Characterization of innate immune signalling receptors in virus‐induced acute asthma

Background The role of toll‐like receptors (TLRs) and innate immune activation in clinical asthma exacerbations and their relationship to virus infection are unclear. Objective This study aimed to characterize TLR expression and innate immune activity during virus infection in acute asthma. Methods Subjects with acute asthma, stable asthma and healthy controls were recruited and underwent spirometry and sputum induction with isotonic saline. Selected sputum was dispersed with dithiothreitol and total and differential leucocyte counts were performed. Selected sputum was also used for quantitative real‐time PCR for TLR2, TLR3, TLR4, IL‐10 and IP‐10mRNA expression. Sputum supernatant was used for the measurement of innate immune markers, including IL‐8, matrix metalloproteinase‐9 and neutrophil elastase activity. Viruses were detected using real‐time and gel‐based PCR. Results Sputum TLR2 mRNA expression was up‐regulated in both acute and stable asthma compared with healthy controls and decreased 4–6 weeks after acute exacerbation. Sputum TLR2 mRNA expression was elevated in viral, compared with non‐viral, acute asthma. Sputum TLR3 mRNA expression was similar in controls, stable and acute asthma. However, in acute asthma, subjects with virus‐induced acute asthma had significantly higher sputum TLR3 mRNA expression. Induced sputum gene expression for IP‐10 and IL‐10 were increased in viral, compared with non‐viral, acute asthma. In virus‐induced acute asthma, levels of IP‐10 and IL‐10 mRNA expression were correlated with the mRNA expression of TLR2 and TLR3. Conclusions and Clinical Relevance Virus‐induced acute asthma leads to specific induction of TLR2, TLR3, IP‐10 and IL‐10, suggesting that signalling via TLRs may play an important role in mediating airway inflammation, via both innate and adaptive pathways, in virus‐induced exacerbations. These mediators may provide potential treatment targets for virus‐induced asthma. They may also be useful in diagnosing the nature of acute asthma exacerbations and monitoring treatment responses, which would be useful in the clinical management of asthma exacerbations. Cite this as: L. G. Wood, J. L. Simpson, P. A. B. Wark, H. Powell and P. G. Gibson, Clinical & Experimental Allergy, 2011 (41) 640–648.     

Allergic airway inflammation is exacerbated during acute influenza infection and correlates with increased allergen presentation and recruitment of allergen-specific T-helper type 2 cells

BACKGROUND: Respiratory viral infections are a leading cause of the hospitalization of asthmatics, however, the cellular immunological interactions which underlie these two diseases remain elusive. OBJECTIVE: We sought to characterize the effect influenza viral infection has on allergic airway inflammation and to identify the cellular pathways involved. METHODS: We have used an ovalbumin (OVA) model of allergic airway inflammation, which involves sensitization of animals with OVA adsorbed in alum adjuvant followed by an intranasal challenge with OVA in phosphate-buffered saline. To study T cell recruitment into the lung, we adoptively transferred in vitro activated T cell receptor-transgenic T cells, which were subsequently identified by fluorescence-activated cell sorting (FACS) analysis. In addition, to study in vivo dendritic cell (DC) migration, we administered fluorescently labelled dextran and identified DCs that had phagocytosed it by FACS analysis. RESULTS: We found that different stages of influenza infection had contrasting effects upon the outcome of OVA-induced allergic airway inflammation. The allergic response against OVA was exacerbated during the acute stage of influenza infection; however, mice were protected against the development of airway eosinophilia at late time-points following infection. We investigated the mechanisms responsible for the virus-induced exacerbation and found that the response was partially independent of IL-4 and that there was increased delivery of inhaled allergens to the draining lymph node during the acute stage of the infection. In addition, virus-induced inflammation in the lung and draining lymph node resulted in the non-specific recruitment of circulating allergen-specific effector/memory cells. CONCLUSION: In addition to virus-mediated damage to the lung and airways, influenza viral infection can also enhance unrelated local allergic responses.     

The incidence of human herpes virus‐8 expression in lymph node biopsies from human immunodeficiency virus‐positive patients

Beukes C A & Thiart J
(2012) Histopathology  61, 942–944 The incidence of human herpes virus‐8 expression in lymph node biopsies from human immunodeficiency virus‐positive patients Aims: Human immunodeficiency virus (HIV)‐related lymphadenopathy is characterized by a wide spectrum of histological changes. Three patterns have been described which correspond to clinical stages of HIV/acquired immune deficiency syndrome (AIDS). Castleman disease is a heterogeneous group of disorders. A recently described variant, multicentric Castleman disease (MCD), of which some cases are associated with human herpes virus‐8 (HHV‐8), has been reported in both HIV‐seropositive and ‐negative patients. Considerable morphological overlap occurs between one of the patterns of HIV lymphadenopathy and this variant. Methods and results: This retrospective histopathological study on 95 cases of HIV‐reactive lymphadenopathy assessed the incidence of the different patterns and HHV‐8 on immunohistochemistry (IHC). Of the 95 cases, 78 (82.1%) were HHV‐8‐negative, of which 46 (59.0%) were classified as pattern A, 20 (25.6%) as pattern B and 12 (15.4%) as pattern C. Nine (31.0%) of 29 cases with pattern B and 8 (40.0%) of 20 cases with pattern C were HHV‐8 positive. In total 15 cases of MCD were diagnosed in this series. Conclusion: This study draws attention to the overlap between HIV lymphadenopathy and MCD. We recommend that cases of HHV‐8‐associated MCD should be investigated for HIV infection.     

牙胚细胞体外培养过程中成牙基因的表达

背景：多项实验表明牙胚组织在不同的时间点有不同的基因发挥作用,共同促进牙胚发育。 目的：观察牙本质基质蛋白1、成釉蛋白、Ⅰ型胶原蛋白和同源异型盒基因1在大鼠牙胚细胞体外培养的不同时间的表达。 方法：对体外培养后第1,3,6天的牙胚细胞提取RNA,反转录后采用实时定量PCR的方法检测牙本质基质蛋白1、成釉蛋白、Ⅰ型胶原蛋白和同源异型盒基因1mRNA相对表达水平的变化。 结果与结论：牙胚细胞中牙本质基质蛋白1、成釉蛋白和Ⅰ型胶原蛋白mRNA的表达随培养时间的延长而增加,在培养3 d时达到峰值(P < 0.05),而同源异型盒基因1 mRNA的表达随培养时间的延长而下降(P < 0.05)。 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程全文链接：     

Over‐expression of the LTC4 synthase gene in mice reproduces human aspirin‐induced asthma

Background The pathogenesis of aspirin‐induced asthma (AIA) is presumed to involve the aspirin/non‐steroidal anti‐inflammatory drug (NSAID)‐induced abnormal metabolism of arachidonic acid, resulting in an increase in 5‐lipoxygenase (5‐LO) metabolites, particularly leukotriene C₄ (LTC₄). However, the role of LTC₄ in the development of AIA has yet to be conclusively demonstrated. Objective The aim of this study was to evaluate the contribution of the lipid product LTC₄ secreted by the 5‐LO pathway to the pathogenesis of AIA. Methods To evaluate antigen‐induced airway inflammation, the concentrations of T‐helper type 2 cytokine in bronchoalveolar lavage fluid (BALF) obtained from LTC₄ synthase‐transgenic (Tg) and wild‐type (WT) mice after challenge with ovalbumin were measured. Subsequently, the ex vivo and in vivo effects of the NSAID sulpyrine were investigated in these Tg and WT mice by measuring the secretion of LTC₄ from sulpyrine‐treated BAL cells and the levels of LTC₄ in BALF following challenge with sulpyrine. Finally, the sulpyrine‐induced airway response by the administration of pranlukast, an antagonist of the cysteinyl (cs)‐LT1 receptor, was analysed. Results The concentrations of IL‐4, ‐5, and ‐13 in BALF from Tg mice were significantly higher than those in WT mice. In addition, sulpyrine augmented the secretion of LTC₄ in BALF and by BAL cells in Tg mice, but not in WT mice. Additionally, the increased airway resistance induced by sulpyrine could be reduced by treatment with pranlukast. Furthermore, the secretion of LTC₄ from mast cells, eosinophils, and macrophages was increased in the allergen‐stimulated LTC₄ synthase gene Tg mice, even in the absence of sulpyrine, as well as in BAL cells after sulpyrine. Conclusion and clinical relevance The over‐expression of the LTC₄ synthase in a mouse asthma model also replicates the key features of AIA. And our study supports that cys‐LTs play a major role in the pathogenesis of AIA in patients with chronic asthma. Cite this as: H. Hirata, M. Arima, Y. Fukushima, K. Honda, K. Sugiyama, T. Tokuhisa and T. Fukuda, Clinical & Experimental Allergy, 2011 (41) 1133–1142.     

丙型肝炎病毒核心蛋白基因重组质粒的构建及其在真核细胞中的表达

目的：构建包含丙型肝炎病毒（ＨＣＶ）核心蛋白（Ｃ）基因片段的重组真核表达载体，并在肝细胞癌细胞株７７２１细胞中表达。方法：将从ｐＢＲＴＭＨＣＶ１－３０１１质粒切下的ＨＣＶＣ基因片段插入ｐｃＤＮＡ３质粒的ＣＭＶ启动子下游，构建真核表达质粒ｐｃＤＮＡＨＣＶ－Ｃ，然后，采用脂质体转染技术，转染７７２１细胞进行瞬时表达，转染细胞裂解煮沸后，通过ＳＤＳ－ＰＡＧＥ及Ｗｅｓｔｅｒｎｂｌｏｔ检测表达的核心抗原。结果：用限制性内切酶酶切后，片段大小与计算值相符。Ｗｅｓｔｅｒｎｂｌｏｔ证实，表达抗原的Ｍｒ约为２２０００。结论：ＨＣＶＣ基因能够插入ｐｃＤＮＡ３真核表达载体，并使其在真核细胞中表达，为进一步ＨＣＶ基因疫苗的研制和探讨抗ＨＣＶ感染打下了基础。