Predicted salivary human protease activity in experimental gingivitis revealed by endoProteo-FASP approach

Gingivitis is a highly prevalent oral condition that can be studied in humans via the 21-d experimental gingivitis model, which allows for investigations into the induction and resolution of gingivitis. In this study, we used the autolysis of saliva as a source of peptides to predict the activity of human proteases in saliva during induction and resolution of inflammation. Healthy volunteers, with no remarkable oral or systemic conditions, were recruited into the study and stimulated saliva samples were collected at days 0, 21, and 35 of experimental gingivitis. Plaque and gingival indices were recorded to ensure clinical induction and resolution. Saliva was auto-digested at 37°C for 18 h before identification of peptides by mass spectrometry. Protease prediction was carried out using Proteasix in silico with the identified peptides. A comparison of day 0 to days 21 and 35 showed changes in predicted protease activity. Correlation network analysis revealed that at day 21 the proteases became less connected and showed a potential for a dysregulated system; by day 35 the connectivity was returning towards similar conditions at day 0. This study demonstrates that changes in predicted proteases are apparent even in saliva collected from donors experiencing inflammation around three teeth.