The mucosae of the respiratory, gastrointestinal, and urogenital tract are permanently exposed to the harmful influence of the oxygen-derived and other kinds of free radicals. Antioxidants found on the luminal epithelium surface are effective free radical scavengers that could represent "the first line of defense" against free radicals. Free radical damage has been implicated in a wide variety of pathological mucosal conditions, including gastric lesions induced by nonsteroidal antiinflammatory drugs 1,2) and ethanol, 2-4) gastric carcinogenesis, 5) intestinal ischaemia/reperfusion injury, 6) Salmonella typhimurium intestinal infection, 7) inflammatory bowel disease, 8,9) and even in a development of calcium oxalate crystals on urothelium.
10)The antioxidative properties of biological systems are frequently studied using a non-destructive spectroscopic method, electron paramagnetic resonance (EPR), and nitroxides as models of biological free radicals. 5,[11][12][13] Nitroxides are stable radicals-paramagnetic substances-which living systems metabolically transform into the non-paramagnetic substances (EPR silent), hydroxylamines. The main metabolic transformation of nitroxides is the reduction by ascorbic acid and thiol reducing agents 11,12,[14][15][16] which can be mediated by enzymatic mechanisms involving also mitochondria and microsomes. 16,17) Therefore, the kinetics of nitroxides metabolism provides useful information about the antioxidative properties of the investigated system.The antioxidative properties of gastrointestinal mucosa are intensively studied and regional differences in the stomach, 5) differences along the gastrointestinal tract, 18) and also species 19) and gender 20) related differences have been established. On the other hand, the studies of antioxidative properties of urogenital tract are very scarce. Therefore, the aim of our study is to evaluate the antioxidative properties of pig urinary bladder mucosa and compare them with those of gastric and intestinal mucosa using three different nitroxides and EPR spectroscopy. Our work focuses also on the types of antioxidants present in the urinary bladder mucosa.
MATERIALS AND METHODS
NitroxidesWe used water soluble nitroxides-spin probes, namely: ASL [N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl)-N-dimethyl-N-hydroxyethylammonium iodide], which has a permanently positive charge in water solutions, GluSL [4-(2,3,4,5,6-pentahydroxyhexanoylamino)-2,2,6,6-tetramethylpiperidinyl-1-oxyl], which is uncharged in water solutions, and PCA [2,2,5,5-tetramethylpyrroline-1-oxyl-3-carboxylic acid], which is partially ionized and therefore negatively charged in water solutions. The chemical structures of nitroxide compounds used are shown in Fig. 1. The nitroxides ASL, PCA, and GluSL were synthesized according to procedures published elsewhere.
21-23)Tissue Preparation Stomachs, small intestines, and urinary bladders from freshly slaughtered 7 month-old pigs of both sexes, weighing 90-110 kg, were obtained from a local slaughterhouse. A small segment of th...