title: An experimental platform using human intestinal epithelial cell lines to differentiate between hazardous and non-hazardous proteins.
abstract: Human intestinal epithelial cell lines (T84, Caco-2, and HCT-8) grown on permeable Transwell™ filters serve as models of the gastrointestinal barrier. In this study, this in vitro model system was evaluated for effectiveness at distinguishing between hazardous and non-hazardous proteins. Indicators of cytotoxicity (LDH release, MTT conversion), monolayer barrier integrity ([(3)H]-inulin flux, horseradish peroxidase flux, trans-epithelial electrical resistance [TEER]), and inflammation (IL-8, IL-6 release) were monitored following exposure to hazardous or non-hazardous proteins. The hazardous proteins examined include streptolysin O (from Streptococcus pyogenes), Clostridium difficile Toxins A and B, heat-labile toxin from enterotoxigenic Escherichia coli, listeriolysin O (from Listeria monocytogenes), melittin (from bee venom), and mastoparan (from wasp venom). Non-hazardous proteins included bovine and porcine serum albumin, bovine fibronectin, and ribulose bisphosphate carboxylase/oxygenase (RuBisco) from spinach. Food allergenic proteins bovine milk β-lactoglobulin and peanut Ara h 2 were also tested as was the anti-nutritive food protein wheat germ agglutinin. Results demonstrated that this model system effectively distinguished between hazardous and non-hazardous proteins through combined analysis of multiple cells lines and assays. This experimental strategy may represent a useful adjunct to multi-component analysis of proteins with unknown hazard profiles.
authors.affiliation: Mucosal Immunology & Biology Research Center, Massachusetts General Hospital, CNY 114 (114-3503), Charlestown, MA, 02129, United States; Department of Pediatrics, Harvard Medical School, Boston, MA, 02129, United States. Electronic address: bphurley@partners.org.
authors.forename: Bryan P
authors.initials: BP
authors.lastname: Hurley
authors.affiliation: Mucosal Immunology & Biology Research Center, Massachusetts General Hospital, CNY 114 (114-3503), Charlestown, MA, 02129, United States.
authors.forename: Waheed
authors.initials: W
authors.lastname: Pirzai
authors.affiliation: Mucosal Immunology & Biology Research Center, Massachusetts General Hospital, CNY 114 (114-3503), Charlestown, MA, 02129, United States.
authors.forename: Alex D
authors.initials: AD
authors.lastname: Eaton
authors.affiliation: DuPont Pioneer, 8325 NW 62nd Avenue, Johnston, IA, 50131, United States.
authors.forename: Marc
authors.initials: M
authors.lastname: Harper
authors.affiliation: DuPont Haskell, 1090 Elkton Road, Newark, DE, 19714, United States.
authors.forename: Jason
authors.initials: J
authors.lastname: Roper
authors.affiliation: DuPont Pioneer, 8325 NW 62nd Avenue, Johnston, IA, 50131, United States.
authors.forename: Cindi
authors.initials: C
authors.lastname: Zimmermann
authors.affiliation: DuPont Haskell, 1090 Elkton Road, Newark, DE, 19714, United States.
authors.forename: Gregory S
authors.initials: GS
authors.lastname: Ladics
authors.affiliation: DuPont Pioneer, 8325 NW 62nd Avenue, Johnston, IA, 50131, United States.
authors.forename: Raymond J
authors.initials: RJ
authors.lastname: Layton
authors.affiliation: DuPont Pioneer, 8325 NW 62nd Avenue, Johnston, IA, 50131, United States.
authors.forename: Bryan
authors.initials: B
authors.lastname: Delaney
mesh_terms: D000485:Allergens / Q000494:pharmacology*; D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology*; D002417:Cattle; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism; D004044:Dietary Proteins / Q000494:pharmacology*; D006801:Humans; D007422:Intestines / Q000187:drug effects / Q000473:pathology*; D037102:Lectins / Q000494:pharmacology*; D009375:Neoplasms, Glandular and Epithelial / Q000473:pathology*; D014688:Venoms / Q000494:pharmacology*
chemical_list: D000485:Allergens; D001427:Bacterial Toxins; D016207:Cytokines; D004044:Dietary Proteins; D037102:Lectins; D014688:Venoms
keywords: Cytotoxicity; Intestinal epithelial cells; Protein toxins