Neomycin sulfate

Mice were treated with antimicrobials in drinking water. Antibiotics used were ampicillin (0.5 mg/ml) (Sigma Aldrich; St. Louis, MO), gentamicin sulfate (0.5 mg/ml) (Corning; Manassas, VA), metronidazole (0.5 mg/ml) (Spectrum Chemical, Gardena, CA), neomycin sulfate (0.5 mg/ml) (EMD Millipore; Billerica, MA), and vancomycin (0.25 mg/ml) (VWR Life Science). All of the antimicrobials were dissolved in non-acidified autoclaved reverse osmosis drinking water. Antimicrobial water was changed every week.

Multidrug-resistant Salmonella (ATCC 700408) was obtained from the American Type Culture Collection (Manassas, VA). Silver nitrate, sodium borohydride, sodium citrate, NaOH, tetracycline hydrochloride (TET), neomycin sulfate (NEO), kanamycin sulfate (KAN), tryptic soy broth (TSB), and tryptic soy agar (TSA) were purchased from Sigma-Aldrich (St. Louis, MO). Penicillin G sodium salt (PEN) and nitric acid (trace metal grade, 67%–70%) were from Fisher Scientific (Houston, TX). Ampicillin sodium salt (AMP) was from Research Products International Corp (Prospect, IL), and enoxacin hydrochloride (ENO) was from MP Biomedicals LLC (Solon, OH). The structures of the antibiotics are shown in Figure SI 1.

Mice were given the following antibiotics in autoclaved drinking water: ampicillin (1g/l; Sigma), vancomycin (500 mg/l; Henry Schein, Inc.) neomycin sulfate (1g/l; Sigma), and metronidazole (1g/l; Sigma; solubilized with 15 ml of 0.1N acetic acid/liter). Fresh antibiotics were supplied every three days. Mice were kept on antibiotics for the indicated time and killed for tumor and tissue analysis. Mice were administered IDENs resuspended in PBS by gavage or intravenous injection as described previously^{39 (link)}. In some experiments, neutralizing antibody (200 µg/mouse) for IL-17A (MAB421 R&D Systems) or control IgG was injected once every week for eight weeks. Apc^Min/+ and Apc^Min/+/MyD88^−/− littermates were euthanized at 3−4 months of age or when the first signs of morbidity appeared. The intestinal tract was removed immediately after euthanasia. The small intestine and colon were examined, and tumors were counted and measured using a stereomicroscope. Most polyps were adenomas, identified by their appearance as grossly visible, preinvasive lesions that were common in the small intestine of APC^Min/+ mice. Colonic lesions that displayed a focally invasive phenotype were categorized as tumors. Five-µm sections of paraffin-embedded specimens were cut and stained with H&E for histology analyses.