Tosyl gly pro arg p nitroanilide

Colonic inflammation was induced by treatment with DSS (MP Biomedicals, Illkirch, France 160110), dissolved in drinking water (5% weight/volume). The animals were free to drink the DSS solution for 7 days. For the whole period of DSS exposure, mice were daily treated by intragastric gavage with 5×10⁹ CFU (in 100 μL PBS) of either wild-type (WT) or recombinant L. lactis strains or PBS (100 μL). Body weight was measured daily after the induction of colitis. On day 7 after adding DSS to their drinking water, mice were sacrificed and colons were analyzed and harvested for measurement of inflammation parameters: macroscopic and microscopic damage score, bowel thickness, myeloperoxydase (MPO) activity as previously described [14 (link),17 (link)]. Upon sacrifice, the entire colon was excised and 1 ml PBS was instilled and washed twice through the lumen. Trypsin-like and elastase/PR-3-like activities were measured in those washes, using tosyl-Gly-Pro-Arg-p-nitroanilide (150 μM, Sigma) and MeO-succinyl-Ala-Ala-Pro-Val- p-nitroanilide (100 μM, Sigma) respectively as substrates. The change in absorbance at 405 nm was determined over 30-minutes at 37°C with a microplate reader NOVOstarTM (BMG Labtech, Champigny-sur-Marne, France). Activity was compared to known standard dilution of trypsin from porcine pancreas (Sigma) or human NE (Sigma) [14 (link),17 (link)].