胰腺导管腺癌和慢性胰腺炎中18q21杂合性缺失及相关性分析

目的 探讨18q21在人胰腺导管腺癌和慢性胰腺炎中杂合性缺失(LOH)的情况及其相关因素.方法 选择18q21上的位点RP11-729G3和RP11-850A17作为目的 片段,选择接近18号染色体着丝粒的位点RP11-621L6作为参照位点,利用细菌人工染色体(BAC)提取、纯化相应位点的DNA,用切口平移法分别标记生物素和地高辛后制成双色探针,应用荧光原位杂交技术(FISH)检测30例胰腺导管腺癌和10例慢性胰腺炎石蜡包埋组织切片中18q21 LOH情况,并收集、整理相应临床病理资料进行相关性分析.结果 30例胰腺导管腺癌中在RP11-729G3位点有25例(83.3%)有LOH,在RP11-850A17位点26例(86.6%)有LOH,其中25例在RP11-729G3和fuP11-850A17两个位点均有LOH,1例仅在RP11-850A17位点有LOH.10例慢性胰腺炎中均未发现18q21 LOH.经统计学分析发现,18q21 LOH在慢性胰腺炎和胰腺导管腺癌中差异有统计学意义(P<0.01),且RP11-729G3和RP11-850A17两个位点LOH有高度相关性(Phj系数=0.877),但和临床病理各因素间未发现明显相关.结论 18q21 LOH在胰腺导管腺癌中属于高频事件,并且位点RP11-729G3和RP11-850A17的LOH有高度相关性,可能在胰腺导管腺癌发生发展中起重要作用.在临床诊断中18q21LOH也能作为较特异的标记辅助诊断.     

Cloning of a Novel Babesia equi Gene Encoding a 158-Kilodalton Protein Useful for Serological Diagnosis

In this study, we characterized a Babesia equi Be158 gene obtained by immunoscreening a B. equi cDNA expression phage library with B. equi-infected horse serum. The Be158 gene consists of an open reading frame of 3,510 nucleotides. The recombinant Be158 gene product was produced in Escherichia coli and used for the immunization of mice. In Western blot analysis, mouse immune serum against the Be158 gene product recognized 75- and 158-kDa proteins from the lysate of B. equi-infected erythrocytes. In an indirect fluorescent-antibody test with the mouse immune serum, the Be158 antigen appeared in the cytoplasm of Maltese cross-forming parasites (which consist of four merozoites) and was located mainly in the extraerythrocytic merozoite body. When the recombinant Be158 gene product was used in an enzyme-linked immunosorbent assay as a serological antigen, it was found to react to B. equi-infected horse sera, indicating that the Be158 gene product is useful as a serologically diagnostic antigen for B. equi infection.     

Expression of Babesia equi merozoite antigen 1 in insect cells by recombinant baculovirus and evaluation of its diagnostic potential in an enzyme-linked immunosorbent assay

The gene encoding the entire Babesia equi merozoite antigen 1 (EMA-1) was inserted into a baculovirus transfer vector, and a recombinant virus expressing EMA-1 was isolated. The expressed EMA-1 was transported to the surface of infected insect cells, as judged by an indirect fluorescent-antibody test (IFAT). The expressed EMA-1 was also secreted into the supernatant of a cell culture infected with recombinant baculovirus. Both intracellular and extracellular EMA-1 reacted with a specific antibody in Western blots. The expressed EMA-1 had an apparent molecular mass of 34 kDa that was identical to that of native EMA-1. The secreted EMA-1 was used as an antigen in an enzyme-linked immunosorbent assay (ELISA). The ELISA differentiated B. equi-infected horse sera from Babesia caballi-infected horse sera or normal horse sera. The ELISA was more sensitive than the complement fixation test and IFAT. These results demonstrated that the recombinant EMA-1 expressed in insect cells might be a useful diagnostic reagent for detection of antibodies to B. equi.     

Wnt2信号通路重要成员在胶质瘤中的表达及其意义

近来发现Wnt信号通路对个体发育和肿瘤生成具有重要作用[1-2].为进一步探讨Wnt通路在胶质瘤中是否存在表达异常及其在胶质瘤生成中的作用,我们采用RT-PcR、组织芯片免疫组织化学染色以及蛋白印迹法,对人脑胶质瘤Wnt2通路相关因子的表达变化及其意义进行了分析.     

Immunization with recombinant surface antigen P50 of Babesia gibsoni expressed in insect cells induced parasite growth inhibition in dogs

This is a report of a vaccine trial directed against Babesia gibsoni infection in dogs with the use of the recombinant antigen P50. Dogs immunized with P50 showed partial protection manifested as a significantly low level of parasitemia. The results indicated that P50 is a primary vaccine candidate molecule against canine B. gibsoni infection.     

Effects of hypoxia and tumor necrosis factor-alpha on production of plasminogen activator inhibitor-1 in human proximal renal tubular cells

Chronic hypoxia has been newly proposed as a common mechanism of tubulointerstitial fibrosis in the progression of various chronic inflammatory renal diseases, where PAI-1 plays an important role in the accumulation of extracellular matrix (ECM) through inhibition of plasmin-dependent ECM degradation. In the present study, we investigated the presence of PAI-1 in renal tubular cells by immunostaining renal biopsy samples. We also closely examined the effects of hypoxia and TNF-alpha on PAI-1 expression in cultured human proximal renal tubular cells (HRCs). Confluent cells growth-arrested in DMEM for 24h were exposed to hypoxia (1% O2) and/or TNF-alpha at 10 ng/ml for 24 hours. Amounts of PAI-1 protein and mRNA after stimulation were measured by ELISA and TaqMan quantitative PCR, respectively and compared to those in cells incubated under control conditions (18% O2 without TNF-alpha). HIF-1alpha was demonstrated by immunoblot analysis. In crescentic glomerulonephritis, clusters of proximal tubules were specifically stained for PAI-1. Treatment of 24 hours with hypoxia, TNF-alpha and their combination induced a 2.7-fold, a 1.8-fold, and a 4.6-fold increase in PAI-1 protein secretion, and produced a 3.6-fold, a 3.3-fold, and a 12.1-fold increase at the PAI-1 mRNA level, respectively. Immunoblot analysis revealed that hypoxia-inducible factor-1alpha (HIF-1alpha) was markedly accumulated in the nuclear fraction after 16-hours exposure of HPTECs to hypoxia but not to TNF-alpha. In conclusion, hypoxia induces PAI-1 expression via remarkable nuclear accumulation of HIF-1alpha in HRCs. TNF-alpha can enhance this hypoxia-induced PAI-1 expression.     

Improved enzyme-linked immunosorbent assay using C-terminal truncated recombinant antigens of Babesia bovis rhoptry-associated protein-1 for detection of specific antibodies

An enzyme-linked immunosorbent assay (ELISA) based on a recombinant rhoptry-associated protein-1 (RAP-1) of Babesia bovis has been previously developed, but it was imperfect because some cross-reactions were still present in Babesia bigemina-infected bovine sera. To improve its accuracy for the specific detection of the antibodies to B. bovis, we constructed three C-terminal truncated recombinant antigens of the RAP-1-rCT1 (amino acids [aa] 301 to 408), rCT2 (aa 388 to 490), and rCT3 (aa 466 to 565)-by using a baculovirus expression system and evaluated their diagnostic potentials using ELISA. rCT1 and rCT2 were better diagnostic antigens in their sensitivities and diagnostic efficiencies than rCT3, although none of the recombinant antigens showed any cross-reactivity to B. bigemina-infected bovine sera. These results confirmed that the N-terminal 300-aa region caused cross-reactivity of the entire RAP-1 antigen, and the C-terminal truncated recombinant antigens were shown to be useful reagents for species-specific serodiagnosis.     

Heterogeneity of feline herpesvirus type 1 strains

Summary Heterogeneity of 9 feline herpesvirus type 1 (FHV-1) strains consisting of the prototype C27 strain, one French isolate, six Japanese isolates, and the attenuated vaccine F2 strain was examined by biological, immunological, and molecular biological methods. No significant difference was observed in virus growth and antigenic properties among the strains in Crandell feline kidney cell cultures. Hemagglutination activity was also detected in all extracts of cells infected with each strain. However, in immunoblot analysis, a virus-structural immunogenic protein with an M_r of 36 kDa was lacking in 2 strains, one of which was the vaccine F2 strain, whereas the other immunogenic proteins including three kinds of major glycoproteins were detected in all strains without differences in electrophoretic mobilities. Furthermore, when restriction endonuclease analysis was performed to examine the genomic heterogeneity of strains, the cleavage patterns with the enzymeMluI showed a genomic heterogeneity between wild and vaccine strains. In contrast, only a slight variation in the sizes of some fragments was shown with most of the 7 other enzymes used. These results indicated that the lack of the 36 kDa protein and theMluI cleavage pattern could be used as markers of the vaccine F2 strain. The specific markers are important not only to control the quality of the vaccine but also to evaluate the vaccine immunity in FHV-1 infection in cats.     

The effect of neonatal BCG vaccination on atopy and asthma at age 7 to 14 years: an historical cohort study in a community with a very low prevalence of tuberculosis infection and a high prevalence of atopic disease

BACKGROUND: There are conflicting reports on the effect of BCG vaccination on the subsequent development of atopy and asthma. There are no data on the effects of neonatal BCG vaccination on cytokine responses of lymphocytes that are exposed in vitro to allergens. OBJECTIVES: We sought to test the hypothesis that neonatal BCG vaccination or, alternatively, evidence of an immunologic memory of this vaccination is associated with a reduced prevalence of allergic sensitization, asthma, eczema, and hay fever during childhood. METHODS: An historical cohort study was conducted among 7- to 14-year-old children who were born in 2 districts in Sydney, Australia, and whose mothers were born in southeast Asia. One district had routinely administered BCG vaccination to infants born to overseas-born mothers and the other had not. Eligible subjects were identified from birth registers. Consenting subjects completed questionnaires, performed spirometric and airway hyperresponsiveness testing, and had allergen skin prick testing and tuberculin skin testing. Blood was collected to measure total serum IgE levels and for in vitro lymphocyte culture in the presence of an extract of house dust mite, the dominant allergen in this region, and purified protein derivative of Mycobacterium tuberculosis (tuberculin). IL-4, IL-5, IL-10, and IFN-gamma were measured in the culture supernatant. RESULTS: The cohort included 309 BCG-vaccinated subjects and 442 non-BCG-vaccinated subjects. BCG-vaccinated subjects did not have a lower rate of allergic sensitization than nonvaccinated subjects. However, among the subgroup of subjects with a family history of rhinitis or eczema, BCG vaccination was associated with a lower prevalence of current asthma (defined as recent wheezing plus airway hyperresponsiveness; relative risk, 0.46; 95% CI, 0.22-0.95). BCG vaccination was also associated with lower levels of allergen-stimulated IL-10 production in vitro. Among the BCG-vaccinated subjects, the 44 (14.3%) who had tuberculin skin test reaction sizes of 5 mm or greater and the 31 (18.3%) who demonstrated an in vitro IFN-gamma response to purified protein derivative of M tuberculosis did not have lower rates of allergic sensitization and, overall, did not have a lower prevalence of allergic disease than tuberculin skin test or IFN-gamma nonreactors. CONCLUSION: We conclude that neonatal BCG vaccination has an effect on T-cell allergen responsiveness 7 to 14 years after vaccination and that among a subgroup of subjects with an inherited predisposition to allergic disease, this is associated with clinically relevant beneficial effects. The findings of this study encourage the view that external influences on the immune system in the neonatal period have consequences that extend into later childhood and influence the expression of asthma. Genetic factors are likely to modify the effect of those external factors.     

Association between nasal and bronchial symptoms in subjects with persistent allergic rhinitis

BACKGROUND: The association between nasal and bronchial symptoms, and the course of bronchial responsiveness and airway inflammation in house dust mite sensitive persistent rhinitis over a prolonged time period has not been thoroughly explored. OBJECTIVE: To determine if nasal symptoms were associated with bronchial symptoms in persistent rhinitic subjects, and to assess their bronchial responsiveness and airway inflammation in comparison to nonrhinitic, nonatopic controls. The additional impact of pollen sensitivity on the lower airways in rhinitic subjects was also addressed. METHODS: Rhinitics and controls answered telephone symptom questionnaires once every 2 weeks for 1 year. Every 3 months, exhaled nitric oxide (eNO) and bronchial responsiveness to histamine were measured. RESULTS: Thirty-seven rhinitics and 19 controls completed the study. High nasal symptom scores in rhinitic subjects were associated with bronchial symptoms (OR = 1.7, 95% CI 1.2-2.5). Bronchial hyper-responsiveness was present in 32.4% of rhinitic subjects on at least one clinical visit during the year. Pollen allergy caused seasonal variation in eNO (P = 0.03). CONCLUSION: In persistent rhinitic subjects, high nasal symptom scores were associated with bronchial symptoms, and many subjects experienced bronchial hyper-responsiveness during the year. Persistent rhinitic subjects were more at risk than healthy adults of bronchial symptoms and airway inflammation, which are likely risk factors for asthma.