血管内皮生长因子及其受体2在卵巢上皮癌中的表达

目的比较血管内皮生长因子（VEGF）及其受体VEGFR-2在卵巢癌上皮组织中的表达。方法研究32例卵巢上皮癌和15例正常对照卵巢组织,采用冷冻切片免疫荧光染色法检测VEGF和VEGFR-2的表达,并通过蛋白免疫印迹Western blot法进一步作半定量分析。结果在正常卵巢上皮细胞中,VEGF和VEGFR-2表达阳性率分别为4/15和5/15;而在卵巢癌细胞中,VEGF和VEGFR-2表达阳性率分别为15/32和29/32,二者对应阳性率比较具有统计学意义（P〈0.01）。在正常卵巢组织中,VEGF和VEGFR-2的表达均较少,而在卵巢癌组织中,VEGF和VEGFR-2的表达均较正常卵巢组织中明显增加。结论卵巢癌中存在显著增强的VEGF/VEGFR-2自分泌作用,这种自分泌在卵巢癌的发病中可能发挥了重要的作用。     

Characterization of fibromodulin isolated from bovine periodontal ligament

Although several proteoglycans (PGs) have been reported in bovine periodontal ligament (PDL), the composition of PGs in PDL has been poorly characterized. In the present study, we isolated and characterized keratan sulfate-substituted PG (fibromodulin) in bovine PDL. Fibromodulin was purified from 4 m guanidine hydrochloride (GdmCl) extracts of bovine PDL tissues using DEAE Sephacel ion-exchange chromatography and preparative electrophoresis. Fibromodulin appeared as a single polydisperse band with an apparent molecular weight (MW) of 80, 000 (80 kDa) on SDS-PAGE. Digestion of fibromodulin with keratanase or neuraminidase reduced the apparent molecular size, and N-glycanase treatment produced core protein bands of around 40 kDa. Fibromodulin reacted with keratan sulfate monoclonal antibody (5D4) and fibromodulin polyclonal antibodies (α-FM). The keratanase-digested fibromodulin reacted with α-FM, but not with 5D4. These data suggest that fibromodulin is one of the small PGs in the PDL-matrix and may fulfill construction and maintenance functions in this tissue.     

Linear IgA disease: the IgA and IgG response to the epidermal antigens demonstrates that intermolecular epitope spreading is associated with IgA rather than IgG antibodies, and is more common in adults

BACKGROUND: Linear IgA disease (LAD; adult and childhood) is a dapsone-responsive, acquired immunobullous disorder mediated by IgA antibodies directed at target antigens within the epithelial basement membrane. These antigens have not been completely characterized. OBJECTIVES: To identify the target antigens in LAD, and to correlate these with the antibody isotype. METHODS: We used 101 LAD sera without IgG antibodies detected by indirect immunofluorescence. The sera were analysed by immunoblotting for IgA (65 adults and 36 children) and IgG (61 adults and 34 children) autoantibodies, on salt-split, urea-extracted epidermal tissue extracts. RESULTS: Antigens were targeted in LAD by IgA antibodies (54 adults and 23 children), IgG antibodies (34 adults and 19 children), and both isotypes (30 adults and 16 children). Three major antigens were recognized by IgA antibodies: LAD285 (22 adults and three children), BP230 (30 adults and eight children) and BP180 (collagen XVII), including the 97-kDa ectodomain (52 adults and 20 children). Seven 'minor' antigens were occasionally detected (18 adults and 13 children). IgA antibodies bound multiple antigens (33 adults and nine children) more frequently than single antigens (21 adults and 14 children), but the binding to multiple antigens was more restricted in children than in adults. IgG antibodies mainly bound a single antigen (29 adults and 16 children), predominantly BP180. CONCLUSIONS: There was variation in the autoantibody response within the disease and the patient, with regard to target molecules and autoantibody class. The finding that IgG as well as IgA autoantibodies predominantly target BP180 supports a pivotal role for collagen XVII in adult and childhood LAD. The IgG response was very restricted compared with IgA autoantibodies (P < 0.01). Autoantibodies from children had a more restricted antigen repertoire than from adults (P < 0.05). Epitope spreading is common in LAD and is affected by the class of autoantibody and age of the patient.     

免疫印迹法、酶联免疫法及放射免疫法对胰岛自身抗体检测的比较

目的：评价免疫印迹法检测胰岛自身抗体（GAD-A、ICA、IAA）与酶联免疫法测ICA、GADA放射免疫法测IAA结果的一致性。方法：采用免疫印迹法测定81例糖尿病患者胰岛自身抗体,将结果与酶联免疫法测定的GAD-A、ICA,放射免疫法测定IAA结果进行比较。结果：免疫印迹法阳性检出率为：GAIN-A51．8％,ICA18．5％,IAA27．1％;酶联免疫法（GAD-A、ICA）、放射免疫法（IAA）阳性检出率：GAD-A32．1％,ICA34．5％,IAA30．8％;上述两组结果进行比较,两组相比ICA和GAD-A有统计学差异（P〈0．05）,IAA无统计学差异。两组结果一致率比较：GAD-A50．6％,ICA64．2％,IAA69．1％。结论：与临床常用酶联免疫法检测GAD-A、ICA,放射免疫法检测IAA比较,免疫印迹法和酶联免疫法在ICA及GAD-A阳性检出率上的差异有显著性,和放射免疫法在IAA阳性检出率上差异无显著性。     

基因重组HCV抗原斑点酶免疫法检测血清中抗-HCV

目的　建立检测抗-HCV的斑点酶免疫法。方法　将基因重组HCV核心、NS     

Pemphigus Vulgaris Autoantibody Response is Linked to HLA-DQB10503 in Pakistani Patients

ABSTRACT: Pemphigus vulgaris (PV) is an autoimmune disease of the skin and mucous membranes characterized by an autoantibody response against an epidermal cadherin. We performed high resolution HLA class II typing in 19 patients with PV from Rawalpindi, Pakistan and 19 non-Jewish European PV patients from Boston by sequence-specific oligonucleotide probe hybridization. The results were compared with two separate ethnically matched control populations. We found that PV patients from Pakistan had significantly increased frequencies of DRB1*1404 ( p = 0.01), DQA1*0101 ( p = 0.02), and DQB1*0503 ( p = 0.01). Among the patients of non-Jewish European ancestry, DRB1*1401 ( p < 10⁻⁶), DQA1*0101 ( p < 10⁻⁵) and DQB1*0503 ( p < 10⁻⁶), were increased in PV patients. Formal linkage analysis between the major histocompatibility complex and the PV antibody was performed in 67 relatives of the 19 Pakistani patients. The results showed strong evidence for linkage of HLA-DRB1*1404, DQA1*0101, DQB1*0503, with the presence of PV antibody in relatives’ families with a significant logarithm of the odds score of 6.06. Based on the three dimensional structure of class II molecules, we propose that HLA-DQA1*0101 and DQB1*0503, encode a negatively charged P9 peptide binding pocket of the DQ molecule and are significantly associated with susceptibility to PV in non-Jewish populations.     

Diagnosis of cypress pollen allergy: in vivo and in vitro standardization of a Juniperus ashei pollen extract

BACKGROUND: Cypress pollen allergy is a major cause of rhinoconjunctivitis and asthma in the Mediterranean area. The nonstandardized cypress allergen extracts currently available for the diagnosis of cypress allergy have a low level of activity. The search for an active material has led to the selection of Juniperus ashei (Ja) pollen because of its very high cross-reactivity with cypress extracts and its superior allergenic activity. The aim of this study was to characterize in vitro and calibrate in vivo an in-house reference extract (IHRS) of J. ashei pollen and determine the specificity and sensitivity of a standardized Ja extract for the prick test diagnosis of cypress allergy. METHODS: Juniperus ashei pollen extract was analysed by 2-D electrophoresis. The IHRS Ja extract was calibrated by skin prick testing in 28 cypress-allergic patients. The sensitivity and specificity of cypress allergy diagnosis using a standardized Ja extract was studied by skin prick test in 42 cypress-allergic patients and 53 nonallergic patients. Jun a 1 content of the IHRS was determined by a monoclonal antibody-based electrophoretic technique. RESULTS: The Jun a 1 content of the 100 IR/ml Ja IHRS extract was 180 microg/ml. For in vivo diagnosis of cypress allergy, Ja pollen extract demonstrated a sensitivity of 95%, a specificity of 100%, a negative predictive value of 96%, and a positive predictive value of 100%. CONCLUSION: Standardized Ja pollen extract is therefore a very appropriate tool for the in vivo diagnosis of cypress pollen allergy and good candidate for specific immunotherapy.     

Lettuce anaphylaxis: identification of a lipid transfer protein as the major allergen

BACKGROUND: Allergy to plant-derived foods is associated with birch pollinosis in central and northern Europe. Symptoms elicited are usually limited to the oropharyngeal system. By contrast, in the Mediterranean area, allergy to the same foods manifests more frequently with systemic reactions caused by nonspecific lipid transfer proteins (nsLTP), independently of an associated pollinosis. OBJECTIVE: We sought to investigate the pattern of immunoglobulin E (IgE) binding protein bands implicated in lettuce allergy, in particular the presence of an nsLTP. METHODS: Consecutive lettuce allergic patients were selected. Determination of serum-specific IgE, immunoblot, and inhibition experiments were performed in order to study the pattern of IgE binding proteins and the potential cross-reactivity to pollens. Inhibition studies with recombinant allergens were conducted to identify the lettuce allergens. The major allergen was subjected to N-terminal amino acid sequencing. RESULTS: Fourteen patients were diagnosed as being allergic to lettuce. All were sensitized to Platanus pollen. Ten of them showed specific IgE to a lettuce protein of 9-kDa. The IgE binding to this protein was completely inhibited by the cherry-LTP and peach extract. The N-terminal sequence of the 9-kDa protein showed a high degree of amino acid sequence identity to other nsLTPs. A clear partial cross-reactivity was observed between lettuce-LTP and Platanus-pollen extract. CONCLUSIONS: An LTP has been demonstrated to be a major allergen in patients suffering from lettuce allergy.     

Bee moth (Galleria mellonella) allergic reactions are caused by several thermolabile antigens

BACKGROUND: Exposure and contact with bee moth (Galleria mellonella) larvae (Gm) can cause an allergic reaction both in anglers and breeders. We described the case of an amateur fisherman who experienced an allergic reaction using Gm but not using heat-treated Gm (h-Gm) (mummies). The aim of this study was to demonstrate by immunoblotting and radioallergosorbent test (RAST)-inhibition experiments the loss of allergenic epitopes in h-Gm extracts. METHODS: Galleria mellonella larvae and h-Gm were homogenized and extracted at 10% (w/v) in 0.5 M phosphate-buffered saline, pH 7.4 containing 0.5% NaN(3) for 16 h at 4 degrees C. Gm and h-Gm extracts were electrophoresed in a 10% polyacrylamide precast Nupage Bis-Tris gel at 180 mA for 1 h and the resolved proteins stained with 0.1% Coomassie brilliant blue and the molecular weight calculated. For the immunoblotting detection of allergenic components the resolved extracts were transferred onto a nitrocellulose membrane and incubated with the patient's serum. Bound specific-IgE was detected by peroxidase-conjugated anti-human IgE. RAST inhibition experiments were performed according to the Ceska method. RESULTS: The protein profile of Gm and h-Gm extracts resulted markedly different in number, intensity and the position of bands, indicating that heat-treatment modifies the chemical-physical characteristics of the protein contents. The Gm extract showed a strong-coloured band at 73 kDa and more than 20 components ranging from 12 to 133 kDa; h-Gm showed two main band at 77 and 38 kDa and about 15 faint bands between 20 and 133 kDa apparently without any correspondence to the bands present in the Gm extract. Immunoblotting with the patient's serum demonstrated several bands of reactivity with the Gm extract ranging from 20 to 100 kDa and no recognizable bands, but only a diffuse smear with h-Gm. When used in a RAST inhibition experiment the h-Gm extract demonstrated an inability to compete with the Gm one for the binding to patient's IgE serum. CONCLUSIONS: The h-Gm seems to lose the allergenic epitopes and has two advantages for anglers: to avoid new possible sensitizations as well as allergic symptoms in sensitized people, without interfering with their skills and satisfaction in their fishing performance.     

Humoral immunity patterns based on antibody reactivity to rotavirus antigens in Brazilian children under 5 years of age

The age distribution of antibody to simian rotavirus (SA-11) was studied in serum specimens obtained from 399 children aged to 5 years and living in the city of Recife (PE), located in the north eastern region of Brazil. Sera were examined for group-specific rotavirus antibody using a blocking enzyme immunoassay (bELISA) and a hemagglutination inhibition antibody (HIA) test, and for anti-VP2, anti-VP4, anti-VP6, and anti-VP7 antibodies using an immunoblotting assay (IBA). Antibody prevalence was similar in all bELISA and HIA assays, showing a steep rise in the 6- to 17-month-old age groups. The results indicate early acquisition of antibody to rotavirus. The majority of children aged 2 to 4 years had bELISA (50% to 60%) and HIA (70% to 81%) antibodies. There was an association in prevalence data obtained by HIA and bELISA with immunoblotting (IBA), revealing four serologic profiles. Children with profiles I and II (60%) respectively had HAI and ELISA antibody or HAI antibody alone and all had immunoprotective antibodies to VP4 and/or VP7. These children were regarded as “immune,” resembling convalescent patients with a rotavirus infection. Children with profile III (4%) had no HIA antibody and only non-protective anti-VP6 and/or VP7 antibody, and were considered to be “partially immune.” Children with profile IV (36%) had no detectable antibody and were classified as “nonimmune.” These children should be considered to be susceptible to rotavirus infection, with the risk of developing clinically severe diarrhea. © 1996 Wiley-Liss, Inc.