Rats from the noncolitic and nontreated colitic groups received water orally. Colitis was induced using the method originally described by Morris et al [17]. After fasting overnight, the animals were anesthetized

with halothane. Under anesthesia, they were given 10 mg of trinitrobenzenesulfonic acid (TNBS) dissolved in 0.25 mL of 50% (vol/vol) ethanol by means of a Teflon (Dupont, Wilmington, Del) cannula inserted 8 cm into the anus. During and after TNBS administration, the rats were kept in a head-down position until they recovered from the anesthesia. Rats from the noncolitic (normal) group received 0.25 mL of saline. Animals from all groups were euthanized 7 days after colitis induction by an overdose of halothane. Animal body weights, the occurrence of diarrhea (as detected by perianal fur Galunisertib research buy soiling), and total food intake for each group were recorded daily. Once the rats were killed, the colon was removed aseptically, placed on an ice-cold plate, and longitudinally opened. The luminal contents were then collected for the microbiological studies (see below). Afterward, the colonic segment was cleaned of fat and mesentery Panobinostat solubility dmso and

blotted on a filter paper. Each specimen was weighed, and its length was measured under a constant load (2 g). The colon was scored for macroscopically visible damage on a 0 to 10 scale by 2 observers who were unaware of the treatment, in accordance with the criteria described by Bell et al [18]. Representative whole-gut specimens were taken from a region of the inflamed colon corresponding to the segment adjacent to the gross macroscopic damage and were fixed in 4% buffered formaldehyde. Cross sections were selected and embedded in paraffin. Equivalent colonic

segments were also obtained from the noncolitic group. Full-thickness sections of 5 mm were obtained at different levels and were stained with hematoxylin and eosin. The histologic damage was evaluated by one observer who was blind to the experimental groups, according to the criteria much described previously by Stucchi et al [19]. The colon was subsequently divided into different longitudinal pieces to be used for the following biochemical determinations: myeloperoxidase (MPO) activity, alkaline phosphatase (AP) activity, and total glutathione (GSH) content. Myeloperoxidase activity was determined using the technique described by Krawisz et al [20]. The results are expressed as MPO units per gram of tissue. One unit of MPO activity was defined as the amount required to degrade 1 mmol of hydrogen peroxide per minute at 25°C. Alkaline phosphatase activity was determined spectrophotometrically using disodium nitrophenylphosphate (5.5 mmol/L) as the substrate buffered in 50 mmol/L glycine with 0.5 mmol/L MgCl2 at pH 10.5 [21]. The enzymatic activity is expressed as milliunits per milligram of protein [22].