IgE binding to unique hazelnut allergens: Identification of non pollen-related and heat-stable hazelnut allergens eliciting severe allergic reactions

Summary Background: Usually hazelnut allergic patients suffer from the tree pollen associated oral allergy syndrome (OAS) caused by cross-reactive structures. Anaphylactic reactions elicited by hazelnuts happen rarely but are of high clinical significance. Considering that hazelnuts are ingredients in processed foods, hazelnuts may play an important role as hidden allergens for these high risk patients. Therefore, we analyzed the IgE reactivity of a young woman with severe allergic reactions after ingestion of hazelnuts without any association to tree pollen allergy. Aim of the study: The aim of this study was to identify and characterize these potent hazelnut-specific allergens. We compared these allergens to structures displayed by sera from patients with a completely or partially non pollen-related hazelnut allergy and with birch pollen-related hazelnut allergy. None of the sera had a clinical history of anaphylaxis. Special emphasis was placed on the heat stability and cross-activity of these allergens. Methods/Results: Using Western blotting with extract from birch pollen and EAST inhibition techniques we were able to show that the allergens in the serum sample of the young woman were not cross-reactive with birch pollen. Immunoblot experiments with extracts from native and heated hazelnuts and EAST inhibition tests further characterized these allergens to be heat-stable. Unlike the IgE binding pattern of the sera from the patients with pollen-related hazelnut allergy, low molecular weight proteins below 10 kDa were identified by the sera from the patients without pollinosis. Conclusions: Since the binding pattern of the serum sample of the young woman was different from that of the sera from patients without pollen allergy but less severe symptoms, we assume an association between single non pollen-dependent hazelnut allergens in the low molecular range and severe allergic reactions. These results enable us to approach a subgroup of hazelnut allergens which we believe to be responsible for anaphylactic reactions in hazelnut allergic patients after ingestion of heat-stable hazelnut structures in processed food stuff, independent of pollinosis. Received: 14 March 2000, Accepted: 13 July 2000     

Genetic engineering of trimers of hypoallergenic fragments of the major birch pollen allergen, Bet v 1, for allergy vaccination

An immunotherapy trial performed in allergic patients with hypoallergenic recombinant fragments, comprising aa 1-74 and 75-160 of the major birch pollen allergen, Bet v 1, has indicated that the induction of allergen-specific IgG responses may be an important mechanism of this treatment. To investigate whether the immunogenicity of the rBet v 1 fragments can be increased, recombinant trimers of the fragments were produced. For this purpose, DNA trimers of rBet v 1 aa 1-74 as well as of rBet v 1 aa 75-160 were subcloned into expression plasmid pET 17b, expressed in Escherichia coli and purified. The fragments as well as the fragment trimers showed a reduced IgE-binding capacity and allergenic activity compared to rBet v 1 wildtype when tested in allergic patients. Both rBet v 1 aa 75-160 monomer and trimer induced high titers of allergen-specific IgG1 Abs in mice. Interestingly, rBet v 1 aa 1-74 trimer induced a much higher IgG₁ response to rBet v 1 than rBet v 1 aa 1-74 monomer. Consequently, IgG Abs induced with the rBet v 1 aa 1-74 trimer inhibited birch pollen allergic patients’ IgE-binding 10-fold more efficiently than IgG Abs induced with the monomer. Our data show that the immunogenicity of allergy vaccines can be increased by oligomerization.     

Neonatal colonization of germ-free mice with Bifidobacterium longum prevents allergic sensitization to major birch pollen allergen Bet v 1

The main goal in reversing the allergy epidemic is the development of effective prophylactic strategies. We investigated the prophylactic effect of neonatal mother-to-offspring mono-colonization with Bifidobacterium longum ssp. longum CCM 7952 on subsequent allergic sensitization. Adult male and female germ-free (GF) mice were mono-colonized with B. longum, mated and their offspring, as well as age-matched GF controls, were sensitized with the major birch pollen allergen Bet v 1. Furthermore, signaling pathways involved in the recognition of B. longum were investigated in vitro. Neonatal mono-colonization of GF mice with B. longum suppressed Bet v 1-specific IgE-dependent β-hexosaminidase release as well as levels of total IgE and allergen-specific IgG2a in serum compared to sensitized GF controls. Accordingly, Bet v 1-induced production of both Th1- and Th2-associated cytokines in spleen cell cultures was significantly reduced in these mice. The general suppression of Bet v 1-specific immune responses in B. longum-colonized mice was associated with increased levels of regulatory cytokines IL-10 and TGF-β in serum. In vitro, B. longum induced low maturation status of bone marrow-derived dendritic cells and production of IL-10 in TLR2-, MyD88-, and MAPK-dependent manner. Our data demonstrate that neonatal mono-colonization with B. longum reduces allergic sensitization, likely by activation of regulatory responses via TLR2, MyD88, and MAPK signaling pathways. Thus, B. longum might be a promising candidate for perinatal intervention strategies against the onset of allergic diseases in humans.     

Efficacy of sublingual vectorized recombinant Bet v 1a in a mouse model of birch pollen allergic asthma

Background

Second generation sublingual allergy vaccines based upon recombinant allergens combined with vector systems are being developed as an alternative to conventional allergen extracts. Herein, we evaluated the efficacy of a recombinant form of the major allergen Bet v 1a (rBet v 1a) formulated as a mucoadhesive particle in a preclinical model of birch pollen (BP) respiratory allergy.

Materials and methods

BALB/c mice were sensitized to BP extracts by intraperitoneal injections followed by aerosol exposures. Sensitized mice underwent sublingual immunotherapy (SLIT) twice a week for eight weeks with either a BP extract or rBet v 1a formulated in amylopectin-based microparticles (MPA). SLIT efficacy was assessed using whole body plethysmography, lung histology and cell counts in broncho-alveolar lavages (BAL) as read outs. BP and/or rBet v 1a-specific T cell and antibody responses were monitored in lung and serum, respectively. IgA levels were measured in saliva.

Results

Mice sensitized to BP exhibit chronic airway hyperresponsiveness (AHR), lung inflammation (documented by compliance and resistance measurements), eosinophil infiltrates in BAL, as well as Bet v 1-specific Th2 biased responses. Both SLIT with soluble rBet v 1a (50 μg/dose) and BP extract (equivalent to 50 μg rBet v 1 per dose) lead to a significant reduction in AHR, lung eosinophilia and Th2 responses. A sub-optimal dose of 5 μg of rBet v 1a displays a similar level of efficacy with a significant decrease of Th2 responses when formulated with MPA microparticles. In addition, allergen vectorization with mucoadhesive particles allows a faster reduction in AHR in sensitized animals.

Conclusion

We demonstrate in a murine model of chronic BP respiratory allergy the efficacy of SLIT with vectorized rBet v 1a. Thus, combining recombinant allergens with mucoadhesive vector systems paves the ground for improved second generation sublingual allergy vaccines.     

Enzyme immunoassays for total and allergen specific IgE in population studies.

OBJECTIVE--Extensive IgE serology in occupational or environmental health studies is often hampered by a lack of technical facilities and finance. The use in population studies of relatively simple and inexpensive enzyme immunoassays (EIAs) was therefore evaluated for the assessment of total serum immunoglobulin E (IgE), and of specific IgE reactions with various common (house dust mites, grass and birch pollen, and cat) or occupational (fungal alpha-amylase and rat urinary protein) allergens. METHODS--Total IgE was measured with a sandwich EIA, calibrated with commercially available IgE standards. Reproducibility was studied by testing pooled normal human serum samples in each of a large series of test plates. A panel of 156 children's serum samples with known IgE values was used to compare the assay with other total IgE assays. A previously developed EIA for anti-yeast IgE was adapted for the measurement of IgE reacting with various common and occupational allergens. Binding of IgE to microwells coated with commercially available allergen extracts, or allergen preparations from our own laboratory, was measured with a monoclonal anti-human IgE antibody and subsequent incubations with biotinylated rabbit anti-mouse Ig and avidin-peroxidase. Panels of serum samples from school children (n = 116), bakery workers (n = 126), and laboratory animal workers (n = 52) were used to study sensitivity and specificity, with reference to skin prick tests as the standard, and to compare the EIAs with commercially available test kits. RESULTS--The detection limit of the EIA for total IgE was 0.5-1 kU/l for undiluted serum samples, and the coefficient of variation between assays was less than 15% at serum concentrations between 1 and 150 kU/l. Results obtained with the panel of 156 children's serum samples were strongly correlated (r2 = 0.86) with IgE concentrations measured previously by radioimmunoassay. The results of the EIA for various occupational allergens correlated very well, both qualitatively and quantitatively, with the results of commercial test kits. Sensitivity and specificity of the EIA results as a predictor of skin prick test reactivity towards common allergens (house dust mite, grass pollen, birch pollen, and cat) were remarkably high (> 80%-90%) in the series of 116 children's serum samples. In a population of bakery workers the specificity of the EIAs was also very high (> 90%). The sensitivity was notably lower (30%-70%) in this adult population, which is, however, in agreement with results reported for conventional IgE tests. CONCLUSION--As the costs were estimated to be at least five to 10-fold lower than those of commercial test kits, the EIAs for total and specific IgE may be very useful tools in epidemiological studies of atopic respiratory or other disorders.     

Mucosal co-application of lactic acid bacteria and allergen induces counter-regulatory immune responses in a murine model of birch pollen allergy

Recent epidemiological studies and clinical trials suggest a possible role of certain lactic acid bacteria (LAB) strains in the prevention of allergic diseases. In this study, we aimed at evaluating the immunomodulatory potential of two LAB strains, Lactococcus lactis and Lactobacillus plantarum, for prophylaxis and therapy of allergic immune responses. Both LAB strains-induced high levels of IL-12 and IFN-gamma in naive murine spleen cell cultures. Intranasal co-application with recombinant Bet v 1, the major birch pollen allergen, prior or after allergic sensitization, led to increased levels of allergen-specific IgG2a antibodies and in vitro IFN-gamma production, indicating a shift towards Th1 responses. Successful immunomodulation by the mucosal pre-treatment was further demonstrated by suppression of allergen-induced basophil degranulation. We conclude that these LAB strains in combination with an allergen could be promising candidates for mucosal vaccination against type I allergy.     

Periplaneta americana arginine kinase as a major cockroach allergen among Thai patients with major cockroach allergies

Periplaneta americana is the predominant cockroach (CR) species and a major source of indoor allergens in Thailand. Nevertheless, data on the nature and molecular characteristics of its allergenic components are rare. We conducted this study to identify and characterize the P. americana allergenic protein. A random heptapeptide phage display library and monoclonal antibody (MAb) specific to a the P. americana component previously shown to be an allergenic molecule were used to identify the MAb-bound mimotope and its phylogenic distribution. Two-dimensional gel electrophoresis, liquid chromatography, mass spectrometry, peptide mass fingerprinting, and BLAST search were used to identify the P. americana protein containing the MAb-specific epitope. We studied the allergenicity of the native protein using sera of CR-allergic Thai patients in immunoassays. The mimotope peptide that bound to the MAb specific to P. americana was LTPCRNK. The peptide has an 83-100% identity with proteins of Anopheles gambiae, notch homolog scalloped wings of Lucilia cuprina, delta protein of Apis mellifera; neu5Ac synthase and tyrosine phosphatase of Drosophila melanogaster, and a putative protein of Drosophila pseudoobscura. This finding implies that the mimotope-containing molecule of P. americana is a pan-insect protein. The MAb-bound protein of P. americana was shown to be arginine kinase that reacted to IgE in the sera of all of the CR-allergic Thai patients by immunoblotting, implying its high allergenicity. In conclusion, our results revealed that P. americana arginine kinase is a pan-insect protein and a major CR allergen for CR-allergic Thai patients.     

Iron-induced chelation alleviates the potential allergenicity of ovotransferrin in a BALB/c mouse model

Ovotransferrin (OVT) is one of the main egg allergens with 2 iron-binding sites. Several studies have demonstrated that iron-chelation decreased the allergenicity of milk allergen and birch pollen allergens. Therefore, we hypothesized that iron-chelation could also reduce the allergenicity of OVT. Apo-OVT (iron-free OVT, the natural state in egg white) and Holo-OVT (iron-chelated OVT) were prepared, and the allergenicity of them were assessed and compared using a BALB/c mouse model as well as dendritic cells (DCs) based on antigen uptake. Mice were orally sensitized with Apo-OVT or Holo-OVT using cholera toxin as adjuvant. Clinical signs of allergy, morphological structure of jejunum, specific antibody levels, mast cell protease-1 (mMCP-1) concentrations, cytokines and antigen uptake by DCs were determined after the mice were challenged with Apo-OVT or Holo-OVT. Results showed that both Apo-OVT and Holo-OVT induced intestinal allergy, but no systematic allergic symbols were observed. Serum levels of mouse mast cell protease-1 (mMCP-1) and specific IgE in Apo-OVT group were lower than in control group, and no significant difference between Apo-OVT group and Holo-OVT group (P > .05). The levels of OVT-specific IgG and IgG1, as well as the Th-1 cytokine interferon gamma and Th2-type cytokine interleukin-13 in Holo-OVT sensitized mice were significantly decreased compared to Apo-OVT group (P < .05), while no significant difference with control group (P > .05). However, DCs took in less Apo-OVT than Holo-OVT. Overall, iron-induced chelation could alleviate the potential allergenicity of OVT in vivo.     

海口地区138例哮喘患儿血清总IgE水平及过敏原检测分析

目的:探讨海口地区哮喘患儿血清中总IgE水平及主要过敏原种类。方法:采用德国欧蒙印迹法分别检测138例哮喘患儿及40例健康儿童的过敏原种类,并采用美国贝克曼化学发光仪DXI800检测血清中总IgE含量。结果:40例健康儿童血清总IgE含量为116.29±91.60μg/L,过敏原检测均为阴性;138例哮喘患儿总IgE的含量为2 126.17±1 942.69μg/L,138例哮喘患儿总IgE阳性率为73.2%,总IgE水平明显高于对照组过敏原阳性率68.1%(P<0.01),且大多数患儿被多种过敏原致敏。吸入组最高为屋尘57例,其次为屋尘螨/粉尘螨46例;食入组最高为虾20例,其次为鸡蛋白、蟹。结论:通过对哮喘患儿测定血清总IgE水平,并结合过敏原的测定,可了解患儿过敏的变应原,寻找病因,尽量避免接触此类过敏原,减少发病。     

Effects of atmospheric pollutants (CO,O3, SO2) on the allergenicity of Betula pendula,Ostrya carpinifolia,and Carpinus betulus pollen

Pollen of Betula pendula, Ostrya carpinifolia, and Carpinus betulus was exposed in vitro to relatively low levels of the air pollutants, namely carbon monoxide, ozone, and sulfur dioxide. The allergenicity of the exposed pollen was compared with that of non-exposed pollen samples to assess if air pollution exposition affects the allergenicity potential of pollen. The immunodetection assays indicated higher IgE recognition by all sera of allergic patients to the pollen protein extracts in all exposed samples in comparison to the non-exposed samples. These results show that the pollen exposition to low pollutants’ levels induces increased allergic reaction to sensitized individuals.     

惠州地区204例儿童慢性荨麻疹血清特异性IgE抗体检测及过敏原分析

目的分析惠州地区儿童慢性荨麻疹患者血清总IgE抗体及特异性IgE抗体水平,了解过敏原分布情况。方法入选2012年1月至2013年12月儿童慢性荨麻疹204例,应用IgE抗体特异性过敏原检测试剂盒(ELISA法)检测血清IgE特异性抗体。结果 204例慢性荨麻疹患儿中总IgE阳性198例,阳性率为97.1%,196例患儿检测出过敏原,阳性率为96.1%,其中食入组主要为蛋黄46例(22.5%),牛奶42例(20.6%),蟹、虾40例(19.6%);吸入组主要为花粉、屋尘、尘螨76例(37.3%),蟑螂52例(25.5%)、猫狗毛发皮屑44例(21.6%)。食入组及吸入组的阳性率无显著性差异(P>0.05)。不同特异性过敏原在各阳性级别均有分布,总IgE均为(+++)级或(++)级。结论多数儿童慢性荨麻疹患者同时出现多种过敏原,其中主要食入因素有蛋黄、牛奶、蟹、虾,主要吸入因素主要为花粉、尘螨、蟑螂、猫狗毛发皮屑。     

Functionality of endogenous folates from rye and orange juice using human <Emphasis Type="Italic">in vivo</Emphasis> model

Summary. Background: Cereals contribute about a quarter of the daily folate intake from a typical diet in several European countries. However, studies on bioavailability of endogenous folates, in particular of cereal sources, are scarce. Aim of the study: We aimed to study how well natural folates from rye (different rye breads and muesli made of malted rye) and orange juice function in improving folate status of human volunteers compared to a diet containing folic acid fortified wheat bread. Methods: Healthy human volunteers aged 20–66 y took part in a four-week intervention trial in which bread, breakfast cereal and juice were provided. The study had a parallel design with two groups, 1) rye and orange juice group (33 volunteers) and 2) fortified wheat bread and apple juice group (31 volunteers). The test foods provided on average 184 μg and 188 μg folate per day in rye and wheat groups, respectively. Test foods were consumed as part of the subjects' normal diet. Results: In both groups statistically significant increases in serum and red cell folates were observed after the intervention period. The serum folate increased 26 % and 31 %, and red cell folate levels increased 17 % and 15 % in rye and orange juice and wheat and apple juice groups, respectively. The effects did not differ between the rye and wheat groups. Increases in serum and red cell folate were more profound among subjects with low starting folate levels. Decrease in the plasma homocysteine concentrations was observed only in the highest tertile of both groups but not in the group means. Conclusions: Endogenous folates incorporated into a healthy diet, even in moderate amounts, is an efficient way to improve folate status among healthy adults. Folates from different rye products and orange juice showed good bioavailability that was similar to folic acid from fortified white bread. Received: 19 July 2002, Accepted: 6 October 2002 Correspondence to: Dr. Liisa Vahteristo     

Impacts of climate change on plant food allergens: a previously unrecognized threat to human health

Global climate change has had, and will continue to have, many significant impacts on biological and human systems. There are now many studies of climate change impacts on aeroallergens, particularly pollen, including a study demonstrating significant increases in the major allergen content of ragweed pollen as a function of rising atmospheric carbon dioxide concentration ([CO₂]). Recent research has also demonstrated more allergenic poison ivy in response to elevated [CO₂]. Here, we suggest, for the first time, the potential for global climate change, and, in particular, increased [CO₂] and temperature, to have an impact on the allergenicity of plant food allergens such as peanut. Such impacts could have significant impacts on associated allergic diseases, and pose a previously unrecognized threat to human health. There is an urgent need for research on the impacts of climate change on plant food allergens.     

Identification of airborne pollen allergens from two avenue trees of India

An attempt has been made to detect airborne pollen of Lagerstroemia speciosa (LS) and Spathodea campanulata (SC) – two common avenue trees of India as potential sources of aeroallergens and also to identify the major IgE-reactive components present in them. The airborne pollen concentration was assessed using a Burkard sampler. A detailed questionnaire on clinical data of 1490 patients was recorded based on hospital data. We assessed the allergenicity of pollen by in vivo and in vitro tests. The correlation among meteorological factors, pollen seasons and allergenic potency of patients was assessed by multiple regression analysis. The sensitivity of patients to pollen antigens was highly correlated with pollen seasons. In SDS–PAGE, 15 protein bands were detected from LS pollen, while 14 bands from SC. The IgE-specific immunoblotting with patients’ sera allergic to LS displayed five major allergens, while four major allergens were detected from SC. This would be the first report from India to prove the allergenic potentiality of airborne pollen of these two common avenue trees of India.     

Multinational study of major breast milk carotenoids of healthy mothers

Summary. Background: Carotenoids in serum vary between countries and within populations with evidence suggesting a qualitative relationship to diet. Breast milk carotenoids furnish a source of vitamin A and potentially provide immunoprotection and other health benefits for infants. There have been numerous studies of milk carotenoid concentrations in undernourished populations; however, carotenoid concentrations have not previously been compared in populations of well-nourished mothers. Aim of Study: To compare concentrations of five major carotenoid groups: α-carotene, β-carotene, β-cryptoxanthin, lutein/zeaxanthin, and lycopene in breast milk of healthy women from Australia, Canada, Chile, China, Japan, Mexico, the Philippines, the United Kingdom, and the United States, and to qualitatively compare patterns of dietary intake with milk carotenoid concentrations. Methods: Breast milk collected from healthy lactating women was analyzed for concentrations of five carotenoids and retinol and quantitated relative to total milk lipid. All determinations were performed in a single research laboratory using standardized methodology. Mothers consumed their usual diets and provided a single 24-h dietary recall. Results: Breast milk carotenoid concentrations varied greatly among countries, with the greatest differences in β-cryptoxanthin (∼ 9-fold) and the least in α-carotene and lycopene (∼ 3-fold). Breast milk retinol concentrations varied ∼ 2-fold across countries. The provitamin A carotenoids α-carotene, β-carotene, and β-cryptoxanthin as a group accounted for > 50 % of the carotenoids measured. Total breast milk carotenoids were highest in Japanese and lowest in Philippine mothers. Breast milk β-carotene concentrations were highest in Chile and lowest in the Philippines. Conclusions: Patterns of breast milk carotenoids were unique to each country and qualitative patterns reflected the dietary carotenoid supply. Received: 24 June 2002, Accepted: 12 December 2002 Supported by a grant from Wyeth Nutrition, Philadelphia, PA, USA. Correspondence to: Louise M. Canfield     

Skin reactivity to grass pollen and the mite Dermatophagoides pteronyssinus in the prick-test using 2 types of allergen preparations

The reliability of prick testing with allergen preparations produced in Yugoslavia was assessed by comparison to the internationally accepted ones (Pharmacia Diagnostica) of 100,000 BU activity. The following allergens were used: the mite Dermatophagoides pteronyssinus in 107 subjects, and grass pollens: Dactylis glomerata, Poa pratensis, Phleum pratense and Secale cereale in 39 subjects. The clinical significance of the response was assessed in relation to the skin reaction to the negative control solution and positive histamine control (in concentration of 1 mg/ml). The results indicate that the allergen extract of Dermatophagoides pteronyssinus (3000 PNU/ml) produced in Yugoslavia does not provoke skin reactions comparable to those provoked by the standardized extract of 100,000 BU in contrast to the grass pollen allergens, with the exception of Poa pratensis, which evokes equivalent skin reactions. Thus grass pollen allergens are reliable extracts and can be applied with good confidence in routine work. The need to set more rigorous criteria for assessing skin prick reactions, when domestic, unstandardized products are used, emerged as a result of this study.     

Use of a genetic cholera toxin B subunit/allergen fusion molecule as mucosal delivery system with immunosuppressive activity against Th2 immune responses

Induction of peripheral tolerance can be facilitated when the antigen is linked to the B subunit of cholera toxin (CTB), an efficient mucosal carrier. In the present study, a genetic fusion molecule of Bet v 1 and CTB was produced to test whether mucosal application of this construct would lead to suppression of Th2 responses. Intranasal pretreatment of BALB/c mice with rCTB-Bet v 1 prior to allergic sensitisation with the allergen significantly decreased IgE but markedly increased allergen-specific IgG2a levels in sera as well as IFN-gamma production of splenocytes. This Th1 shift was supported by an increased T-bet/GATA3 mRNA ratio. IL-5 production within the airways was suppressed after the pretreatment with rCTB-Bet v 1, while local allergen-specific IgA antibodies were markedly enhanced by pretreatment with the construct. Upregulation of Foxp3, IL-10 and TGF-beta mRNA expression was detected in splenocytes after pretreatment with unconjugated allergen but not with the fusion molecule, indicating that antigen conjugation to a mucosal carrier modifies the immunomodulating properties of an antigen/allergen.     

A side-by-side comparison of sampling methods for settled, indoor allergens

Exposure to indoor allergens is associated with asthma, but there is no standardized sampling method for measuring allergens. We compared the association of measured allergen exposure and serum-specific IgE levels and the precision of three sampling methods (Cyclone, Mighty Mite, and Readivac II) to identify a standardized sampling method for indoor allergens. A random sample of 72 children, 5 to 17 years old, with doctor-diagnosed asthma who lived in the same residence >or=2 years were enrolled. Composite, side by side floor samples were obtained with all three methods. Dust allergen concentrations and serum-specific IgE levels were measured for Der f I, Fel d I, and Bla g I. Mean allergen concentration did not differ significantly by sampling method. Cat allergen was significantly correlated with serum-specific IgE for Cyclone (P=0.003) and Mighty Mite (P=0.008), but only marginally for Readivac II (P=0.07). Dust mite allergen was significantly correlated with serum-specific IgE for Readivac II (P=0.02) and Cyclone (P=0.038), but not for Mighty Mite (P=0.12). Cockroach allergen was not correlated with serum-specific IgE for any sampling method. In multiple linear regression, cat allergen was associated with serum-specific IgE for Cyclone (P=0.007) and Mighty Mite (P=0.02), but not for Readivac II (P=0.06). In contrast, dust mite allergen was marginally associated with serum-specific IgE for Readivac II (P=0.07), but not for Mighty Mite (P=0.64) or Cyclone (P=0.27). The Cyclone and Mighty Mite were more precise than Readivac II for cat allergen, but there was no difference for dust mite allergen (P>0.05). No single method is superior for measurement of indoor allergens. In general, cat allergen collected with the Cyclone was a better predictor of serum-specific IgE levels to Fel d I, whereas dust mite allergen collected with the Readivac II was a better predictor of serum-specific IgE levels to Der f I.     

Segregation analysis of the specific response to allergens: a recessive major gene controls the specific IgE response to Timothy grass pollen

Segregation analysis of the specific response to allergens (SRA) was performed in a sample of 234 randomly selected Australian families using the regressive models. Various SRA phenotypes were considered using broad and narrow definitions of these phenotypes, according to the type of test used, skin test or RAST test, and the specificity of the response to allergen. Strong evidence for familial dependencies among blood relatives was shown for most SRA phenotypes, especially when using a broad definition. There was no evidence for a Mendelian factor accounting for the familial transmission of these broadest phenotypes, which may involve multiple factors preventing the clear detection of a major effect with Mendelian transmission. However, segregation of a Mendelian recessive major gene was detected for one SRA sub-phenotype, the IgE response to a single allergen, Timothy grass pollen, measured by the RAST test. Identification of a specific SRA phenotype controlled by a major gene may have important implications for further linkage studies.     

Antiproliferative effect of antioxidant polyphenols from grape in murine Hepa-1c1c7

Summary. Background: Grapes and wine contain high concentrations of polyphenolic compounds. Although their cancer protective effect has been well documented, their activity as anticarcinogens should be cautiously considered since the molecular bases of action and their applicability to human cancer prevention are still unclear. Aim of the study: We studied the antioxidant/antiradical activity and the antiproliferative effect in vitro of different polyphenolic mixtures, extracted from grapes and fractionated through RP-HPLC. Methods: The polyphenolic fractions were chemically characterized and their antioxidant/antiradical activity was determined by the DPPH assay. Mouse hepatoma Hepa-1c1c7 cells were used to study the cell growth inhibition capacity of these fractions by MTT assay. Their capacity of altering cell cycle and possible induction of apoptosis was examined using FACS analysis. Results: The original polyphenolic fraction OW, which contained gallic acid (GA), (+)-catechin (Cat), (−)-epicatechin (Ec), glycosylated flavonols (F) and procyanidin oligomers was fractionated into fraction I, composed of monomers and small oligomers, and fraction II that included flavonols and procyanidin oligomers of higher molecular weight. The three polyphenolic fractions tested showed quite similar antiradical activity, although fraction I was the most potent antiradical agent (lowest ED₅₀ value: 9 μg). Fraction II was the least potent cell growth inhibitor (highest IC₅₀ value: 100 μg/ml) but showed the strongest effect on the cell cycle of Hepa-1c1c7, inducing apoptosis in those cells. The original fraction OW was demonstrated to have the most potent cell growth inhibition effect (lowest IC₅₀ value: 43 μg/ml). However, it only appeared to alter cell cycle of Hepa-1c1c7 at concentrations higher than its IC₅₀ and did not induce apoptosis in those cells. A similar effect on cell cycle and apoptosis was encountered for fraction I. Conclusions: The polyphenolic fractions tested in this study were potent antiradical agents and exerted an antiproliferative effect in mouse hepatoma Hepa-1c1c7 cells; the fraction with the highest degree of polimerization and galloylation (fraction II) had the most influence on the cell cycle and induction of apoptosis on Hepa-1c1c7. Received: 6 July 2002, Accepted: 24 July 2002 Correspondence to: Dra. Marta Cascante Serratosa