Harish Chandra, Bethany Ahlers, Ying Wai Lam, Jagjit S. Yadav
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引用次数: 0
Abstract
Background: Occupational exposure to industrial Metalworking Fluid (MWF) colonized by Mycobacterium immunogenum (MI) has been associated with immune lung disease hypersensitivity pneumonitis (HP) in machinists. This warrants regular fluid monitoring for early detection of mycobacterial proteins, especially those with antigenic potential. Objective: To detect and identify dominant MI proteins and antigens directly from the field-drawn in-use MWF using an integrated immunoproteomic and immunoinformatic approach. Methods: An MI-positive MWF selected by DNA-based screening of several field-drawn MWF samples were cultured to isolate the colonizing strain and profiled for dominant circulating cell- free (ccf) MI proteins, including antigens using an integrated immunoproteomic (1D- and 2Dgel fractionation of seroreactivity proteins combined with shotgun proteomic analysis using LC-MS/ MS) and immunoinformatic strategy. Results: A new MI strain (MJY-27) was identified. The gel fractionated MI protein bands (1Dgel) or spots (2D-gel) seroreactive with anti-MI sera probes (Rabbit and Patient sera) yielded 86 MI proteins, 29 of which showed peptide abundance. T-cell epitope analysis revealed high (90-100%) binding frequency for HLA-I& II alleles for 13 of the 29 proteins. Their antigenicity analysis revealed the presence of 6 to 37 antigenic determinants. Interestingly, one of the identified candidates corresponded to an experimentally validated strong B- and T-cell antigen (AgD) from our laboratory culture-based studies. result: A new MI strain (MJY-27) was identified. The gel fractionated MI protein bands (1D-gel) or spots (2D-gel) seroreactive with anti-MI sera probes (Rabbit and Patient sera) yielded 86 MI proteins, 29 of which showed peptide abundance. T-cell epitope analysis revealed high (90-100%) binding frequency for HLA-I& II alleles for 13 of the 29 proteins. Their antigenicity analysis revealed presence of 6 to 37 antigenic determinants. Interestingly, one of the identified candidates corresponded to an experimentally validated strong B- and T- cell antigen (AgD) from our laboratory culture-based studies. Conclusion: This first report on dominant proteins, including putative antigens of M. immunogenum prevalent in field in-use MWF, is a significant step towards the overall goal of developing fluid monitoring for exposure and disease risk assessment for HP development in machining environments. conclusion: This first report on dominant proteins including putative antigens of M. immunogenum prevalent in field in-use MWF is a significant step towards the overall goal of development of fluid monitoring for exposure and disease risk assessment for HP development in machining environments.
背景：机械师在工作中接触到由免疫分枝杆菌（MI）定植的工业金属加工液（MWF）与免疫性肺部疾病超敏性肺炎（HP）有关。因此，有必要对金属加工液进行定期监测，以便及早发现分枝杆菌蛋白，尤其是具有抗原潜力的分枝杆菌蛋白。目标：检测并确定主要的分枝杆菌蛋白采用免疫蛋白组学和免疫形式学综合方法，直接从现场抽取的使用中的 MWF 中检测和鉴定主要的分枝杆菌蛋白和抗原。研究方法：通过对多个野外提取的 MWF 样品进行 DNA 筛选，筛选出 MI 阳性的 MWF，对其进行培养以分离出定植菌株，并使用综合免疫蛋白组学（血清反应蛋白的一维和二维凝胶分馏，结合使用 LC-MS/ MS 的霰弹枪蛋白质组分析）和免疫形式化策略分析主要的循环游离细胞 (ccf) MI 蛋白，包括抗原。结果：确定了一种新的 MI 株系（MJY-27）。用抗 MI 血清探针（兔血清和患者血清）进行血清反应的凝胶分馏 MI 蛋白带（1D-凝胶）或点（2D-凝胶）得到了 86 个 MI 蛋白，其中 29 个显示了肽丰度。T细胞表位分析显示，29种蛋白质中有13种与HLA-I& II等位基因的结合频率很高（90%-100%）。其抗原性分析表明存在 6 至 37 个抗原决定因子。有趣的是，其中一个候选抗原与我们实验室培养研究中实验验证的强 B 细胞和 T 细胞抗原（AgD）相对应：确定了一种新的 MI 菌株（MJY-27）。用抗 MI 血清探针（兔血清和患者血清）对凝胶分馏的 MI 蛋白带（1D-凝胶）或点（2D-凝胶）进行血清反应，得到了 86 个 MI 蛋白，其中 29 个显示了肽丰度。T细胞表位分析显示，29种蛋白质中有13种与HLA-I&II等位基因的结合频率很高（90%-100%）。其抗原性分析表明存在 6 至 37 个抗原决定因子。有趣的是，其中一种候选蛋白与我们实验室培养研究中实验验证的强 B 细胞和 T 细胞抗原（AgD）相对应。结论：这份关于主要蛋白质（包括现场使用中的 MWF 中普遍存在的免疫原疟原虫的假定抗原）的首次报告，是朝着为机械加工环境中的 HP 开发流体接触监测和疾病风险评估的总体目标迈出的重要一步：该报告首次介绍了野外使用的 MWF 中普遍存在的主要蛋白质（包括免疫原疟原虫的假定抗原），这是为机械加工环境中的 HP 开发开发流体监测暴露和疾病风险评估总体目标迈出的重要一步。
期刊介绍：
Accounts of Chemical Research presents short, concise and critical articles offering easy-to-read overviews of basic research and applications in all areas of chemistry and biochemistry. These short reviews focus on research from the author’s own laboratory and are designed to teach the reader about a research project. In addition, Accounts of Chemical Research publishes commentaries that give an informed opinion on a current research problem. Special Issues online are devoted to a single topic of unusual activity and significance.
Accounts of Chemical Research replaces the traditional article abstract with an article "Conspectus." These entries synopsize the research affording the reader a closer look at the content and significance of an article. Through this provision of a more detailed description of the article contents, the Conspectus enhances the article's discoverability by search engines and the exposure for the research.
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