4-hydroxy-2-nonenal and pimonidazole

Although the mechanisms responsible for acute liver failure (ALF) have not yet been fully elucidated, studies have indicated that intrahepatic macrophage activation plays an important role in the pathogenesis of ALF through intrahepatic microcirculatory disorder and consequent parenchymal cell death. Intrahepatic microcirculatory disorder has been demonstrated in animal models using intravital microscopy; however, the limitations of this method include simultaneously evaluating blood flow and the surrounding pathological changes. Therefore, in this study, we devised a novel method involving tetramethylrhodamine isothiocyanate (TRITC)-dextran administration for the pathological assessment of hepatic microcirculation. In addition, we aimed to elucidate the mechanisms through which intrahepatic microcirculatory disorder progresses with relation to activated macrophages. ALF was induced in Wistar rats by exposure to lipopolysaccharide and D-galactosamine. Intrahepatic microcirculation and microcirculatory disorder in zone 3 (pericentral zone) of the livers of rats with ALF was observed. Immunohistochemical examinations in conjunction with TRITC-dextran images revealed that the macrophages were mainly distributed in zone 2 (intermediate zone), while cleaved caspase-3-positive hepatocytes, pimonidazole and hypoxia-inducible factor 1-α were abundant in zone 3. We also found that 4-hydroxy-2-nonenal and nicotinamide adenine dinucleotide phosphate oxidase (NOX)4-positive cells were predominantly located in the zone 3 parenchyma. The majority of apoptotic hepatocytes in zone 3 were co-localized with NOX4. Our results revealed that the apoptotic cells in zone 3 were a result of hypoxic conditions induced by intrahepatic microcirculatory disorder, and were not induced by activated macrophages. The increased levels of oxidative stress in zone 3 may contribute to the progression of hepatocyte apoptosis.

Topics: Aldehydes; Animals; Apoptosis; Caspase 3; Disease Models, Animal; Fluconazole; Galactosamine; Hepatocytes; Hypoxia-Inducible Factor 1; Lipopolysaccharides; Liver; Liver Failure, Acute; Macrophages; Male; Microcirculation; NADPH Oxidase 4; NADPH Oxidases; Nitroimidazoles; Oxidative Stress; Rats; Rats, Wistar; Rhodamines; Up-Regulation

The effect of chronic enteral ethanol on pancreatic hypoxia was investigated using the hypoxia marker, pimonidazole. Male Wistar rats were fed an ethanol-containing diet for 3 weeks using an enteral model shown to cause pancreatic damage; pimonidazole (120 mg/kg i.v.) was injected 1h before sacrifice. Pimonidazole and 4-hydroxynonenal (an index of lipid peroxidation) adducts were detected immunochemically. Breathing air with low oxygen content (8% O(2)) for 1h increased pimonidazole adduct accumulation approximately 2-fold in pancreata of nai;ve rats, confirming that this technique will detect increases in hypoxia in pancreata. Pancreata of rats fed ethanol began to show signs of damage after 3 weeks. Ethanol feeding also significantly increased pimonidazole adducts in pancreas approximately 2-fold (1 or 3 weeks of ethanol produced similar values). Concomitant with increasing hypoxia in the pancreas, alcohol also caused a significant increase in 4-hydroxynonenal adducts, indicative of increased oxidative stress. These results indicate that chronic ethanol causes hypoxia at the cellular level in the pancreas in vivo; further, the data support the hypothesis that hypoxia is involved in mechanisms of chronic alcoholic pancreatitis.

Topics: Aldehydes; Amylases; Animals; Blood Pressure; Cell Hypoxia; Diet; Ethanol; Lipase; Male; Models, Biological; Nitroimidazoles; Oxidative Stress; Pancreas; Rats; Rats, Wistar; Time Factors