BACKGROUND: The incidence and severity of Clostridium difficile colitis have increased dramatically in the last decade. Disease severity is related to C. difficile virulence factors, including toxins A and B, as well as the patient's immune status. The intestinal mucus is an important component of innate barrier function in the intestine. Phosphatidylcholine (PC) is a key constituent of the intestinal mucus barrier, and exogenous PC administration has had therapeutic efficacy in patients with ulcerative colitis. We studied the protective function of exogenous PC on C. difficile toxin effects on the intestinal barrier in vitro. METHODS: Mucus-producing (HT29-MTX strain) and non-mucus-producing (HT29 strain) intestinal epithelial monolayers were cocultured with PC and C. difficile toxin A added to the apical media. Basal chamber culture supernatants were subsequently obtained, and tumor necrosis factor and interleukin 6 were quantitated by enzyme-linked immunosorbent assay. In other experiments, HT29 toxin A uptake, intestinal monolayer permeability, necrosis, and actin microfilament disruption were determined. RESULTS: There was a threefold to fourfold decrease in tumor necrosis factor and interleukin 6 levels and similar decreases in toxin A uptake and permeability changes in intestinal epithelial cells with mucus or PC versus control. Intestinal epithelial cell necrosis was reduced by more than 50% with either mucus or PC versus control. The integrity of HT29 cell cytoskeleton was demonstrated by both the mucus layer of the HT29-MTX strain and by exogenous PC administration by phalloidin staining of actin microfilaments. CONCLUSION: PC supplementation was effective in improving intestinal barrier defense against C. difficile toxin A challenge. PC administration may be a useful therapeutic adjunct in severe cases of C. difficile colitis or in patients who do not improve with conventional treatment.
BACKGROUND: The incidence and severity of Clostridium difficilecolitis have increased dramatically in the last decade. Disease severity is related to C. difficile virulence factors, including toxins A and B, as well as the patient's immune status. The intestinal mucus is an important component of innate barrier function in the intestine. Phosphatidylcholine (PC) is a key constituent of the intestinal mucus barrier, and exogenous PC administration has had therapeutic efficacy in patients with ulcerative colitis. We studied the protective function of exogenous PC on C. difficile toxin effects on the intestinal barrier in vitro. METHODS: Mucus-producing (HT29-MTX strain) and non-mucus-producing (HT29 strain) intestinal epithelial monolayers were cocultured with PC and C. difficile toxin A added to the apical media. Basal chamber culture supernatants were subsequently obtained, and tumor necrosis factor and interleukin 6 were quantitated by enzyme-linked immunosorbent assay. In other experiments, HT29 toxin A uptake, intestinal monolayer permeability, necrosis, and actin microfilament disruption were determined. RESULTS: There was a threefold to fourfold decrease in tumor necrosis factor and interleukin 6 levels and similar decreases in toxin A uptake and permeability changes in intestinal epithelial cells with mucus or PC versus control. Intestinal epithelial cell necrosis was reduced by more than 50% with either mucus or PC versus control. The integrity of HT29 cell cytoskeleton was demonstrated by both the mucus layer of the HT29-MTX strain and by exogenous PC administration by phalloidin staining of actin microfilaments. CONCLUSION:PC supplementation was effective in improving intestinal barrier defense against C. difficile toxin A challenge. PC administration may be a useful therapeutic adjunct in severe cases of C. difficilecolitis or in patients who do not improve with conventional treatment.
Authors: Julajak Limsrivilai; Krishna Rao; Ryan W Stidham; Shail M Govani; Akbar K Waljee; Andrew Reinink; Laura Johnson; Emily Briggs; Peter D R Higgins Journal: Dig Dis Sci Date: 2018-04-12 Impact factor: 3.199
Authors: Michael G Dieterle; Rosemary Putler; D Alexander Perry; Anitha Menon; Lisa Abernathy-Close; Naomi S Perlman; Aline Penkevich; Alex Standke; Micah Keidan; Kimberly C Vendrov; Ingrid L Bergin; Vincent B Young; Krishna Rao Journal: mBio Date: 2020-05-05 Impact factor: 7.867