T1807 Bacterial Proteolytic Profile in Inflammatory Bowel Diseases

Background: Proteolytic degradation of the extracellular matrix is a serious consequence of intestinal inflammation. It is hypothesized that, in addition to host matrix metalloproteases, the proteases produced by gut microflora might contribute to the pathogenesis of inflammatory bowel disease (IBD). Aim & Methods: To determine whether proteolytic activity is a feature of the human microflora, we screened the following sources for protease activity against various substrates: A metagenomic library constructed from the gut microflora of a healthy adult, an Escherichia coli strain collection, an extensive biobank of human commensal bacteria, and bacteria isolated from fecal samples of 23 patients with ulcerative colitis (UC), 18 with Crohn's disease (CD) and 20 healthy controls. Results: Gelatinolytic activity was detected primarily in association with gram-positive bacteria from the feces of patients with CD, UC, and healthy volunteers (HV). However, while this was predominantly linked with Clostridium perfringens in controls and UC, a different profile was found in CD, with a diversity of bacteria exhibiting gelatinolytic activity. Culture supernatants from C. perfringens were able to degrade gelatin, azocoll, type I collagen and basement membrane type IV collagen, but different isolates of C. perfringens varied in their proteolytic activity. In addition, although detection of the colA gene encoding collagenase in C. perfringens showed that this was not specific for either CD or UC vs controls, quantification found the highest levels in UC patients (UC > CD > HV). To demonstrate the functional significance of microbialderived proteolytic activity, rat distal colon was exposed to culture supernatants of C. perfringens in Ussing chambers. Culture supernatants of C. perfringens caused a decrease in transepithelial resistance (TER) and responsiveness to bethanechol. Conclusion: (i) proteolytic activity is associated with human gut microbiota and is not exclusively host-derived in IBD; (ii) bacteria-derived proteolytic activity may contribute to the pathogenesis of disease, but the profile differs between UC and CD.