Proteomic Profiling and Targeted Validation Reveal a More Extensive Salivary than Serum Response to a Feed-Deprivation Protocol in Horses with Gastric Lesions

Gastric ulceration is common in horses and remains challenging to diagnose, as gastroscopy is the gold standard. This study aimed to characterise the salivary and serum protein changes associated with a feed-deprivation protocol. Eight horses underwent this protocol, and gastric lesions were assessed by gastroscopy at the end of the experimental period. Saliva and blood samples were collected pre- and post-protocol, and pooled samples from each biological fluid and condition were analysed by two-dimensional gel electrophoresis to compare protein profiles. Fifteen salivary and five serum proteins showed differential abundance and were identified using mass spectrometry. Nine proteins were subsequently validated by Western blotting in individual samples, confirming the proteomic findings. The validated proteins are involved in multiple pathways, including inflammatory and immune responses (interleukins 1-F5 and -F6, prolactin-inducible protein, alpha-1-antitrypsin), metabolic and transport processes (albumin, serotransferrin, apolipoprotein A-IV, fatty acid-binding protein 5), and mucosal protection mechanisms (carbonic anhydrase VI, apolipoprotein A-IV, prolactin-inducible protein, fatty acid-binding protein 5). Compared with serum, saliva showed a more dynamic post-protocol proteomic response, with more proteins displaying altered abundance, supporting its versatility as a non-invasive matrix. Overall, these findings characterise salivary and serum responses associated with the feed-deprivation protocol in horses with gastric lesions confirmed post-protocol.