猫主要过敏原Fel d 1的二级结构及B细胞抗原表位预测

目的预测猫主要过敏原Fel d 1的二级结构和B细胞抗原表位。方法以Fel d 1肽链Ⅰ和肽链Ⅱ的氨基酸序列为基础,通过DNASIar Protean软件,采用Chou—Fasman方法预测蛋白质的二级结构;用Kyte—Doolillle法预测亲水性;用Karplus—Sehulz方法预测柔韧性;用Emini方法预测表面可及性;用Jaineson—Wolf方法预测抗原性指数。结果对Fel d 1的二级结构和B细胞抗原表位预测的结果表明,第15～21、48～57、103～111、138～143、151～161区段是潜在的B细胞抗原表位。结论本研究有助于确定Fel d 1的B细胞优势抗原表位,为Fel d 1的抗体设计以及抗原性改造提供理论依据     

Validity of the interview on pets kept at home for predicting the actual domestic expsoure to their specific allergens. Krakow inner city area study

The aim of the study was to describe the exposure to dog (Can f 1) and cat (Fel d 1) allergens within homes of very young children living with and without pets, and to assess the validity of the interview on pets for predicting the actual exposure to pet allergens in house dust. House dust samples were collected in 275 dwellings from the mattresses, children’s bedroom and kitchen floors. In the laboratory, dust samples were analyzed for Can f 1 and Fel d 1 using monoclonal antibody enzyme-linked immunosorbent assays (ELISA). The majority of households (79.3%) had neither a dog nor a cat living in the home over the past 6 months preceding the survey. Dog allergen above 2 μg/g dust were found in 22.5% of homes and 14.2% of homes contained dog allergen above 10 μg/g of house dust. In the total study sample, cat allergen above 1 μg/g of dust were found in 12.7% of homes, and 3.3% of homes contained Fel d 1 levels greater than 8 μg/g of dust. The majority of children (75.0%) with reported ownership of dogs were exposed to Can f 1 levels above 2 μg/g of house dust, and 73.1% of children with cats at home were exposed to Fel d 1 concentrations above 1 μg/g house dust. The results of the study showed that post-test probability of the true exposure to Can f 1 above 2 μg/g dust in houses with positive interview on indoor dogs was 75.0% (95%CI: 61.7–84.8%). On the other hand, the prediction of exposure estimated from the interview data on indoor dogs produced 12.6% of false negatives (95% CI: 9.9–15.8%). Similarly, the post-test probability of the true exposure to Fel d 1 above 1 μg/g dust in houses with positive interview on indoor cats was 73.1% (95%CI: 55.1–85.7%). On the other hand, the interview data produced 6.4% false negatives (95% CI: 4.6–9.0%). In conclusion, the study demonstrated that homes in Poland with pet ownership are important reservoir of Can f 1 and Fel d 1 allergens with levels that might induce allergic symptoms. Even in homes of children without a dog or cat indoors, there was a higher prevalence of pet allergens at the levels above allergic sensitisation thresholds. This may have an important implication for epidemiologic studies on pet related allergy and prevention practice.     

Divergent antibody isotype responses induced in mice by systemic exposure to proteins: a comparison of ovalbumin with bovine serum albumin.

Whereas many foreign proteins are immunogenic, only a proportion is associated commonly with allergy, having the potential to induce the quality of immune response necessary for IgE antibody production and the development of immediate type hypersensitivity reactions in the gastrointestinal and/or respiratory tracts. In the context of toxicological evaluations there is a need to identify those properties that confer on proteins the ability to provoke allergic reactions. The characteristics of antibody responses induced in BALB/c strain mice following administration of ovalbumin (OVA), a significant human allergen, have been compared with those provoked by bovine serum albumin (BSA), a protein considered to have more limited allergenic potential. Intranasal or intraperitoneal (ip) administration of BSA or OVA elicited vigorous IgG and IgG1 antibody responses. Differential IgE antibody production was observed, however, with OVA stimulating relatively high IgE antibody titres at all doses tested whereas no or low titre IgE antibody was detected following exposure to BSA. Furthermore, a differential capacity for IgG2a antibody responses was observed, with only BSA provoking high titres of this IgG subclass. The relative quality of induced responses was equivalent following administration of these proteins via mucosal (in) tissue or via a non-mucosal (ip) route of exposure. IgG2a antibody production is promoted by the type 1 cytokine interferon gamma (IFN-gamma), whereas IFN-gamma and the type 2 cell product interleukin 4 exert reciprocal antagonistic effects on IgE antibody responses. Although cytokine expression patterns were not analysed in this series of experiments, the differential IgE and IgG subclass antibody responses induced by BSA and OVA are consistent with the preferential activation of T helper (Th) 1- and Th2-type cells, respectively. These data indicate that proteins can provoke in mice characteristic antibody (IgE and IgG) isotype profiles suggestive of discrete T lymphocyte responses and that such differences may be associated with variable allergenic activity.     

Dietary perilla oil lowers serum lipids and ovalbumin-specific IgG1, but increases total IgE levels in ovalbumin-challenged mice

Our previous studies indicated that α-linolenic acid (ALA)-rich perilla oil might alleviate bronchoalveolar inflammation. However, it failed to modulate the Th1/Th2 balance toward the Th1 pole during Th2-skewed allergic airway inflammation in mice. This study attempts to further investigate the effects of dietary perilla oil on serum lipids and immunoglobulin profiles using an ovalbumin (OVA)-challenged mouse model. The inbred female BALB/c mice were randomly divided into four groups and fed different AIN-76 feeds containing 5% corn oil (rich in linoleic acid, 18:2n-6 polyunsaturated fatty acids (PUFA), as a control diet), 5% perilla oil (rich in α-linolenic acid, 18:3n-3 PUFA) or 5% compound oil containing 50% corn oil and 50% perilla oil, respectively, for 35 consecutive days ad libitum. Experimental mice were sensitized by an intraperitoneal injection of alum-precipitated antigen containing ovalbumin on 7, 14 and 21 days after supply of the specified experimental diets. One week later, the mice were then challenged by aerosolized OVA. The results showed that dietary perilla oil administration significantly (P < 0.05) decreased the relative liver tissue weight (RTW) and serum lipid levels including triglycerides, total cholesterol, HDL- and LDL-cholesterol. However, the HDL/LDL ratio was also significantly lowered by dietary perilla oil. Dietary perilla oil markedly decreased serum OVA-specific IgG1 level and total IgA antibodies (Th2 antibodies). Unfortunately, it also increased non-specific serum IgE (Th2 antibody) levels. The results suggest that dietary perilla oil might have a moderately beneficial effect on asthmatic allergy via lowering serum lipids and OVA-specific IgG1, as well as total IgA levels. However, it failed to obviously modulate Th1/Th2 antibody levels via isotype switching of B cells from Th2 antibody to Th1 antibody.     

青霉素过敏病人血清特异性IgE和IgG抗体

采用放射过敏原吸附试验(RAST)和酶联免疫吸附试验(ELISA)测定52例青霉素过敏病人血清特异性IgE、IgG抗体，进一步探讨青霉素过敏反应机制。结果 52例过敏病人特异性IgE、IgG抗体的阳性率分别为50％和44．2％，若RAST与ELISA联合检测，IgE和IgG抗体总阳性率增至63.5％。荨麻疹组BPO—IgG水平高于过敏性休克组(P＜0.01)，过敏性休克病人BPA—IgG水平明显高于BPO—IgG(P＜0．01)，荨麻疹组内BPO—IgE水平与BPA—IgE无显著差异(P＞0．05)。但均比过敏性休克组高。研究结果提示，荨麻疹与BPO—IgE和BPA—IgE关系密切，过敏性休克与BPO—IgE和BPA—IgG关系密切；同时检测IgE和IgG抗体，可提高诊断阳性率。     

The ex vivo study of synergistic effects of polycyclic aromatic hydrocarbon, benzo(a)pyrene with ovalbumin on systemic immune responses by oral route

The present study was undertaken in order to examine whether oral administration of soluble antigen together with one of polycyclic aromatic hydrocarbons (PAHs) which is present in diesel exhaust particles (DEPs) called benzo(a)pyrene (BP), induced the systemic immune response in mice or not. Mice were orally given 1mg of ovalbumin (OA), a common food allergen, every 3 days over a period of 15 days. The results showed that oral administration of OA plus BP produced anti-OA IgE antibodies in serum, whereas either OA or BP alone failed to show the antigen-specific IgE antibody production. Production of anti-OA IgE antibody, which is dependent on Th2 CD4(+) T cells, was seen in mice fed with combined OA and BP was significantly higher than that of other groups. The anti-OA antibody production was associated with marked secretion of the Th1 cytokines, IFN-gamma and IL-12p70 as well as the Th2 cytokines IL-4, and IL-10. These results suggest that BP may act as a mucosal adjuvant in the gut enhancing systemic Th1 and Th2 immune responses and might play a role in oral immunization and food allergy.     

Ambient air particles from four European cities increase the primary cellular response to allergen in the draining lymph node

In the RAIAP (respiratory allergy and inflammation due to ambient particles) project, qualitative properties of ambient air particles from Amsterdam, Oslo, Lodz and Rome were investigated in relation to inflammation and allergy. Most collected particle fractions were found to increase the allergen-specific IgE and IgG2a responses after subcutaneous injection of particles with allergen in mice. However, some fractions appeared to skew the antibody response towards more Th1- or Th2-associated antibody isotypes, and the fine fractions were found to be more potent than the coarse fractions with regard to IgE adjuvant activity. In the present study we investigated the cellular response in the draining lymph node 5 days after a subcutaneous injection of selected RAIAP particle fractions. The particles (100 microg) were injected into both hind footpads of BALB/cA mice, in the presence or absence of the allergen ovalbumin (OVA, 50 microg). We also studied if the coarse and fine RAIAP particle fractions affected the cellular responses to OVA differently. The number of lymph node cells, as well as the relative number of B and T lymphocytes and T helper cells were determined. Expression of cell surface molecules (MHC class II, CD86 and CD23) and ex vivo cytokine production (IL-4, IL-10 and IFN-gamma) by the lymph node cells were measured. Overall, particles in the presence of allergen enhanced the levels of the various cellular parameters compared to allergen alone or particles alone. In the absence of allergen, ambient air particles, in contrast to diesel exhaust particles, marginally affected some cellular parameters. By histological examination of the lymph node, the particles appeared to be scattered between the lymphocytes, often localised within macrophage-like (acid phosphatase positive) cells. The cell parameters measured could, for the individual sample, neither predict the degree of a Th2- or Th1-skewed antibody response, nor the stronger antibody adjuvant capacity of the fine than the coarse particle fractions. In conclusion, we have shown that coarse and fine ambient air particles from different European cities enhance the cellular response in the draining lymph node after injection with an allergen. In the absence of allergen, ambient particles only marginally affected the cellular parameters.     

The fungal cell wall component beta-1,3-glucan has an adjuvant effect on the allergic response to ovalbumin in mice

The polyglucose beta-1,3-D-glucan is a major structural component of the cell wall of yeasts and fungi. In the present study, the adjuvant activity of beta-1,3-glucan from the fungus Sclerotinia sclerotiorum (SSG) on the response to the model allergen ovalbumin (OA) was studied, using the popliteal lymph node assay (PLNA) in BALB/c mice. The adjuvant activity on the local cellular response was determined by measuring the weight, cell number, and proliferation of the extracted PLNs. The levels of OA-specific immunoglobulin (Ig)E, IgG1, and IgG2a in serum were measured by enzyme-linked immunosorbent assay (ELISA). Groups of 8 mice were given either SSG + OA, SSG alone, or OA alone on d 0. Thereafter they were exsanguinated on d 20, or reinjected with OA on d 21, before exsanguination on d 26 or 33. Only on d 26 was SSG + OA found to significantly increase the PLN weight and cell numbers, but not cell proliferation (thymidine incorporation), compared with OA or SSG alone. SSG + OA was also found to significantly increase both the anti-OA IgE and IgG1 levels on d 20, 26, and 33 compared to OA alone. Compared to SSG alone, SSG + OA increased the OA-specific IgE and IgG 1 levels significantly on d 26 and 33, but not on d 20. A similar increase was not found for IgG2a. Our results show that beta-1,3-D-glucan provides a clear Th2-dependent (allergic) immune response to OA, indicated by elevated levels of IgE and IgG1 and not IgG2a, in the mouse model used.     

Aqueous extract of peanut skin and its main constituent procyanidin A1 suppress serum IgE and IgG1 levels in mice-immunized with ovalbumin

In this study, we investigated the effects of an aqueous extract of peanut (Arachis hypogaea L.) seed skin (PSE) and its main constituent procyanidin A1 (PA) on the allergic response to allergen ovalbumin (OVA) in a mouse model. Mice immunized interaperitoneally with OVA dramatically increased anti-OVA IgE and total IgG1 levels in serum compared with non-treated control mice. Oral injection of PSE at doses ranging from 10 to 100 mg/kg/d (for 21 consecutive days) decreased anti-OVA IgE and IgG1 levels 21 d after OVA-immunization. OVA-induced increments in spleen weight and peripheral white blood cell count were also suppressed by this PSE administration. Polyphenol-enriched fractions from apple (30 mg/kg) and grape seed (30 mg/kg) also decreased anti-OVA IgE level but did not affect total IgG1 levels. Oral injection of PA (1 to 10 mg/kg/d) purified from PSE resulted in a suppression of IgE and total IgG1 levels in serum. An increment of serum interleukin-4 level in mice that were immunized with OVA was reduced by all tested samples, whereas PSE and PA were the only compounds that could reverse the reduced interferon-gamma level by OVA. These findings suggest that intake of PSE or its main active constituent PA may prevent an allergic reaction by inhibiting immunoglobulin synthesis, and the mechanism of this action of PSE and PA is in part due to their regulation of T helper cytokine production.     

Cry-consensus peptide, a novel peptide for immunotherapy of Japanese cedar pollinosis, induces Th1-predominant response in Cry j 1-sensitized B10.S mice

Cry-consensus peptide (CCP) is a newly designed peptide for peptide-based immunotherapy of Japanese cedar pollinosis but its mechanism of efficacy is unknown. We investigated the effect of CCP on Cry j 1-specific Th1/Th2 response in a mice model. Subcutaneous injection of CCP decreased Cry j 1-specific IgE and IgG1 in blood slightly, but the IgG2a level was increased significantly in a dose dependent manner. Splenocytes from these mice were stimulated with Cry j 1 in vitro. This inhibited IL-4, IL-5 and IL-10 secretion significantly, but IFN-gamma secretion was increased. In vitro CCP stimulation of splenocytes from Cry j 1-sensitized mice induced more marked Th1-predominancy of cytokine production than native allergen stimulation. Taken together, these data suggest that one of the mechanisms of CCP is dependent on the modulation of the antigen-specific Th1/Th2 response.     

Specific IgE and IgG1 Responses to Subtilisin Carlsberg (Alcalase) in Mice: Development of an Intratracheal Exposure Model

The primary objective of this study was to examine the feasibilityof using a mouse model to evaluate the immunogenicity of proteinsas a potential method to determine occupational exposure guidelines.Mice were intratracheally administered a benchmark protein allergen,subtilisin Carlsberg (Alcalase) in detergent matrix once a weekfor 4 to 6 weeks and specific IgE and IgG1 levels were determined.In all experiments, specific IgE levels were determined by usinga rat basophilic leukemia cell (RBL) release assay, while specificIgG1 was measured by an ELISA. A good correlation was observedbetween IgE titers determined by the RBL assay and rat passivecutaneous anaphylaxis assay. Intratracheal administration ofprotease with detergent matrix was found to result in significantIgE and IgG1 responses that were dose related. Detergent matrixwas found to enhance the Alcalase-specific IgE and IgG1 responsewhen administered by the intratracheal route. The IgG1 responsewas much more robust, easier to measure, and found to followthe IgE response. These results suggest that a mouse intratrachealmodel is a feasible approach to examining the immunogenic potencyof enzymes using specific IgE or IgG1 as the end points. Additionaldevelopment and validation of the mouse model with other typesof proteins will be pursued.     

THE FUNGAL CELL WALL COMPONENT b-1,3-GLUCAN HAS AN ADJUVANT EFFECT ON THE ALLERGIC RESPONSE TO OVALBUMIN IN MICE

The polyglucose b -1,3-D-glucan is a major structural component of the cell wall of yeasts and fungi. In the present study, the adjuvant activity of b-1,3-glucan from the fungus Sclerotinia sclerotiorum (SSG) on the response to the model allergen ovalbumin (OA) was studied, using the popliteal lymph node assay (PLNA) in BALB/c mice. The adjuvant activity on the local cellular response was determined by measuring the weight, cell number, and proliferation of the extracted PLNs. The levels of OA-specific immunoglobulin (Ig)E, IgG1, and IgG2a in serum were measured by enzyme-linked immunosorbent assay (ELISA). Groups of 8 mice were given either SSG + OA, SSG alone, or OA alone on d 0. Thereafter they were exsanguinated on d 20, or reinjected with OA on d 21, before exsanguination on d 26 or 33. Only on d 26 was SSG + OA found to significantly increase the PLN weight and cell numbers, but not cell proliferation (thymidine incorporation), compared with OA or SSG alone. SSG + OA was also found to significantly increase both the anti-OA IgE and IgG1 levels on d 20, 26, and 33 compared to OA alone. Compared to SSG alone, SSG + OA increased the OA-specific IgE and IgG1 levels significantly on d 26 and 33, but not on d 20. A similar increase was not found for IgG2a. Our results show that b -1,3-D-glucan provides a clear Th2-dependent (allergic) immune response to OA, indicated by elevated levels of IgE and IgG1 and not IgG2a, in the mouse model used.     

Adjuvant effect of quaternary ammonium compounds in a murine model

It has been suggested that occupational exposure to quaternary ammonium compounds (QACs) may promote the development of allergic airway diseases. In this study, hazard identifications of the adjuvant effect of cetylpyridinium chloride (CPC), dimethyldioctadecylammonium bromide (DDA), hexadecyltrimethylammonium bromide (HTA), and tetraethylammonium chloride (TEA) were performed in a screening bioassay. Female BALB/c mice were injected subcutaneously with the model allergen ovalbumin (OVA) alone or together with different quantities of one of the QAC test compounds. After one or two boosters, levels of OVA-specific IgE, IgG1 and IgG2a antibodies were measured in sera. CPC and DDA increased IgE and IgG1 antibody production, respectively, compared to the OVA control group, whereas HTA and TEA showed no adjuvant effect. Nevertheless, when TEA was given in combination with DDA, the adjuvant effect was up to six-fold higher than the adjuvant effect of DDA alone. Only DDA had a statistically significant adjuvant effect on IgG2a antibody levels.     

Effect of diesel exhaust particles and their components on the allergen-specific IgE and IgG1 response in mice

Increased antigen-specific IgE expression is a hallmark of the allergic response in mice. IgG1 may also be involved. Co-injection of mice with diesel exhaust particles (DEP) and ovalbumin three times over a 2 week period lead to a rapid and marked elevation of ovalbumin-specific IgE, IgG1 and also IgG2a, compared with ovalbumin alone. When DEP were injected 1 day before or after ovalbumin on each occasion, their adjuvant effect was considerably muted, suggesting that the adjuvant effect of DEP is short-lived, or that a physical interaction between ovalbumin and DEP is required. DEP were extracted with methylene chloride. Both the resulting core carbon particles and the organic extract enhanced ovalbumin specific IgE and IgG1 levels. Thus the adjuvant effect of DEP in this model is due both to the physical and the chemical attributes of the particles. The tricyclic hydrocarbons phenanthene (the most prevalent polycyclic aromatic hydrocarbon in DEP) and anthracene were both capable of enhancing antigen-specific IgE and IgG1 production. The phenolic antioxidant, butylated hydroxyanisole, which can affect gene expression via the antioxidant responsive element (ARE), had a lesser effect. Two agonists for the aryl hydrocarbon receptor, 3-methychloranthrene and 2,3,7,8-tetrachlorodibenzo-p-dioxin, either were without effect or suppressed the response, suggesting that DEP adjuvancy may not be mediated by this receptor.     

IgG and IgE antibody responses following exposure of Brown Norway rats to trimellitic anhydride: comparison of inhalation and topical exposure

A variety of chemicals can cause sensitisation of the respiratory tract and occupational asthma, including certain acid anhydrides, diisocyanates and reactive dyes. As yet, no well-validated methods are available for the toxicological evaluation of the respiratory sensitising potential of chemicals. One approach which has been explored recently is the evaluation of induced IgE responses or cytokine expression patterns in rats or mice following topical exposure to chemical. Thus, it has been demonstrated that topical exposure of rodents to respiratory sensitising chemicals, but not to contact allergens, causes a dose-dependent and time-related increase in the concentration of total IgE. Using the reference respiratory allergen trimellitic anhydride (TMA), we have considered here the influence of route of exposure on the nature of induced immune responses. Specific IgG and IgE antibody responses and changes in total serum concentration of IgE have been measured following exposure of Brown Norway (BN) starin rats to TMA by topical administration or by inhalation. Exposure to TMA by both routes resulted in the stimulation of specific IgG and IgE antibody, although responses were considerably more vigorous after dermal exposure. Topical treatment also provoked marked and sustained increases in total serum IgE levels, whereas exposure via the respiratory tract stimulated a more transient elevation of this immunoglobulin in a minority of animals which reached statistical significance only at the highest dose group. The lesser vigour of the immune response following inhalation exposure is likely to be related to the considerably lower total antigenic dose which is delivered by this route. Nevertheless, these results show that the nature of immune response with respect to antibody isotype profile provoked by topical administration of TMA is qualitatively comparable with that stimulated by inhalation exposure to the same chemical. For the purposes of hazard assessment and identification of potential chemical respiratory allergens as a function of induced changes in serum IgE concentration, however, the evidence is that topical administration of test material is the preferred route of exposure.     

8-Mercaptoguanosine-mediated enhancement of in vivo IgG1, IgG2 and IgG3 antibody responses to polysaccharide antigens in normal and xid mice

We have examined the effects of the immune adjuvant 8-mercaptoguanosine (8sGuo) on the in vivo antibody response to the T-cell-independent type 2 antigen, TNP-Ficoll. While 8sGuo enhanced the IgG1, IgG2 and IgG3 antibody responses, it was without effect on the IgM antibody responses. Increasing the dose of injected 8sGuo from 30 to 300 mg or the frequency or its injection led to greater enhancement in the antibody response, which varied from 20 to 100 times that of control responses. The effect of 8sGuo was relatively early acting in that it no longer enhanced anti-TNP antibody responses when given 3 days after antigen injection. Its ability to mediate an adjuvant effect on antibody responses was demonstrable even under conditions where the injected antigen by itself stimulated either no or low-level antibody responses. Thus, it enhanced the antibody response to the very weak antigen, pneumococcal polysaccharide, and restored the antibody response of nonresponder immune defective xid mice to TNP-Ficoll. These results extend the earlier observations of Goodman and coworkers by demonstrating that in vivo IgG response to type 2 polysaccharide antigens can be enhanced in normal mice and restored in xid immune-deficient mice.     

Immune Modulation by Adjuvants Combined with Diphtheria Toxoid Administered Topically in BALB/c Mice After Microneedle Array Pretreatment

Purpose  In this study, modulation of the immune response against diphtheria toxoid (DT) by various adjuvants in transcutaneous immunization (TCI) with microneedle array pretreatment was investigated. Methods  TCI was performed on BALB/c mice with or without microneedle array pretreatment using DT as a model antigen co-administrated with lipopolysaccharide (LPS), Quil A, CpG oligo deoxynucleotide (CpG) or cholera toxin (CT) as adjuvant. The immunogenicity was evaluated by measuring serum IgG subtype titers and neutralizing antibody titers. Results  TCI with microneedle array pretreatment resulted in a 1,000-fold increase of DT-specific serum IgG levels as compared to TCI. The immune response was further improved by co-administration of adjuvants, showing a progressive increase in serum IgG titers when adjuvanted with LPS, Quil A, CpG and CT. IgG titers of the CT-adjuvanted group reached levels comparable to those obtained after DT-alum subcutaneous injection. The IgG1/IgG2a ratio of DT-specific antibodies decreased in the following sequence: plain DT, Quil A, CT and CpG, suggesting that the immune response was skewed towards the Th1 direction. Conclusions  The potency and the quality of the immune response against DT administered by microneedle array mediated TCI can be modulated by co-administration of adjuvants.     

Intranasal IL-12 produces discreet pulmonary and systemic effects on allergic inflammation and airway reactivity

IL-12 modulates T cell responses between helper T cells Th2 and Th1; however, the therapeutic potential of IL-12 for allergic diseases either directly or as an adjuvant in allergen therapy has been controversial. The role of intranasal IL-12 as an adjuvant in modulating the grass pollen allergen (GAL) therapy-induced systemic immune response and lung-specific inflammation and airway reactivity was examined in this study using a mouse model of established allergic asthma. The effects of intranasal or nebulized IL-12 with or without intranasal anti-IFN-gamma antibody were examined in groups of control and allergen-sensitized or -challenged mice. T cell cytokine patterns, antibody response profiles, pulmonary inflammation and airway reactivity were examined. Intranasal IL-12 was found to be more effective in the Th2-Th1 shifting of immune response and anti-inflammatory activity in the lung compared to nebulized IL-12 at the given doses. Intranasal IL-12 significantly decreased production of IFN-gamma, eotaxin and LTC4/D4/E4 in the lung and decreased eosinophil infiltration, resulting in attenuated airway hyper-responsiveness in GAL-sensitized (GS) mice. In contrast, intranasal IL-12 significantly increased IFN-gamma production in the thoracic lymph node cultures and decreased the IL-5/IFN-gamma ratio, suggesting a Th2-Th1 shift. Also, intranasal IL-12 increased GAL-specific IgG2a antibody response, while the IgE response remained unaffected. The systemic effects of IL-12 were IFN-gamma dependent. IL-12 induces differential expression of its own receptor beta1 and beta2 subunits in the lung tissues to augment IL-12 responsiveness. Together, these results demonstrate that intranasal IL-12 is effective in shifting the systemic immune response in the direction of Th1 in IFN-gamma-dependent manner, while decreasing pulmonary inflammation and airway reactivity independent of IFN-gamma. Thus, intranasal delivery of IL-12 may provide an approach for the treatment of asthma and may be useful as an adjuvant in local nasal immunotherapy (IT) and in asthma.     

Surface-linked liposomal antigen induces IgE selective unresponsiveness in a T-cell independent fashion

We previously reported that surface-linked liposomal antigen induced IgE-selective unresponsiveness. The results were consistent even when different coupling procedures for antigen with liposomes, or for liposomes with different lipid components, were employed. During the course of an investigation intended to clarify the mechanism of IgE-selective unresponsiveness induced by surface-coupled liposomal antigens, we discovered an alternative approach to regulate the production of IgE, one that is independent of the activity of T-cells. Immunization of mice with OVA-liposome conjugates induced IgE- selective unresponsiveness without apparent Th1 polarization. Neither interleukin-12 (IL-12), IL-10, nor CD8(+) T-cells participated in the regulation. Further, CD4(+) T-cells of mice immunized with OVA-liposome were capable of inducing antigen-specific IgE synthesis in athymic nude mice immunized with alum-adsorbed OVA. On the other hand, immunization of the recipient mice with OVA-liposome did not induce anti-OVA IgE production, even when CD4(+) T-cells of mice immunized with alum-adsorbed OVA were transferred. In the secondary immune response, OVA-liposome enhanced anti-OVA IgG antibody production but not the ongoing IgE production, suggesting that the IgE-selective unresponsiveness induced by the liposomal antigen involved direct effects on IgE but not IgG switching in vivo. These results suggest the role of an alternative mechanism, one not involving T-cells, in the regulation of IgE synthesis, and raise the possibility that surface-linked liposomal antigen is potentially applicable for the development of a novel vaccine that induces the least IgE synthesis. Moreover, given the relatively low allergic response to and increased antigenicity of the allergen, this form of antigen preparation would be applicable to allergen immunotherapy.     

The Role of IgG1 and IgG2 in Trimellitic Anhydride-Induced Allergic Response in the Guinea Pig Lung

Trimellitic anhydride (TMA) is a small molecular weight chemical used in the paint and plastics industry that can cause asthma-like symptoms in humans. Guinea pigs sensitized intradermally with TMA will respond to antigen challenge with asthma-like symptoms, including an immediate bronchoconstriction and a delayed cellular infiltration into the lung, particularly eosinophil infiltration. Sensitized guinea pigs produce TMA-specific IgG1, which is thought to be important in asthmatic reactions in this animal model; however, they also produce TMA-specific IgG2 antibody. The purpose of the present study was to determine the role of IgG1 and IgG2 in the TMA-induced immediate bronchoconstriction and delayed cellular infiltration in the guinea pig. Guinea pigs were passively sensitized by intratracheal instillation of TMA-specific IgG2, an antibody preparation enriched with TMA-specific IgG1, or a combination of the two. The allergic response was induced by intratracheal instillation of TMA conjugated to guinea pig serum albumin (TMA–GPSA). A significantly greater bronchoconstrictor response was observed in animals sensitized with a combination of the IgG2 and IgG1 preparation compared to those sensitized with IgG2 or the IgG1 preparation alone. Cellular infiltration was quantified 24 h after antigen challenge by differential cell counts of bronchoalveolar lavage (BAL) cells as well as by using eosinophil peroxidase (EPO) and myeloperoxidase (MPO) activity as a measure of the numbers of eosinophils and neutrophils, respectively. In the BAL, passively sensitizing with IgG2 alone resulted in an increase in both TMA-induced MPO and EPO activity. In contrast, in the lung, passively sensitizing with a partially purified preparation of TMA-specific IgG1 alone resulted in a significant increase in TMA-induced EPO activity. Passively sensitizing with IgG2 in conjunction with the IgG1 preparation resulted in an enhanced cellular infiltration and lung injury over that seen with either antibody preparation alone. These data demonstrate an augmentation of IgG1-mediated responses by the addition of IgG2 and suggest a significant role for both subclasses of IgG antibodies in this guinea pig model of TMA-induced occupational asthma.