Abstract Background Enhanced intestinal proteolytic activity contributes to the pathophysiology of inflammatory bowel disease (IBD) through multiple mechanisms, including increased intestinal barrier permeability and activation of immune responses. The relative contribution of host- and microbe-derived proteases to total fecal protease activity, and the specific identity of the overactive proteases, are not sufficiently understood. Moreover, limited therapeutic strategies targeting these proteases exist. Potatoes naturally produce pathogen-defensive protease inhibitors and current technologies enable the production of a high-yield protease inhibitor (PI) rich fraction. The aim of this study is to identify the overactive faecal proteases present in IBD and to assess the inhibitory capacity of potato PI fraction. Methods Faecal water from 17 healthy controls (HC), 15 Crohn’s disease (CD), and 14 ulcerative colitis (UC) patients were prepared from frozen stools. Protease activity of faecal water was assessed against gelatin, elastin, fibronectin and OmniMMP, and was further characterized by gelatin zymography. The human and bacterial protein composition was determined by label-free quantitative LC-MS/MS analysis. Correlation analyses between protease activity and proteomic profiles were performed to identify overactive human and bacterial proteases. The anti-protease activity of potato PI fraction on IBD faecal water was compared to aprotinin, elafin and α1-antitrypsin. Results Compared to HC, protease activity was significantly enhanced in IBD fecal water against all substrates tested and caused by serine proteases of 25-30, 37-50 and 100-150 kDa. While human versus bacterial protein counts were approximately equal in HC fecal water, the human protein fraction was significantly increased to ∼80% in CD and UC and were 2.5- and 4.2-fold higher than in HC, respectively. Thirty-one (31) proteases were differentially abundant in IBD fecal water. Human proteases strongly correlated (r ≥ 0.60) with protease activity, most significantly for CELA2A, ELANE, CPA5, PRTN3, CTRC, CPA1, CTSC and PRSS2 in CD, and PRSS2, CELA2A, CTRC, PRSS1 and CTSC in UC. Only 5 bacterial proteases correlated moderately with protease activity in CD (r = 0.48-0.57), while none were found in UC. Of the four protease inhibitors evaluated, potato PI fraction demonstrated the strongest inhibitory effect, reducing the gelatinolytic activity to HC levels. Conclusion The enhanced protease activity in IBD faeces is mainly driven by human proteases and is efficiently suppressed by a protease inhibitor-rich fraction from potato, which may have therapeutic potential to reduce intestinal inflammation in IBD patients by restoring proteolytic homeostasis. Conflict of interest: Mrs. Herreman, Laure: L.Herreman is employed by Royal Avebe U.A. and has receveid a grant from the Carbohydrate Competence Center (CCC) supported by Royal Avebe U.A. Gacesa, Ranko: Grant: Partially funded by Janssen Pharmaceuticals (for projects unrelated to any of research presented at this conference) Personal Fees: Paid freelance consulting for Esox biologicals ltd. (for projects unrelated to any of research presented at this conference) Pibiri, Paola: No conflict of interest Jansen, Bernadien Honorin: No conflict of interest Vandooren, Jennifer: No conflict of interest Horvatovich, Péter: No conflict of interest Laus, Marc Christian: M. Laus is an employee of Royal Avebe U.A. Dijkstra, Gerard: Grant: Royal DSM Personal Fees: Consultancy fee from Astra-Zeneca and Speakers fee from Abbvie Faber, Klaas-Nico: K.N. Faber has received a grant from the Carbohydrate Competence Center (CCC) supported by Royal Avebe U.A.
Herreman et al. (Thu,) studied this question.