Formaldehyde

The chemicals used in this study were obtained from the following sources: resveratrol from CTMedChem (Bronx, NY); U0126 and D609 from Tocris Bioscience (Ellisville, MO); alachlor and lindane from Chem Service (West Chester, PA); arachidonic acid from Cayman Chemical (Ann Arbor, MI); 1-monolaurin (Lauricidin®) from Med-Chem Laboratories (Galena, IL); TPA from Biomol International (Plymouth Meeting, PA); 1-methylanthracene, 1-methylfluorene, benzoylperoxide (Luperox® A98), 18-β-glycyrrhetinic acid, dicumylperoxide, EGF, fluoranthene, fluorene, Lucifer Yellow CH dilithium salt(MW 457.25), pentachlorophenol, perfluorodecanoic acid (PFDA), perfluorooctane sulfonic acid (PFOSA), phenanthrene, pyrene and thrombin receptor activator peptide 6 (TRAP-6) from Sigma-Aldrich (St. Louis, MO); 9,10-dimethylanthracene, 1-methylpyrene, PFOA and 2,2',4,4',5,5'-hexachlorobiphenyl (PCB153) from Fluka (Buchs, Switzerland); 4,4'-(2,2,2-trichloroethane-1,1-diyl)bis(chlorobenzene) (DDT) from Supelco (Bellefonte, PA); R59022 from Calbiochem (La Jolla, CA); acetonitrile, dimethylsulfoxide (DMSO), ethanol and formaldehyde from Mallinckrodt Baker (Phillipsburg NJ).

Mice were sacrificed 2 days after infection, and their colonic tissues were harvested for histopathologic examination. The colons were cut longitudinally and fixed with 10% formaldehyde (Mallinckrodt Pharmaceuticals, St Louis, MO, USA). The colonic tissue was embedded in paraffin and sectioned (5 µm) prior to hematoxylin and eosin staining (H&E stain). The stained, sectioned tissue was imaged through microscopy, and the integrity of the tissue was assessed. For pathological scoring, six fields per sample were examined and scored. Average counts of neutrophils in the six high-power fields (HPFs) in tissues were determined. The severity of colitis was scored, ranging between zero and three points for each of the following parameters: (1) polymorphonuclear infiltrate; (2) mononuclear infiltrate; (3) edema; (4) erosion and ulcerations; (5) crypt abscess; (6) crypt destruction; and (7) distribution of inflammation (mucosa = 1, mucosa and submucosa = 2, transmural inflammation = 3). Based on the total score (ranging from 0 to 21), the extent of tissue inflammation was graded as mild (1–5 points), moderate (6–10 points), or severe (>10 points) [25 (link)]. The markers of inflammation, including neutrophil infiltration, epithelial damage, and irregular shape of colon mucosa, were analyzed.

The passage 0 generation cells were grown in media until 90% confluence was achieved and reseeded at 10 × 10³, 5 × 10³, and 1 × 10³ cells into new culture disks and incubated for 7–14 days. When the cell colonies grew to include approximately 50 cells, the media were removed, washed twice with PBS, and fixed with 10% formaldehyde (H121-08; Mallinckrodt Chemicals, Phillipsburg) for 30 min. After washing twice with PBS, the cells were stained with 0.1% crystal violet (C3886; Sigma-Aldrich) for 15 min and then washed 3–5 times with double distilled water. The dishes were subsequently photographed to determine the number of colony-forming units.