Laboratory of Comparative Perinatal Development, Department of Veterinary Sciences, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Antwerp, Belgium.
Laboratory of Protein Chemistry, Proteomics and Epigenetic Signalling (PPES), Department of Biomedical Sciences, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Antwerp, Belgium.
PLoS One. 2023 May 26;18(5):e0286455. doi: 10.1371/journal.pone.0286455. eCollection 2023.
Monitoring chronic stress in pigs is not only essential in view of animal welfare but is also important for the farmer, given that stress influences the zootechnical performance of the pigs and increases their susceptibility to infectious diseases. To investigate the use of saliva as a non-invasive, objective chronic stress monitoring tool, twenty-four 4-day-old piglets were transferred to artificial brooders. At the age of 7 days, they were assigned to either the control or the stressed group and reared for three weeks. Piglets in the stressed group were exposed to overcrowding, absence of cage enrichment, and frequent mixing of animals between pens. Shotgun analysis using an isobaric labelling method (iTRAQ) for tandem mass spectrometry performed on saliva samples taken after three weeks of chronic stress identified 392 proteins, of which 20 proteins displayed significantly altered concentrations. From these 20 proteins, eight were selected for further validation using parallel reaction monitoring (PRM). For this validation, saliva samples that were taken one week after the start of the experiment and samples that were taken at the end of the experiment were analysed to verify the profile over time. We wanted to investigate whether the candidate biomarkers responded fast or rather slowly to the onset of chronic exposure to multiple stressors. Furthermore, this validation could indicate whether age influenced the baseline concentrations of these salivary proteins, both in healthy and stressed animals. This targeted PRM analysis confirmed that alpha-2-HS-glycoprotein was upregulated in the stressed group after one and three weeks, while odorant-binding protein, chitinase, long palate lung and nasal epithelium protein 5, lipocalin-1, and vomeromodulin-like protein were present in lower concentrations in the saliva of the stressed pigs, albeit only after three weeks. These results indicate that the porcine salivary proteome is altered by chronic exposure to multiple stressors. The affected proteins could be used as salivary biomarkers to identify welfare problems at the farm and facilitate research to optimise rearing conditions.
监测猪的慢性应激不仅对动物福利至关重要，而且对农民也很重要，因为应激会影响猪的生产性能并增加它们对传染病的易感性。为了研究唾液作为一种非侵入性、客观的慢性应激监测工具的用途，将 24 头 4 天大的仔猪转移到人工保育器中。在 7 天大时，它们被分配到对照组或应激组，并饲养三周。应激组的仔猪暴露于过度拥挤、无笼内丰容和频繁混合动物。使用同位标记法（iTRAQ）进行的唾液样本的串联质谱分析，对三周慢性应激后的样本进行分析，共鉴定出 392 种蛋白质，其中 20 种蛋白质的浓度明显改变。从这 20 种蛋白质中，选择了 8 种蛋白质进行平行反应监测（PRM）的进一步验证。对于这种验证，分析了实验开始一周后和实验结束时采集的唾液样本，以验证随时间的变化情况。我们想研究候选生物标志物是否对慢性暴露于多种应激源的开始快速或缓慢反应。此外，这种验证可以表明年龄是否影响这些唾液蛋白质的基础浓度，无论是在健康动物还是应激动物中。这种靶向 PRM 分析证实，应激组在一周和三周后α-2-HS-糖蛋白上调，而气味结合蛋白、几丁质酶、长腭肺和鼻上皮蛋白 5、脂钙蛋白-1 和鼻后神经分泌素样蛋白的浓度较低，但仅在三周后。这些结果表明，猪唾液蛋白质组受慢性暴露于多种应激源的影响。受影响的蛋白质可以作为唾液生物标志物，以识别农场的福利问题，并促进优化饲养条件的研究。
