title: Characterisation of faecal protease activity in irritable bowel syndrome with diarrhoea: origin and effect of gut transit.
abstract: OBJECTIVE
  Faecal serine proteases (FSPs) may play a role in irritable bowel syndrome with diarrhoea (IBS-D), but their origin is unclear. We aimed to structurally characterise them and define the impact of colonic cleansing and transit time.
  
  
  METHODS
  Faecal samples were obtained from 30 healthy volunteers (HV) and 79 patients with IBS-D participating in a trial of ondansetron versus placebo. Colonic transit was measured using radio-opaque markers. Samples were also obtained from 24 HV before and after colonic cleansing with the osmotic laxative MoviPrep. FSPs were purified from faecal extracts using benzamidine-Sepharose affinity chromatography. SDS-PAGE profiled components were identified using trypsinolysis and tandem mass spectrometry. Functional protease activity in faecal extracts was measured using a colorimetric assay based on the proteolysis of azo-casein.
  
  
  RESULTS
  Protein analysis identified the most abundant FSPs as being of human origin and probably derived from pancreatic juice. Functional assays showed increased faecal protease (FP) and amylase in patients with IBS-D compared with HV. Those with higher amylase had significantly higher FP and greater anxiety. FP activity correlated negatively with whole gut transit in patients with IBS-D (Spearman r=-0.32, p=0.005) and HV (r=-0.55, p=0.014). Colon cleansing caused a significant rise in FP activity in HV from a baseline of median (IQR) 253 (140-426) to 1031 (435-2296), levels similar to those seen in patients with IBS-D. FSP activity correlated positively with days/week with urgency.
  
  
  CONCLUSIONS
  The most abundant FSPs are of human origin. Rapid transit through the colon and/or decreased (possibly bacterial) proteolytic degradation increases their faecal concentration and could contribute to visceral hypersensitivity in patients with IBS-D.
  
  
  BACKGROUND
  NCT00745004.
authors.affiliation: School of Biomedical Sciences, University of Nottingham, , Nottingham, Nottinghamshire, UK.
authors.forename: David
authors.initials: D
authors.lastname: Tooth
authors.forename: Klara
authors.initials: K
authors.lastname: Garsed
authors.forename: Gulzar
authors.initials: G
authors.lastname: Singh
authors.forename: Luca
authors.initials: L
authors.lastname: Marciani
authors.forename: Ching
authors.initials: C
authors.lastname: Lam
authors.forename: Imogen
authors.initials: I
authors.lastname: Fordham
authors.forename: Annie
authors.initials: A
authors.lastname: Fields
authors.forename: Rawinder
authors.initials: R
authors.lastname: Banwait
authors.forename: Melanie
authors.initials: M
authors.lastname: Lingaya
authors.forename: Robert
authors.initials: R
authors.lastname: Layfield
authors.forename: Maggie
authors.initials: M
authors.lastname: Hastings
authors.forename: Peter
authors.initials: P
authors.lastname: Whorwell
authors.forename: Robin
authors.initials: R
authors.lastname: Spiller
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D015415:Biomarkers / Q000378:metabolism; D016022:Case-Control Studies; D003106:Colon / Q000187:drug effects / Q000201:enzymology / Q000382:microbiology; D003967:Diarrhea / Q000201:enzymology* / Q000382:microbiology / Q000503:physiopathology; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005243:Feces / Q000201:enzymology*; D005260:Female; D005772:Gastrointestinal Transit; D006801:Humans; D043183:Irritable Bowel Syndrome / Q000188:drug therapy / Q000201:enzymology* / Q000382:microbiology / Q000503:physiopathology; D054368:Laxatives / Q000008:administration & dosage / Q000494:pharmacology; D016015:Logistic Models; D008297:Male; D008875:Middle Aged; D010196:Pancreatic Elastase / Q000378:metabolism; D011092:Polyethylene Glycols / Q000008:administration & dosage / Q000494:pharmacology; D057057:Serine Proteases / Q000378:metabolism*; D053719:Tandem Mass Spectrometry; D055815:Young Adult; D000516:alpha-Amylases / Q000378:metabolism
chemical_list: D015415:Biomarkers; D054368:Laxatives; C520497:MoviPrep; D011092:Polyethylene Glycols; D000516:alpha-Amylases; D057057:Serine Proteases; D010196:Pancreatic Elastase
keywords: DIARRHOEA; IRRITABLE BOWEL SYNDROME
references.citation: Gut. 2007 Dec;56(12):1770-98
references.pmid: 17488783
references.citation: Am J Physiol Gastrointest Liver Physiol. 2007 Jul;293(1):G198-203
references.pmid: 17510196
references.citation: Clin Gastroenterol Hepatol. 2007 May;5(5):550-5
references.pmid: 17336590
references.citation: Scand J Gastroenterol. 1977;12(5):525-9
references.pmid: 918546
references.citation: Digestion. 1983;27(1):8-15
references.pmid: 6554206
references.citation: Gastroenterology. 1987 Jan;92(1):40-7
references.pmid: 3023168
references.citation: Gastroenterology. 1995 Jul;109(1):40-52
references.pmid: 7797041
references.citation: J Clin Invest. 2007 Mar;117(3):636-47
references.pmid: 17304351
references.citation: Pain. 2005 Jan;113(1-2):141-7
references.pmid: 15621374
references.citation: Clin Chem Lab Med. 2008;46(9):1275-80
references.pmid: 18597588
references.citation: Am J Pathol. 1998 Sep;153(3):937-44
references.pmid: 9736042
references.citation: Clin Chim Acta. 2000 Feb 15;291(2):201-21
references.pmid: 10675724
references.citation: J Biol Chem. 2004 Apr 2;279(14):13532-9
references.pmid: 14726524
references.citation: Anal Biochem. 1976 May 7;72:248-54
references.pmid: 942051
references.citation: Pain. 2009 Nov;146(1-2):41-6
references.pmid: 19595511
references.citation: Gut. 2010 Sep;59(9):1213-21
references.pmid: 20650926
references.citation: Gastroenterology. 2009 Oct;137(4):1425-34
references.pmid: 19596012
references.citation: J Appl Bacteriol. 1988 Jan;64(1):37-46
references.pmid: 3127369
references.citation: Scand J Gastroenterol. 1977;12(7):801-9
references.pmid: 594651
references.citation: Am J Pathol. 2002 Nov;161(5):1903-15
references.pmid: 12414536
references.citation: Gut. 2008 May;57(5):591-9
references.pmid: 18194983
references.citation: Neurogastroenterol Motil. 2008 Aug;20(8):900-7
references.pmid: 18363639
references.citation: Gastroenterology. 2007 Jan;132(1):26-37
references.pmid: 17241857
references.citation: Int J Proteomics. 2012;2012:735132
references.pmid: 23008771
references.citation: Anal Biochem. 1995 Jan 1;224(1):451-5
references.pmid: 7710111