Changes in the saliva proteome analysed by gel-proteomics in horses diagnosed with Equine gastric ulcer Syndrome (EGUS) at diagnosis and after successful treatment

Abstract

Equine gastric ulcer syndrome (EGUS) has a high prevalence, being currently one of the more frequent diseases in horses. Saliva is a source of biomarkers that change in EGUS and can potentially be used for treatment monitoring. We aimed to identify changes in the salivary proteome by using gel proteomics in horses with EGUS and also the possible changes that could occur after successful treatment for this disease. Saliva samples were collected from 9 horses with EGUS before and after treatment and 9 matched healthy con-trols. SDS-PAGE (1DE) and two-dimensional gel electrophoresis (2DE) were performed in these samples, and significantly different protein bands and spots were identified by mass spectrometry. In 1DE, increases in bands containing containing family A member 2 and immunoglobulin-heavy constant mu) and decreased in bands containing prolac-tin-inducible protein and Ribosomal proteinS9 were observed in EGUS horses compared to healthy ones. In addition decreases in bands containing Major allergen Equ c 1, Gluta-thione transferase and 6-phosphogluconate dehydrogenase, decarboxylating decreased in EGUS after successful treatment compared to pre-treatment were observed in EGUS horses after treatment. For the 2DE analysis, sets of saliva samples were organized into (a) three pools from a cohort of nine horses afflicted with EGUS before undergoing treatment, (b) another three pools were constituted from the same horses with EGUS after the treatment and (c) three pools from 9 healthy horses. Thioredoxin (TRX), a protein showing signifi-cant changes at 2DE before and after EGUS treatment, was validated by an ELISA kit in 12 horses with EGUS before and after treatment. Results of 2DE showed the hoses with EGUS have changes in saliva proteins compared to healthy horses, with increases in ADA, tri-osephosphate isomerase, keratins and immunoglobulin heavy constant mu and decreases in CA and albumin. These changes would indicate various physiopathological mecha-nisms such as the activation of the immune system, decrease in the stomach defence mechanisms and inflammation involved in this disease. In addition, the successfully treated horses presented lower expression levels of TRX after a successful treatment in proteomics and also measured with a commercially available ELISA kit. Overall, horses with EGUS have protein changes in their saliva when measured with gel proteomics compared with healthy horses, and they also showed changes after successful treatment. These proteins could be potential biomarkers for detection and monitoring treatment re-sponse in EGUS.