Impaired luminal processing of human defensin-5 in Crohn's disease: persistence in a complex with chymotrypsinogen and trypsin

Am J Pathol. 2008 Mar;172(3):702-13. doi: 10.2353/ajpath.2008.070755. Epub 2008 Feb 7.

Abstract

Human defensin (HD)-5 is an antimicrobial peptide expressed in small intestinal Paneth cells, and alterations in HD-5 expression may be important in Crohn's disease (CD) pathogenesis. Levels of HD-5 in Paneth cells and ileostomy fluid from control and CD patients were studied by quantitative immunodot analysis, immunohistochemistry, acid urea-polyacrylamide gel electrophoresis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis Western blotting, reverse phase-high performance liquid chromatography, N-terminal amino acid sequencing, and ES-QToF mass spectrometry. In both control and CD patients, HD-5 in Paneth cell extracts was present almost exclusively in the precursor form. HD-5 levels in ileostomy fluid were lower in CD patients (n = 51) than in controls (n = 20): median (range), 7.9 (5.5 to 35.0) microg/ml versus 10.5 (6.0 to 30.4) microg/ml; P = 0.05; this difference was most marked in CD patients with homozygous/compound heterozygous mutations in NOD2 (P = 0.03). In control ileostomy fluid, HD-5 was present in the mature form only. In contrast, CD patient ileostomy fluid contained both precursor and mature forms of HD-5, with the majority present in a complex with trypsin, chymotrypsinogen/chymotrypsin, and alpha1-anti-trypsin. Pro-HD-5 was not associated with trypsin or chymotrypsinogen in Paneth cell extracts. In conclusion, pro-HD-5 in the intestinal lumen is processed by trypsin in a complex in which chymotrypsinogen is also cleaved for activation. The persistence of this complex in CD may be attributable to increased luminal levels of proteinase inhibitors such as alpha1-anti-trypsin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Amino Acid Sequence
  • Body Fluids / chemistry
  • Body Fluids / metabolism
  • Case-Control Studies
  • Chromatography, High Pressure Liquid
  • Chymotrypsinogen / metabolism*
  • Crohn Disease / metabolism*
  • Defensins / metabolism*
  • Defensins / pharmacokinetics
  • Female
  • Humans
  • Ileostomy
  • Intestinal Mucosa / metabolism*
  • Intestine, Small / metabolism
  • Male
  • Middle Aged
  • Models, Biological
  • Molecular Sequence Data
  • Multiprotein Complexes / metabolism
  • Protein Precursors / metabolism
  • Protein Processing, Post-Translational*
  • Tissue Extracts / metabolism
  • Trypsin / metabolism*
  • alpha 1-Antitrypsin / metabolism
  • alpha-Defensins / metabolism*
  • alpha-Defensins / pharmacokinetics

Substances

  • DEFA5 protein, human
  • Defensins
  • Multiprotein Complexes
  • Protein Precursors
  • Tissue Extracts
  • alpha 1-Antitrypsin
  • alpha-Defensins
  • Chymotrypsinogen
  • Trypsin