Phillips Ida L, Danelishvili Lia, Bermudez Luiz E
Department of Biomedical Sciences, Carlson College of Veterinary Medicine, Oregon State University, Corvallis, OR 97331, USA.
Department of Microbiology, College of Sciences, Oregon State University, Corvallis, OR 97331, USA.
Proteomes. 2021 Apr 30;9(2):20. doi: 10.3390/proteomes9020020.
Johne's disease is a chronic and usually fatal enteric infection of ruminants caused by subspecies (MAP) and is responsible for hundreds of millions of dollars in losses for the agricultural industry. Natural infection typically begins with bacterial uptake and translocation through the epithelium of the small intestine, followed by ingestion by tissue macrophages and dissemination via the lymphatic or blood system throughout the body. To gain insights into the host responses and adaptation of MAP within phagocytic cells, we utilized the previously developed cell culture passage model, and mass spectrometric-based quantitative proteomic approach. Using the cell culture system, which mimics an in vivo interaction of MAP with intestinal epithelium and tissue macrophages, bacteria were passed through the bovine epithelial cells and, subsequently, used for macrophage infection (termed indirect infection), while uninfected cells and macrophage infection initiated with the culture grown bacteria (termed direct infection) served as controls. Approximately 3900 proteins were identified across all studied groups. The comparison within the subset of proteins that showed synthesis for more than two-fold in the direct infection over the uninfected control revealed an enrichment for the pro-inflammatory pathways such as the NF-κB and cytokine/chemokine signaling, positive regulation of defense response, cell activation involved in the immune response and adaptive immune system. While these responses were absent in the indirect infection, cellular pathways such as cell cycle, healing, regulation of cell adhesion, ensemble of core extracellular matrix proteins, cell surface integrins and proteins mediating the integrin signaling were remarkably high within the indirect infection. In addition to global analysis of the macrophage proteome, we further validated the proteomics data and confirmed that MAP passage through epithelial cells modulates the expression and signaling of integrins in phagocytes. In this study, we demonstrate that predominant expression of integrins in the indirectly infected macrophages allows phagocytic cells to initiate stronger binding and efficient translocation through the endothelial cells, suggesting the important role of integrins in the spread of MAP infection.
约内氏病是一种由副结核分枝杆菌亚种（MAP）引起的反刍动物慢性且通常致命的肠道感染，给农业产业造成了数亿美元的损失。自然感染通常始于细菌通过小肠上皮的摄取和转运，随后被组织巨噬细胞摄取，并通过淋巴或血液系统在全身传播。为了深入了解吞噬细胞内宿主对MAP的反应和适应性，我们利用了先前开发的细胞培养传代模型以及基于质谱的定量蛋白质组学方法。使用模拟MAP与肠上皮和组织巨噬细胞体内相互作用的细胞培养系统，细菌先通过牛上皮细胞，随后用于感染巨噬细胞（称为间接感染），而未感染的细胞以及用培养生长的细菌起始的巨噬细胞感染（称为直接感染）作为对照。在所有研究组中鉴定出约3900种蛋白质。在直接感染组中合成量比未感染对照高出两倍以上的蛋白质子集中进行比较，发现促炎途径如核因子κB和细胞因子/趋化因子信号传导、防御反应的正调控、免疫反应和适应性免疫系统中涉及的细胞活化等途径得到富集。虽然这些反应在间接感染中不存在，但细胞周期、愈合、细胞粘附调节、核心细胞外基质蛋白集合、细胞表面整合素以及介导整合素信号传导的蛋白质等细胞途径在间接感染中显著升高。除了对巨噬细胞蛋白质组进行全局分析外，我们还进一步验证了蛋白质组学数据，并证实MAP通过上皮细胞会调节吞噬细胞中整合素的表达和信号传导。在本研究中，我们证明间接感染的巨噬细胞中整合素的主要表达使吞噬细胞能够启动更强的结合并通过内皮细胞进行有效转运，这表明整合素在MAP感染传播中起着重要作用。
