植物过敏蛋白研究进展

过敏反应是人类常见的一类自身免疫疾病 ,由于症状复杂 ,种类繁多 ,危害广泛且很难根治 ,所以长期以来一直是医学界的一大难题。自然界的过敏原种类很多 ,花粉蛋白质和食物蛋白质是植物过敏原中最为常见的两种过敏原。许多人在直接或间接接触这些过敏蛋白质之后会因过敏而患上支气管哮喘、上呼吸道结膜炎、湿疹、皮炎等症。特别是一些谷类食品加工业的工人 ,由于经常呼吸带食物蛋白质的粉末而极易患过敏性疾病。1 .过敏原引起过敏反应的过敏原大都是外源分子 ,并且能刺激产生显著的免疫反应。许多已知的植物过敏原与病理发生相关蛋白质(ｐａ…     

高等植物的过敏原研究

高等植物中的过敏原是引起人类过敏反应的重要来源,人们对于这个领域的研究已有较长的历史。目前,研究手段正日趋成熟和多样化,研究方向已着眼于改造植物过敏原。本文结合本实验室的初步工作,介绍了这个研究领域的历史、现状、最新进展以及发展前景。     

花生过敏原iso-Ara h 3的克隆、表达和免疫学鉴定

采用Touchdown PCR技术从花生cDNA中克隆到花生的过敏原iso-Ara h 3基因,并进行重组蛋白表达,再用Western blot技术鉴定重组蛋白过敏原性。结果显示,构建的pET44a-iso—Ara h 3重组菌能表达iso—Ara h 3蛋白。用8例花生过敏的阳性血清鉴定表明,重组的iso—Ara h 3蛋白血清IgE识别率为12．5％,是一种低过敏原性的过敏原蛋白。     

牛奶过敏症状明显而特异性IgE抗体呈阴性样本的研究

以对牛奶有强烈临床过敏症状而特异性IgE抗体呈阴性的样本为研究对象，探究临床症状与测试结果的相关性。通过梯度稀释样本，检测粗提取物的过敏原及其常见组分过敏原项目，分析结果的相关性。牛奶粗提取物的测试结果与临床症状及组分的测试结果明显不符，呈阴性，其各组分的测试值也反映粗提取物是假阴性。临床判断患者是否对某种过敏原过敏，不仅要依靠粗提取物的测试结果，而且还需要结合临床症状进行分析，有条件的情况下对粗提取物的组分进行过敏原测试，才能获得正确的临床诊断。     

食物过敏原研究进展

食物过敏反应是世界各地普遍存在的一个严重问题。本文综述了目前有关食物过敏原研究所取得的进展。包括食物过敏反应流行病学数据,食物过敏原,新过敏原的引物及其监控,通过消除内源基因根除过敏性,移去过敏性抗原基等。     

西安地区儿童支气管哮喘吸入性过敏原的调查分析

目的:探讨西安地区儿童支气管哮喘吸入性过敏原的分布情况。方法:选择950例来自西安地区的支气管哮喘患儿为研究对象,采用过敏原皮肤点刺试验检测,以组胺作为阳性对照,生理盐水为阴性对照,分析不同年龄和性别的患儿过敏原的分布情况。结果:950例支气管哮喘患儿中,384例皮肤点刺过敏原检测呈阳性,占40.4%,男女患儿过敏原检测阳性分布无明显差异(P0.05);尘螨为主要的过敏原,其次为艾蒿和霉菌类;随着患儿年龄的增加,其过敏原检测的阳性率明显升高(P0.05),且大多数过敏原检测阳性患儿至少合并2-3种过敏原阳性。结论:西安地区支气管哮喘患儿吸入性过敏原阳性率与其性别无关,但与其年龄有关,过敏原以尘螨类为主,大多数检测阳性的患儿对至少一种以上的过敏原阳性。     

实时荧光PCR检测食品中致敏原芥末成分

对植物原料产品和植物源性食品中芥末成分的快速鉴定是避免过敏性疾病发生的重要措施。依据芥末Sin A1管家基因的核酸序列设计特异性引物和探针,对3种芥末样品和21种非芥末植物样品进行实时荧光PCR检测,结果显示,过敏原芥末管家基因的样品FAM通道有荧光信号检出,非芥末样品FAM通道均无荧光信号检出。灵敏度实验表明,植物原料产品中对芥末的检测低限可达到1 mg/kg。此外对市售的芥菜籽等样品和深加工的芥末致敏原参考物质(葡萄糖)进行实际样品的检测,均能很好检出致敏原芥末成分。     

东亚飞蝗主要过敏原的分析、鉴定与纯化

通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分离东亚飞蝗Locusta migratoria manilensis(Meyen)的蛋白质组分并测定其分子量,收集过敏病人血清,采用免疫印迹(Western—blotting)法鉴定其过敏原成分,通过凝胶过滤层析对东亚飞蝗过敏原进行分离纯化。结果表明:东亚飞蝗蛋白粗提液条带大概有30条左右,其中主带大约有10条,相对分子量约为13、15、25、28、40、45、55、70、100、110ku,其中蛋白含量最丰富的约在70ku左右。免疫印迹结果显示,蝗虫过敏条带主要有5条,相对分子量分别约为19、29、38、70、130ku。通过凝胶过滤层析对东亚飞蝗过敏原进行分离纯化,得到了一个高纯度相对分子质量约为70ku东亚飞蝗过敏原,并且发现了一个相对分子质量约为130ku的蝗虫新过敏原。本研究为临床上蝗虫食物变态反应性疾病的诊断和治疗奠定基础。     

实验动物致敏研究进展

实验动物致敏（laboratory Animal Allergy,LAA）,是一种职业过敏性疾病,造成人的呼吸道及皮肤发生炎症。该病在国外研究较多,过敏源是动物皮毛、尿液、唾液中的一类酸性小分子蛋白质;可以通过皮肤试验、放射过敏吸附试验及ELISA等方法检测易感人员。对易感人员可以通过控制环境中的过敏原来保护。目前国内尚无专门机构研究此病症,笔者综述了该职业性疾病的症状、机理、控制方法及国内外对该病症认识上的差异。     

尘螨过敏原的交叉反应性

尘螨是最主要的室内过敏原之一,随着社会生活的现代化,人们对室内居住环境的要求越来越高,由于环境因素而引起的一些过敏性疾病越来越受到人们的关注。本文主要综述了近几年来尘螨过敏原与其他多种螨类、软体动物（蜗牛）和甲壳类动物（虾）以及昆虫（蟑螂、衣鱼、摇蚊、石蚕蛾）等过敏原之间的交叉反应性,对临床过敏性疾病的诊断具有参考价值。     

植物过敏性蛋白质及其生物学功能

在引起I型超敏反应的变应原中 ,植物的花粉、果实和汁液可以分别作为吸入性变应原 (inhalentallergen)、食入性变应原 (ingestentallergen)、接触性变应原 (contactentallergen)使过敏者患上鼻炎、哮喘、枯草热等疾病。而其中引起这些超敏反应的植物类蛋白质本身在植物体内亦行使着特定的生物学功能。对这些植物类过敏性蛋白质的研究不仅在植物学本身研究中具有一定意义 ,同时在变态反应性疾病的免疫治疗中亦具有重要的应用价值。目前 ,这类涉及植物学、免疫学和变态反应学的研究逐渐形成了一个新的交叉研究领域。     

Mapping of fruit allergens by 2D electrophoresis and immunodetection

Proteomic analyses of fruits are confronted with a series of specific obstacles: a general low protein content in plant tissues, allergen extraction from highly complex matrices and protein determination in the presence of interfering compounds. Different methods are currently being introduced to achieve higher protein yields and a simultaneous removal of interfering substances, such as polyphenols and polysaccharides. However, no universal protocol suitable for protein purification from any given plant species is available. Protein profiling by 2DE-western blotting offers a powerful tool for the detection and characterization of known and novel plant allergens. Moreover, the detection of IgE-reactive proteins from fruits is improved by combining western blot and alternative visualization techniques. The recent developments in bioinformatics and databases facilitate the interpretation of profiling studies with regard to novel potential fruit allergens.     

Recombinant food allergens

Allergenic (glyco)proteins are the elicitors of food allergies and can cause acute severe hypersensitivity reactions. Recombinant food allergens are available in standardised quantity and constant quality. Therefore, they offer new perspectives to overcome current difficulties in the diagnosis, treatment and investigation of food allergies. This review summarises the expression strategies and characteristics of more than 40 recombinant food allergens that have been produced until today. Their IgE-binding properties are compared to those of their natural counterparts, in addition their application as diagnostic tools, the generation of hypoallergenic recombinant isoforms and mutants for therapeutic purposes, the determination of epitopes and cross-reactive structures are described.     

Pyr c 1, the major allergen from pear (Pyrus communis), is a new member of the Bet v 1 allergen family

Pear is known as an allergenic food involved in the ‘oral allergy syndrome’ which affects a high percentage of patients allergic to birch pollen. The aim of this study was to clone the major allergen of this fruit, to express it as bacterial recombinant protein and to study its allergenic properties in relation to homologous proteins and natural allergen extracts. The coding region of the cDNA was obtained by a PCR strategy, cloned, and the allergen was expressed as His-Tag fusion protein. The fusion peptide was removed by treatment with cyanogen bromide. Purified non-fusion protein was subjected to allergenicity testing by the enzyme allergosorbent test (EAST), Western blotting, competitive inhibition assays, and basophil histamine release. The deduced protein sequence shared a high degree of identity with other major allergens from fruits, nuts, vegetables, and pollen, and with a family of PR-10 pathogenesis related proteins. The recombinant (r) protein was recognised by specific IgE from sera of all pear-allergic patients (n=16) investigated in this study. Hence, the allergen was classified as a major allergen and named Pyr c 1. The IgE binding characteristics of rPyr c 1 appeared to be similar to the natural pear protein, as was demonstrated by EAST-inhibition and Western blot-inhibition experiments. Moreover, the biological activity of rPyr c 1 was equal to that of pear extract, as indicated by basophil histamine release in two patients allergic to pears. The related major allergens Bet v 1 from birch pollen and Mal d 1 from apple inhibited to a high degree the binding of IgE to Pyr c 1, whereas Api g 1 from celery, also belonging to this family, had little inhibitory effects, indicating epitope differences between Bet v 1-related food allergens. Unlimited amounts of pure rPyr c 1 are now available for studies on the structure and epitopes of pollen-related food allergens. Moreover, the allergen may serve as stable and standardised diagnostic material.     

Decrease in Rice Allergenic Proteins of Polished Rice Grains by Incubating with a Miso Solution

The effect of miso on allergenic proteins in rice seeds was investigated. When polished rice grains were incubated at 37°C for 30-120 min with a 10% miso solution, but not with heat-treated miso or 1% NaCl, the amount of soluble proteins extracted from the rice grains with 1 M NaCl markedly decreased. SDS-PAGE, immunoblotting and densitometric analyses of these soluble proteins and insoluble proteins indicate that 26 kDa globulin and 14-16 kDa allergens in the grains were decreased to 15-60% during incubation with the miso solution, especially soybean-koji miso, without any large change in the content of major insoluble proteins.     

Primary structure, recombinant expression, and molecular characterization of Phl p 4, a major allergen of timothy grass (Phleum pratense)

Grass pollen allergy is one of the most important allergic diseases world-wide. Several meadow grasses, like timothy grass and rye grass, contribute to allergic sensitizations, but also allergens from extensively cultivated cereals, especially rye, make a profound contribution. The group 4 allergens are well known as important major allergens of grasses. We have cloned for the first time group 4 sequences from Phleum pratense, Lolium perenne, Secale cereale, Triticum aestivum, and Hordeum vulgare, and investigated the IgE-reactivity of recombinant Phl p 4 as a candidate for allergy diagnostic and therapeutic applications.     

Isolation of pathogen/stress-inducible cDNAs from alfalfa by mRNA differential display

Differential display of mRNA was used to isolate a full-length (SRG1) and a partial (SRG2) alfalfa cDNA induced during infection with the fungal pathogen Colletotrichum trifolii. The deduced amino acid sequences are similar to each other and resemble plant defense-related proteins and tree pollen allergens. SRG1 is a member of a gene family in alfalfa, which may also include the putative defense-related gene PR10. Unlike many defense-related genes described in similar systems, expression of SRG1-like genes does not correlate with resistance to C. trifolii. We speculate SRG1 is induced in response to plant stress.     

Baculoviral infection reduces the expression of four allergen proteins of silkworm pupa

Silkworm (Bombyx mori) larvae are widely used to express exogenous proteins. Moreover, some silkworm pupal proteins can be used as drug‐loading materials for selfexpressed oral tolerance drugs. However, several proteins expressed in silkworm pupae cause severe allergic reactions in humans and animals. Interestingly, some baculovirus vectors have been shown to alter the host gene and its expression in insect cells, but this has not been confirmed in silkworm. Here, we analyzed the effects of infection with an empty B. mori baculovirus (BmNPV) vector on silkworm pupal protein expression. Using a proteomics approach, the allergens thiol peroxiredoxin (Jafrac1), 27‐kDa glycoprotein (p27k), arginine kinase, and paramyosin as well as 32 additional differentially expressed proteins were identified. Downregulation of the messenger RNA expression of the four known allergens was observed after BmNPV infection; subsequent changes in protein expression were confirmed by the western blot analysis using polyclonal antibodies prepared with recombinant proteins of the four allergens. Collectively, these data indicate that the four known allergens of silkworm pupae can be reduced by infection ith an empty BmNPV vector to increase the safety of silkworm pupa‐based exogenous protein expression and drug delivery of oral pharmaceuticals. In addition, the four recombinant allergen proteins may contribute to the diagnosis of allergic diseases of silkworm pupa.     

Aerobiology, biodiversity and chemistry of plant trichomes in the tropics at Bodh Gaya, India – a biopollutant and the suspected human allergen

Aerobiology, diversity and chemistry of some air borne plant trichomes were studied at Magadh University campus, Bodh Gaya, India between September 1995 and August 1996. Thirty-three morphotypes of trichomes were tapped using a Gravity Air sampler. The different types of trichomes showed pronounced seasonality in their distribution pattern. The simple non-septate trichomes of grasses were predominant throughout the year. The other commonly observed trichomes were comparable to that of Chrysanthemum, Tridax, Solanum, Amaranthus and Martynia. Trichomes. The Shannon diversity index of trichomes fluctuated widely (1.063–2.867) during different months of the year. This suggests that seasonality had definite bearing on the composition of aerobiota. Relative humidity was found to play greater role in the case of a number of morphotypes (67%), diversity index (50%) and species richness (61%) but in the case of index of dominance (62%), rainfall played a greater role at P < 0.01 level. The trichomes of a few selected plant species, the pollen grains of which are reported allergens, were chemically analysed by collecting them in bulk. They were found to be fairly rich in their total protein (9.58 to 93.60 mg/g) and carbohydrate (10.0 to 20.20 mg/g) contents.