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Environmental heat stress is associated with an age-related increase in hepatic oxidative damage and an exaggerated state of oxidative stress. The purpose of this investigation was to evaluate the regulation of hepatic iron after heat stress. A secondary aim was to determine a potential role for iron in heat stress-induced liver injury. Hyperthermia-induced alterations in hepatic iron were evaluated in young (6 mo) and old (24 mo) Fischer 344 rats by exposing them to a two-heat stress protocol. Livers were harvested at several time points after the second heating and assayed for labile and nonheme iron. In the control condition, there was no difference in labile iron between age groups. Both labile iron and storage iron were not altered by hyperthermia in young rats, but both were increased immediately after heating in old rats. To evaluate a role for iron in liver injury, hepatic iron content was manipulated in young and old rats, and then both groups were exposed to heat stress. Iron administration to young rats significantly increased hepatic iron content and ferritin but did not affect markers of lipid peroxidation under control conditions or after heat stress. In old rats, iron chelation with deferoxamine prevented the increase in nonheme iron, labile iron, ferritin, and lipid peroxidation after heat stress. These results suggest that iron may play a role in hepatic injury after hyperthermia. Thus, dysregulation of iron may contribute to the gradual decline in cellular and physiological function that occurs with aging. reactive oxygen species; deferoxamine; hyperthermia; electron paramagnetic resonance spectroscopy; inflammation

Recent studies have implicated dysregulation of the endocannabinoid system in various liver diseases and their complications (e.g., hepatitis, fibrosis, cirrhosis, cirrhotic cardiomyopathy, and ischemia-reperfusion), and demonstrated that its modulation by either cannabinoid 2 ([CB.sub.2]) receptor agonists or [CB.sub.1] antagonists may be of significant therapeutic benefits. This review is aimed to focus on the triggers and sources of endocannabinoids during liver inflammation and on the novel role of [CB.sub.2] receptors in the interplay between the activated endothelium and various inflammatory cells (leukocytes, lymphocytes, etc.), which play pivotal role in the early development and progression of inflammatory and other liver diseases. ischemia-reperfusion; endocannabinoids; cannabinoid 2 receptor; inflammation; endothelium

CCAAT/enhancer-binding protein (C/EBP) homologous protein (CHOP) is a key component in endoplasmic reticulum (ER) stress-mediated apoptosis. The goal of the study was to investigate the role of CHOP in cholestatic liver injury. Acute liver injury and liver fibrosis were assessed in wild-type (WT) and CHOP-deficient mice following bile duct ligation (BDL). In WT livers, BDL induced overexpression of CHOP and Bax, a downstream target in the CHOP-mediated ER stress pathway. Liver fibrosis was attenuated in CHOP-knockout mice. Expression levels of co-smooth muscle actin and transforming growth factor-[beta]1 were reduced, and apoptotic and necrotic hepatocyte death were both attenuated in CHOP-deficient mice. Hepatocytes were isolated from WT and CHOP-deficient mice and treated with 400 [micro]M glycochenodeoxycholic acid (GCDCA) for 8 h to examine bile acid-induced apoptosis and necrosis. GCDCA induced overexpression of CHOP and Bax in isolated WT hepatocytes, whereas CHOP-deficient hepatocytes had reduced cleaved caspase-3 expression and a lower propidium iodide index after GCDCA treatment. In conclusion, cholestasis induces CHOP-mediated ER stress and triggers hepatocyte cell death, and CHOP deficiency attenuates this cell death and subsequent liver fibrosis. The results demonstrate an essential role of CHOP in development of liver fibrosis due to cholestatic liver damage. apoptosis; necrosis; endoplasmic reticulum stress; transforming growth factor-[beta]1; bile duct ligation

Sepsis is associated with an activation of the coagulation system and multiorgan failure. The aim of the study was to examine the effects of selective thrombin inhibition with melagatran on renal hemodynamics and function, and liver integrity, during early endotoxemia. Endotoxemia was induced in thiobutabarbital-anesthetized rats by an intravenous bolus dose of lipopolysaccharide (LPS; 6 mg/kg). Sham-Saline, LPS-Saline, and LPS-Melagatran study groups received isotonic saline or melagatran immediately before (0.75 [micro]mol/kg iv) and continuously during (0.75 [micro]mol*[kg.sup.-1] iv) 4.5 h of endotoxemia. Kidney function, renal blood flow (RBF), and intrarenal cortical and outer medullary perfusion (OMLDF) measured by laser-Doppler flowmetry were analyzed throughout. Markers of liver injury and tumor necrosis factor (TNF)-[alpha] were measured in plasma after 4.5 h of endotoxemia. In addition, liver histology and gene expression were examined. Melagatran treatment prevented the decline in OMLDF observed in the LPS-Saline group (P < 0.05, LPS-Melagatran vs. LPS-Saline). However, melagatran did not ameliorate reductions in mean arterial pressure, RBF, renal cortical perfusion, and glomerular filtration rate or attenuate tubular dysfunctions during endotoxemia. Melagatran reduced the elevated plasma concentrations of aspartate aminotransferase (-34 [+ or -] 11%, P < 0.05), alanine aminotransferase (-21 [+ or -] 7%, P < 0.05), bilirubin (-44 [+ or -] q%, P < 0.05), and TNF-[alpha][(-32 [+ or -] 14%, P < 0.05) in endotoxemia. Melagatran did not diminish histological abnormalities in the liver or the elevated hepatic gene expression of TNF-[alpha], intercellular adhesion molecule-1, and inducible nitric oxide synthase in endotoxemic rats. In summary, thrombin inhibition with melagatran preserved renal OMLDF, attenuated liver dysfunction, and reduced plasma TNF-[alpha] levels during early endotoxemia. acute kidney failure; acute liver failure; coagulation; multiple organ failure; sepsis

We used a congenic C57B1/6J cystic fibrosis transmembrane conductance regulator [(Cftr).sup.-/-] mouse model, which develops cystic fibrosis (CF)-like pathology in all organs, to evaluate the short- and long-term therapeutic effects of dietary docosahexaenoic acid (DHA). Thirty-day-old [(Cftr).sup.-/-] mice and wild-type littermates were randomized to receive a liquid diet with or without DHA (40 mg/day). Animals were killed for histological and lipid analysis after 7, 30, and 60 days of therapy. DHA had no significant therapeutic or harmful effect on the lung, pancreas, or ileum of the [(Cftr).sup.-/-] mice or their wild-type littermates. In contrast, dietary DHA resulted in highly significant amelioration of the severity of liver disease in the [(Cftr).sup.-/-] mice, primarily a reduction in the degree of peri-portal inflammation. Additionally, these detailed measurements confirm our previous findings that [(Cftr).sup.-/-] mice have significant alterations in the pancreas (except external acinar diameter), ileum, liver, lung, and salivary (except sublingual) glands at all ages compared with their age-matched wild-type littermates. In conclusion, inhibition of cytokines and/or eicosanoid metabolism and release of endogenous inhibitors of inflammation by DHA may account for the anti-inflammatory effects in the liver of this congenic murine model of CF. The potential therapeutic benefits of DHA in severe CF-associated liver disease remain to be explored. cystic fibrosis transmembrane conductance regulator; postnatal organ development; anti-inflammatory effect; cystic fibrosis-associated liver disease

Nonalcoholic fatty liver disease is the most common reason for abnormal liver chemistries in the United States. The factors that lead from benign steatosis to nonalcoholic steatohepatitis are poorly understood. Transthyretin-Abcb11 (TTR-Abcb11) transgenic mice overexpress the bile salt transporter Abcb11 and hypersecrete biliary lipids. Thus the aim of this study is to employ feeding of the methionine-choline-deficient (MCD) diet to TTR-Abcb11 transgenic mice to further determine the mechanisms responsible for the development of steatohepatitis. FVB/NJ and TTR-Abcb11 mice were fed control or MCD diets for up to 30 days. Serum aminotransferase levels, serum and hepatic triglyceride content, cytokines, markers of oxidative stress, and expression of selective genes were examined. MCD diet-fed TTR-Abcb11, but not wild-type, mice have elevated serum aminotransferase levels when compared after 7 days. They also have significantly lower hepatic triglyceride levels at all time points studied. After 14 days on the MCD diet, TTR-Abcb11 mice have 3-fold increases in TNF-[alpha] mRNA and 3.9-fold increases in IL-6 mRNA compared with FVB/NJ mice. TTR-Abcb11 mice also had a greater increase in cytochrome P-450 2E1 expression. A greater decrease in sterol regulatory element binding protein-1c and fatty acid synthase mRNA expression was also seen in TTR-Abcb11 compared with wild-type mice fed an MCD diet. They also have enhanced TNF-[alpha] IL-6, and cytochrome P-450 2E1 expression. We conclude that TTR-Abcb11 mice develop a more rapid hepatitis with less steatosis. nonalcoholic steatohepatitis; oxidative stress; cytokines

Staphylococcal enterotoxin B potentiates LPS-induced hepatic dysfunction in chronically catheterized rats. Am J Physiol Gastrointest Liver Physiol 280: G866-G872, 2001.--Most models of liver dysfunction in sepsis use endotoxin (lipopolysaccharide; LPS) to induce a pathophysiological response. In our study published in this issue (Beno DWA, Uhing MR, Goto M, Chen Y, Jiyamapa-Serna VA, and Kimura RE. Am J Physiol Gastrointest Liver Physiol 280: G858-G865, 2001), the adverse effect of LPS on hepatic function in vivo was only significant at relatively high LPS doses despite high tumor necrosis factor-[Alpha] concentrations. However, many patients with sepsis are exposed to multiple bacterial toxins that may augment the immune response, resulting in increased hepatic dysfunction. We have developed a model of polymicrobial sepsis by parentally administering a combination of staphylococcal enterotoxin B (SEB) and LPS. Using this model, we demonstrate that SEB (50 [micro]g/kg) potentiates the effect of LPS-induced hepatic dysfunction as measured by decreased rates of biliary indocyanine green clearance and bile flow. These increases were most pronounced with doses of 10 and 100 [micro]g/kg LPS, doses that by themselves do not induce hepatic dysfunction. This may explain the seemingly increased incidence and severity of liver dysfunction in sepsis, and it suggests that the exclusive use of LPS for replicating septic shock may not be relevant for studies of hepatic dysfunction. interferon-[Gamma]; hepatocellular dysfunction; sepsis; cholestasis; in vivo studies

Teckman, Jeffrey H., Reid Gilmore, and David H. Perlmutter. Role of ubiquitin in proteasomal degradation of mutant [[Alpha].sub.1]-antitrypsin Z in the endoplasmic reticulum. Am. J. Physiol. Gastrointest. Liver Physiol. 278: G39-G48, 2000.--A delay in intracellular degradation of the mutant [[Alpha].sub.1]-antitrypsin ([[Alpha].sub.1]AT)Z molecule is associated with greater retention within the endoplasmic reticulum (ER) and susceptibility to liver disease in a subgroup of patients with [[Alpha].sub.1]AT deficiency. Recent studies have shown that [[Alpha].sub.1]ATZ is ordinarily degraded in the ER by a mechanism that involves the proteasome, as demonstrated in intact cells using human fibroblast cell lines engineered for expression of [[Alpha].sub.1]ATZ and in a cell-free microsomal translocation assay system programmed with purified [[Alpha].sub.1]ATZ mRNA. To determine whether the ubiquitin system is required for proteasomal degradation of [[Alpha].sub.1]ATZ and whether specific components of the ubiquitin system can be implicated, we have now used two approaches. First, we overexpressed a dominant-negative ubiquitin mutant (UbK48RG76A) by transient transfection in the human fibroblast cell lines expressing [[Alpha].sub.1]ATZ. The results showed that there was marked, specific, and selective inhibition of [[Alpha].sub.1]ATZ degradation mediated by UbK48R-G76A, indicating that the ubiquitin system is at least in part involved in ER degradation of [[Alpha].sub.1]ATZ. Second, we subjected reticulocyte lysate to DE52 chromatography and tested the resulting well-characterized fractions in the cell-free system. The results showed that there were both ubiquitin-dependent and -independent proteasomal mechanisms for degradation of [[Alpha].sub.1]ATZ and that the ubiquitin-conjugating enzyme E2-F1 may play a role in the ubiquitin-dependent proteasomal mechanism. [[Alpha].sub.1]-antitrypsin deficiency; liver disease; emphysema; quality-control apparatus; protein degradation; endoplasmic reticulum