and methylmannoside. IgA and IgM were eluted by methylmannoside only. This provides a useful technique in the purification of dog immunoglobulins, especially dog IgE.

Inflammatory bowel disease (IBD) is characterized by increasing morbidity and, if suboptimally treated, poor prognosis. Recent evidence strongly suggests that dysfunctional immune responses play an important role in the pathogenesis of IBD. Therefore, immunologically downregulating the overactivated innate and adaptive immune responses may be a better approach to treat IBD than currently used pharmaceutical therapies. In recent years, many new biological therapies have been developed. These therapies are shown to be effective for inducing remission, preventing complications, improving life quality of the patients, and reducing hospitalization and surgical rates. This article introduces and discusses these new biological agents that have been used effectively in clinic for IBD patients.

AMP-activated protein kinase (AMPK), a cellular energy sensor, has been reported to participate in modulating inflammatory responses, but its role in intestinal inflammation remains unclear. IBD has been characterized by excessive innate and adaptive immune responses. Here, the roles of 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), an agonist of AMPK, in regulating immune responses of experimental colitis were investigated. In vitro effects of AICAR on LPS-induced macrophage activation and Th1 and Th17 differentiation, as well as in vivo effects of AICAR in mice with 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis, were explored. In acute colitis, daily AICAR treatment commenced 2 days after TNBS delivery (day 1), while in relapsing colitis, AICAR treatment commenced after three weekly TNBS administrations. Colon inflammation, production of proinflammatory cytokines and NF-κB activation in colon tissues, and Th1 and Th17 cell populations in lamina propria mononuclear cells (LPMCs) and mesenteric lymph node cells (MLNs) were assayed. Results show that AICAR significantly inhibited in vitro LPS-induced macrophage activation and Th1 and Th17 cell differentiation. Administration of AICAR was therapeutically effective in ameliorating acute and relapsing experimental colitis, as shown by reduced body weight loss and significant attenuation in colon histological inflammation. Moreover, this treatment inhibited NF-κB activation in macrophages, and reduced levels of TNF, Th1- and Th17-type cytokines, and Th1 and Th17 cell populations in LPMCs and MLNs. AICAR-initiated AMPK activation may act as a central downregulator in ongoing innate and adaptive immune responses of murine colitis, providing a novel therapeutic approach in the treatment of IBD.

We studied the quantitative responses of short ragweed (RW)-pollen-specific serum antibodies in 22 patients with RW immunotherapy (IT) and in a different set of 31 patients, 16 of whom stopped RW IT after more than 5 years of treatment. Serum was assayed before and after season, 1 year before and 1 and 2 years after starting IT, and 1 year and 2 years after stopping IT. RW pan-IgG, RW IgG1, and RW IgG4 were measured by ELISA, and RW IgE by RAST. Absolute quantities of RW IgG1 and RW IgG4 in reference sera were estimated by least-squares multiple regression analysis of 223 sera with the equation RW pan-IgG = RW IgG1 + RW IgG4. IgG1 is dominant in the early immune response of IT and disappears relatively slowly when IT is stopped. In contrast, IgG4 appears in significant quantities only after prolonged IT and disappears rapidly when IT is stopped. The apparent average half-life of RW IgG4 (9 months) was significantly shorter than that of RW IgG1 (29 months) (p less than 0.001). Before IT, mean RW IgE rose 180% (p less than 0.01) during the RW pollination season (August to November). This seasonal rise in RW IgE was ablated after IT from 1 year up to 8 years, but returned the year after IT was stopped. After 2 years of IT, the RW IgG1 and IgG4 levels were significantly correlated with RW IgE (r = 0.94 and 0.81; p = 0.0001 and 0.005).

We studied the binding of dog immunoglobulins G, A, M and E to protein A and protein G. Passive cutaneous anaphylaxis (PCA) testing was used for the measurement of dog IgE and enzyme-linked immunosorbent assays (ELISA) were used for the measurements of dog IgG, IgA and IgM. Protein A from lyophilized cells of Staphylococcus aureus bound 97% of IgE, 98% of IgG, 81% of IgA, and 97% of IgM. Protein A-Sepharose CL-4B bound 87% of IgE, 100% of IgG and IgA, and 98% of IgM. In a stepwise elution with varying pH, a small amount of IgE was eluted at pH 5 and pH 6 and all the remaining Igs were eluted at pH 3 from the protein A column. In contrast to protein A, dog IgE was not bound to Protein G-Sepharose, while 100% of IgG, 95% of IgA, and 44% of IgM were bound to Protein G-Sepharose.

We have developed sensitive amplified immunoassays for measurement of IgE and IgG4 ragweed (RW) antibodies in unconcentrated nasal washes. IgE to Amb a I (formerly antigen E) can be assayed to less than 0.1 ng/ml using IgE capture by anti-IgE on microtitre plates and an alkaline phosphatase-conjugated Amb a I with an amplification substrate technique. IgG4 to whole RW extract was assayed to less than 0.01 ng/ml by amplification ELISA using monoclonal anti-IgG4. Nasal washes (NW) (10 ml) and serum were obtained in December from 22 RW-sensitive patients before and after 1 and 2 yr of RW immunotherapy (IT), and assayed for Amb a I IgE or RW IgE and RW IgG4 antibodies Amb a I IgE could be measured in the NW of 15/22 pre IT, 19/22 at 1 yr IT, but only 3/10 at 2 yr IT (compared with pre-IT, P less than 0.05). Mean Amb a I IgE in NW was 0.66, 0.36 and 0.21 ng/ml at pre, 1 yr and 2 yr IT (P-values greater than 0.05). Mean serum RW IgE, was 76, 55 and 27 ng/ml at pre, 1 yr and 2 yr IT (P-values greater than 0.05). Amb a I IgE in nasal washes was correlated with RW IgE in serum (r = 0.56, P less than 0.001, n = 44). RW IgG4 was detectable in NW of 15/22 pre-IT, 18/22 at 1 yr IT and 9/10 at 2 yr IT (P-values greater than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Interleukin (IL)-13 is critical in asthma pathogenesis. Previously, we have developed an IL-13 peptide-based vaccine and confirmed that subcutaneous immunization with the vaccine suppressed airway allergic inflammatory responses in a mouse asthma model. In the present study, we sought to test if mucosal immunization with the vaccine could be a potential approach, by inducing specific autoantibodies of both local IgA in the airway and systemic IgG in serum, to provide an overall suppression of redundant IL-13 effects. The results show that intranasal vaccination induces IL-13-specific IgA responses in multiple mucosal tissues and higher titers of IgG in serum than subcutaneous vaccination. This approach leads to a more effective suppression of ovalbumin-driven Th2 patterns of antibody responses and airway IL-13 and eosinophil accumulation than subcutaneous immunization, even when the induced IL-13 IgG responses were at a similar level. In conclusion, mucosal vaccination may be an innovative potential approach in the treatment of asthma.

Purpose of review Allergic reactions, including severe local and systemic reactions to mosquito bites, are immunological in nature, and involve immunoglobulin E, immunoglobulin G, and T-lymphocyte-mediated hypersensitivities in response to allergens in mosquito saliva. Naturally acquired desensitization to mosquito saliva may occur during childhood or during long-term exposure to mosquitoes. Due to the lack of availability of mosquito salivary preparations for use in skin tests and in-vitro tests, allergic reactions to mosquito bites are under diagnosed and under treated. Recent findings Recombinant saliva allergens with biological activity are being developed. Recombinant Aedes aegypti salivary allergen rAed a 2 has been expressed, purified, characterized and used in in-vitro diagnosis of mosquito allergy. Mosquito saliva-induced non-immunoglobulin E-mediated skin mast cell degranulation was found to induce macrophage-inflammatory protein 2 in the skin and interleukin-10 in draining lymph nodes. Summary In this review, we discuss the allergic reactions to mosquito salivary allergens, the immune mechanisms involved, natural desensitization and immunotherapy with mosquito extracts, characteristics of salivary allergens and their recombinant forms, and prevention and treatment of allergic reactions to mosquito bites. Eventually, recombinant salivary allergens will significantly improve the diagnosis of mosquito allergy, and will also improve specific immunotherapy for patients with systemic reactions to mosquito bites.

1. Dabrowski AJ, Van Der Brempt X, Soler M, Seguret N, Luciani P, Charpin D, et al. Cat skin as an important source of Fel d 1 allergen. J Allergy Clin Immunol 1990;36:462-5. 2. Brown PR, Leitermann K, Ohman JL Jr. Distribution of cat allergen 1 in cat tissues and fluids. Int Arch Allergy Appl Immunol 1984;74: 67-70. 3. Wentz PE, Swanson MC, Reed CE. Variability of cat-allergen shedding. J Allergy Clin Immunol 1990;85:94-8. 4. Jalil-Colome J, Dornelas de Andrade A, Birnbaum J, Casanova D, Mege JL, Lanteaume A, et al. Sex difference in Fel d 1 allergen production. J Allergy Clin Immunol 1996;98:165-8. 5. Miller JD, Bell JB, Vandernoot D. Castration of pets does not prevent allergy to pets. Clin Exp Allergy 1995;25:574-5.