Abstract：
The overall well-being of organisms is widely recognized to be closely intertwined with their intestinal health. The intestinal mucosal layer plays a pivotal role in ensuring the proper functioning of the intestine, a fact observed not only in humans but also in animals like pigs. Any alterations to the mucosal layer of a pig\'s intestine can potentially disrupt its functionality, thereby impacting the animal\'s health and productivity. Mass spectrometry-based proteome analysis serves as a valuable tool in investigating the intricate dynamics of the proteome within the intestinal mucosa. Such studies hold promise in uncovering causal relationships between mucosal changes and overall health outcomes in pigs. It is anticipated that insights gathered from proteome studies will inform future strategies aimed at enhancing the health and productivity of pigs. However, the research field lacks a standardized and detailed method to extract proteins from pig intestinal mucosa and prepare proteins for proteome analysis. In the present study, we evaluated three alternative S-Trap-based protocols for analyzing ileal mucosal scrapings from pigs. Samples were either freeze-dried and treated as solid samples or ground in liquid nitrogen, categorized as either solid or liquid samples. In our analysis, a total of 2840 proteins were identified across all samples. Through statistical analysis and gene ontology examinations, we investigated potential differences between the three approaches. Even though our findings revealed no significant differences among the three methods, we propose the use of the protocol wherein samples are freeze-dried and treated as solid for protein extraction. This protocol stands out as the most convenient and practical option, offering ease of use and ensuring consistent and reliable results. By establishing a standardized approach, we aim to advance research efforts in understanding pig intestinal health. SIGNIFICANCE: The development of an optimized protocol for protein extraction of intestinal mucosal scrapings in pigs addresses a gap in the field and enhances future research on pig intestinal health. By use of the protocol and mass spectrometry-based proteome analysis, valuable insights for improving the health and productivity of pigs can be presented. Studying the complex dynamics of the proteome within the intestinal mucosa, potentially identifying links between mucosal changes and health outcomes, provides us with information about the critical connection between intestinal health and the overall well-being and productivity of pigs. By creating a standardized approach, consistent, reliable, and reproducible results can be obtained for this type of research.
摘要：
生物体的整体健康状况被广泛认为与它们的肠道健康密切相关。肠粘膜层在确保肠道正常运作中起着关键作用，这一事实不仅在人类身上观察到，也在像猪这样的动物身上观察到。猪肠粘膜层的任何改变都可能破坏其功能，从而影响动物的健康和生产力。基于质谱的蛋白质组分析是研究肠粘膜内蛋白质组复杂动力学的有价值的工具。这些研究有望揭示猪粘膜变化与整体健康结果之间的因果关系。预计从蛋白质组研究中获得的见解将为旨在提高猪的健康和生产力的未来策略提供信息。然而,该研究领域缺乏从猪肠粘膜中提取蛋白质和制备蛋白质用于蛋白质组分析的标准化和详细方法。在本研究中,我们评估了三种基于S-Trap的替代方案,用于分析猪的回肠粘膜刮片.将样品冷冻干燥并作为固体样品处理或在液氮中研磨，分为固体或液体样品。在我们的分析中,在所有样本中总共鉴定出2840种蛋白质.通过统计分析和基因本体检验,我们调查了三种方法之间的潜在差异.尽管我们的发现表明这三种方法之间没有显着差异，我们建议使用该方案，其中样品被冷冻干燥并作为固体进行蛋白质提取。该协议是最方便和实用的选择，提供易用性和确保一致和可靠的结果。通过建立标准化的方法，我们的目标是推进了解猪肠道健康的研究工作。意义：猪肠道粘膜刮片蛋白质提取的优化方案的开发解决了该领域的空白，并增强了未来对猪肠道健康的研究。通过使用协议和基于质谱的蛋白质组分析，可以提出改善猪的健康和生产力的有价值的见解。研究肠粘膜内蛋白质组的复杂动态，潜在识别粘膜变化和健康结果之间的联系，为我们提供有关肠道健康与猪的整体健康和生产力之间的关键联系的信息。通过创建标准化方法，一致,可靠,和可重复的结果可以获得这种类型的研究。
