Shotgun Proteomic Analysis of Plasma from Dairy Cattle Suffering from Footrot: Characterization of Potential Disease-Associated Factors

The plasma proteome of healthy dairy cattle and those with footrot was investigated using a shotgun LC-MS/MS approach. In total, 648 proteins were identified in healthy plasma samples, of which 234 were non-redundant proteins and 123 were high-confidence proteins; 712 proteins were identified from footrot plasma samples, of which 272 were non-redundant proteins and 138 were high-confidence proteins. The high-confidence proteins showed significant differences between healthy and footrot plasma samples in molecular weight, isoelectric points and the Gene Ontology categories. 22 proteins were found that may differentiate between the two sets of plasma proteins, of which 16 potential differential expression (PDE) proteins from footrot plasma involved in immunoglobulins, innate immune recognition molecules, acute phase proteins, regulatory proteins, and cell adhesion and cytoskeletal proteins; 6 PDE proteins from healthy plasma involved in regulatory proteins, cytoskeletal proteins and coagulation factors. Of these PDE proteins, haptoglobin, SERPINA10 protein, afamin precursor, haptoglobin precursor, apolipoprotein D, predicted peptidoglycan recognition protein L (PGRP-L) and keratan sulfate proteoglycan (KS-PG) were suggested to be potential footrot-associated factors. The PDE proteins PGRP-L and KS-PG were highlighted as potential biomarkers of footrot in cattle. The resulting protein lists and potential differentially expressed proteins may provide valuable information to increase understanding of plasma protein profiles in cattle and to assist studies of footrot-associated factors.

本研究采用鸟枪法液相色谱-串联质谱（shotgun LC-MS/MS）技术，对健康奶牛与患牛蹄腐病奶牛的血浆蛋白质组展开分析。健康奶牛血浆样本中共鉴定出648种蛋白质，其中非冗余蛋白质234种、高置信度蛋白质123种；患牛蹄腐病奶牛血浆样本中共鉴定出712种蛋白质，其中非冗余蛋白质272种、高置信度蛋白质138种。高置信度蛋白质在分子量、等电点以及基因本体（Gene Ontology）分类层面，于健康组与患病组血浆样本间存在显著差异。研究筛选出22种可用于区分两组血浆蛋白质的潜在标志物，其中来自患病组血浆的16种潜在差异表达（potential differential expression，PDE）蛋白质涉及免疫球蛋白、先天免疫识别分子、急性期蛋白、调控蛋白以及细胞黏附与细胞骨架蛋白；来自健康组血浆的6种PDE蛋白质则涉及调控蛋白、细胞骨架蛋白与凝血因子。在上述PDE蛋白质中，触珠蛋白（haptoglobin）、SERPINA10蛋白、运白蛋白前体（afamin precursor）、触珠蛋白前体、载脂蛋白D（apolipoprotein D）、预测性肽聚糖识别蛋白L（predicted peptidoglycan recognition protein L, PGRP-L）以及硫酸角质素蛋白聚糖（keratan sulfate proteoglycan, KS-PG）被认定为潜在的牛蹄腐病相关因子。其中PDE蛋白质PGRP-L与KS-PG被确定为奶牛牛蹄腐病的潜在生物标志物。本研究得到的蛋白质列表与潜在差异表达蛋白质数据集，可为解析奶牛血浆蛋白质组特征以及开展牛蹄腐病相关因子研究提供极具价值的参考信息。