Baseline data for distribution of contaminants by natural disasters: results from a residential Houston neighborhood during Hurricane Harvey flooding.

Hurricane Harvey made landfall in Texas August 25, 2017, bringing massive rains and flooding that impacted soils in a residential neighborhood in East Houston. Trace elements, organochlorine pesticides, polycyclic aromatic hydrocarbons (PAHs), polybrominated diphenyl ether fire retardants (PBDEs) and polychlorinated biphenyls (PCBs) were determined in 24 soil samples. The highest concentrations found in soils were total PAHs, which ranged from 1,310 µg/kg to 85,700 µg/kg with a mean of 12,600 µg/kg. Analysis of specific PAH ratios indicate the source of the PAHs were dominated by pyrogenic rather than petrogenic sources. Chlordanes were detectable in the area where the likely local source is for ant control. The trace metal concentrations were below any environmental health concern concentrations but As, Cd, Hg, Pb, Se, Ag, Zn were enriched over the crustal abundance. While Hurricane Harvey was responsible for the redistribution of many contaminants, the large volume of rain and floodwater likely transported contaminants from the land areas and into the Houston Ship Channel and Galveston Bay. The findings from this study will serve as baseline data for determining the mobilization of contaminants caused by natural disasters.

[Ethanol into acetaldehyde bioconversion by mutant strains of Hansenula polymorpha Felcao de Morais & Dália Maia].

The influence of some factors (tris-HCl buffer, pH 8.0, concentration, use of different binding agents aeration modes, genetically determined peroxisome degradation damage) on biotransformation efficiency of 0.217 M (1%) ethanol to acetaldehyde at 30 degrees C by Hansenula polymorpha 7-4A (gcrl EAO) strain cells with glucose repression block was investigated. Optimal cultivation conditions for cells were selected. Bioconversion efficiency using 1 M tris-HCl buffer, pH 8.0, was found the highest one as compared with using the buffer in concentrations from 0.1 M to 3 M. The process efficiency when using tris(hydroxymethyl)aminomethane as binding acetaldehyde agent proved much higher than when using sodium bisulfite both at aeration by air stream and incubation on shaker. Using 146 and 179 mutants cells for bioconversion with defects in alcohol oxidase inactivation during macropexophagy stimulated efficiency increase by 5.58% and 8.10%, respectively, as compared with the use of parental 7-4A strain cells.

[Biodegradation of oil hydrocarbons by Candida yeast]. / Biodegradatsiia vuhlevodniv nafty drizhdzhamy rodu Candida.

Capability of 14 yeast species to utilize oil hydrocarbons has been analyzed. All strains utilized oil hydrocarbons as a single carbon source. Four strains-destructors that are characterized by higher growth in the presence oil in cultivation medium have been chosen among them. Peroxisomes participation in utilization of oil hydrocarbons by strains-destructors has been shown. Availability of peroxisome key enzymes are characteristic of these strains grown in cultivation medium with oil. Numerous peroxisomes available in the cells of some strains grown in oil cultivation medium have been demonstrated. Utilization of a wide spectrum of oil hydrocarbons has been revealed in all four strains. Two strains are promising to be used for environment purification from oil pollution.

The role of Kupffer cell oxidant production in early ethanol-induced liver disease.

Considerable evidence for a role of Kupffer cells in alcoholic liver disease has accumulated and they have recently been shown to be a predominant source of free radicals. Several approaches including pharmacological agents, knockout mice, and viral gene transfer have been used to fill critical gaps in understanding key mechanisms by which Kupffer cell activation, oxidant formation, and cytokine production lead to liver damage and subsequent pathogenesis. This review highlights new data in support of the hypothesis that Kupffer cells play a pivotal role in hepatotoxicity due to ethanol by producing oxidants via NADPH oxidase.

Alcohol-induced free radicals in mice: direct toxicants or signaling molecules?

Tumor necrosis factor alpha (TNF-alpha) and free radicals are produced in early alcohol-induced liver injury. Recently, pathology caused by alcohol was blocked nearly completely in tumor necrosis factor alpha receptor 1 (TNF-R1) knockout mice. With this model, it is now possible to evaluate whether free radicals are directly toxic or act as redox regulators of TNF-alpha production. Specifically, if free radicals were directly toxic, a parallel decrease in free radicals and pathology in TNF-R1 knockout mice would be predicted. If they only affect TNF-alpha production, radicals would be expected to remain high while pathology is diminished. Accordingly, free radical production in TNF-R1 knockout mice was studied here. The enteral alcohol delivery model used mice lacking TNF-R1 (p55) and wild-type control C57Bl/6J mice. Animals received a liquid diet continuously with either ethanol or isocaloric maltose-dextrin as control for 4 weeks. Urine ethanol levels fluctuated from 10 to 500 mg/dL in a cyclic pattern in mice receiving ethanol. Ethanol elevated liver:body weight ratios, serum alanine transaminase (ALT) levels, and pathology scores in wild-type mice. These parameters were blunted nearly completely in TNF-R1 knockout mice. Ethanol treatment increased free radical production in wild-type mice compared with animals fed a high-fat control diet. There were no differences in intensity of free radical signals regardless of the presence or absence of TNF-R1; however, pathology differed markedly between these groups. These findings are consistent with the hypothesis that free radicals act as redox signals for TNF-alpha production and do not directly damage cells in early alcohol-induced hepatic injury.

PPARalpha-dependent induction of liver microsomal esterification of estradiol and testosterone by a prototypical peroxisome proliferator.

Fatty acyl-coenzyme A:estradiol acyltransferase in liver microsomes catalyzes the formation of estradiol fatty acid esters. These estrogen esters are extremely lipophilic and have prolonged hormonal activity because they are slowly metabolized and slowly release estradiol. Our previous studies showed that treatment of female rats with clofibrate or gemfibrozil (peroxisome proliferators commonly used as hypolipidemic drugs) markedly stimulated the liver microsomal esterification of estradiol. Although clofibrate administration is a potent inducer of liver microsomal fatty acyl-coenzyme A:estradiol acyltransferase in rats, it is a poor inducer in mice. In contrast to these observations, Wy-14,643 (an exceptionally potent prototypical peroxisome proliferator) is a strong inducer of fatty acyl-coenzyme A:estradiol acyltransferase in mice. To explore the role of PPARalpha in the induction of fatty acyl-coenzyme A:estradiol acyltransferase and fatty acyl-coenzyme A:testosterone acyltransferase activities by peroxisome proliferators, we fed 0.1% Wy-14,643 to female wild-type and PPARalpha null mice for 11 d. The liver microsomal acyl-coenzyme A:estradiol acyltransferase and acyl-coenzyme A:testosterone acyltransferase activities were increased 4- to 5-fold in wild-type mice fed Wy-14,643, but no increase was observed in null mice. These results demonstrate that induction of acyl-coenzyme A:estradiol acyltransferase and acyl-coenzyme A:testosterone acyltransferase activities by a prototypical peroxisome proliferator is dependent on PPARalpha.

Development of an animal model of chronic alcohol-induced pancreatitis in the rat.

This study was designed to develop an animal model of alcoholic pancreatitis and to test the hypothesis that the dose of ethanol and the type of dietary fat affect free radical formation and pancreatic pathology. Female Wistar rats were fed liquid diets rich in corn oil (unsaturated fat), with or without a standard or high dose of ethanol, and medium-chain triglycerides (saturated fat) with a high dose of ethanol for 8 wk enterally. The dose of ethanol was increased as tolerance developed, which allowed approximately twice as much alcohol to be delivered in the high-dose group. Serum pancreatic enzymes and histology were normal after 4 wk of diets rich in unsaturated fat, with or without the standard dose of ethanol. In contrast, enzyme levels were elevated significantly by the high ethanol dose. Increases were blunted significantly by dietary saturated fat. Fibrosis and collagen alpha1(I) expression in the pancreas were not detectable after 4 wk of enteral ethanol feeding; however, they were enhanced significantly by the high dose after 8 wk. Furthermore, radical adducts detected by electron spin resonance were minimal with the standard dose; however, the high dose increased carbon-centered radical adducts as well as 4-hydroxynonenal, an index of lipid peroxidation, significantly. Radical adducts were also blunted by approximately 70% by dietary saturated fat. The animal model presented here is the first to demonstrate chronic alcohol-induced pancreatitis in a reproducible manner. The key factors responsible for pathology are the amount of ethanol administered and the type of dietary fat.

ICAM-1 is involved in the mechanism of alcohol-induced liver injury: studies with knockout mice.

To test the hypothesis that leukocyte infiltration mediated by intercellular adhesion molecule (ICAM)-1 is involved in early alcohol-induced liver injury, male wild-type or ICAM-1 knockout mice were fed a high-fat liquid diet with either ethanol or isocaloric maltose-dextrin for 4 wk. There were no differences in mean urine alcohol concentrations between the groups fed ethanol. Alcohol administration significantly increased liver size and serum alanine aminotransferase levels in wild-type mice over high-fat controls, effects that were blunted significantly in ICAM-1 knockout mice. Dietary ethanol caused severe steatosis, mild inflammation, and focal necrosis in livers from wild-type mice. Furthermore, livers from wild-type mice fed ethanol showed significant increases in the number of infiltrating leukocytes, which were predominantly lymphocytes. These pathological changes were blunted significantly in ICAM-1 knockout mice. Tumor necrosis factor (TNF)-alpha mRNA expression was increased in wild-type mice fed ethanol but not in ICAM-1 knockout mice. These data demonstrate that ICAM-1 and infiltrating leukocytes play important roles in early alcohol-induced liver injury, most likely by mechanisms involving TNF-alpha.

Diphenyleneiodonium sulfate, an NADPH oxidase inhibitor, prevents early alcohol-induced liver injury in the rat.

The oxidant source in alcohol-induced liver disease remains unclear. NADPH oxidase (mainly in liver Kupffer cells and infiltrating neutrophils) could be a potential free radical source. We aimed to determine if NADPH oxidase inhibitor diphenyleneiodonium sulfate (DPI) affects nuclear factor-kappaB (NF-kappaB) activation, liver tumor necrosis factor-alpha (TNF-alpha) mRNA expression, and early alcohol-induced liver injury in rats. Male Wistar rats were fed high-fat liquid diets with or without ethanol (10-16 g. kg(-1). day(-1)) continuously for up to 4 wk, using the Tsukamoto-French intragastric enteral feeding protocol. DPI or saline vehicle was administered by subcutaneous injection for 4 wk. Mean urine ethanol concentrations were similar between the ethanol- and ethanol plus DPI-treated groups. Enteral ethanol feeding caused severe fat accumulation, mild inflammation, and necrosis in the liver (pathology score, 4.3 +/- 0.3). In contrast, DPI significantly blunted these changes (pathology score, 0.8 +/- 0.4). Enteral ethanol administration for 4 wk also significantly increased free radical adduct formation, NF-kappaB activity, and TNF-alpha expression in the liver. DPI almost completely blunted these parameters. These results indicate that DPI prevents early alcohol-induced liver injury, most likely by inhibiting free radical formation via NADPH oxidase, thereby preventing NF-kappaB activation and TNF-alpha mRNA expression in the liver.

Adenoviral gene delivery can inactivate Kupffer cells: role of oxidants in NF-kappaB activation and cytokine production.

Kupffer cells play a significant role in the pathogenesis of several liver diseases; therefore, a potential therapeutic strategy would be to inactivate the Kupffer cell with a gene-delivery system. Although recombinant adenovirus provides robust, transgene expression in parenchymal cells, whether adenovirus transduces Kupffer cells is unclear. Thus, the purpose of this study was to evaluate this possibility. In animals infected with adenovirus, Kupffer cells were identified positively to express adenoviral transgenes by immunohistochemical techniques and Western blot analysis, indicating that Kupffer cells are transduced in vivo. Indeed, isolated Kupffer cells were transduced in vitro with recombinant adenovirus in a dose-dependent manner. Moreover, adenoviral transduction of Kupffer cells was blocked by inhibitors of alphaVbeta5 integrin, the co-receptor for adenovirus binding, supporting the hypothesis that adenovirus transduces Kupffer cells via an alphaVbeta5 integrin-dependent mechanism. Indeed, it is shown here that Kupffer cells express alphaVbeta5 integrins. In a functional assay, infection of isolated Kupffer cells with adenovirus containing superoxide dismutase or IkappaB alpha super-repressor blunted LPS-induced nuclear transcription factor kappa B (NF-kappaB) activation and tumor necrosis factor alpha (TNF-alpha) production but not IL-10 production. Moreover, superoxide production was blocked by expression of superoxide dismutase. These data support the hypothesis that LPS-induced NF-kappaB activation and TNF-alpha production in Kupffer cells are oxidant-dependent. These findings suggest that Kupffer cell-targeted approaches may be a potential therapeutic strategy against many inflammatory diseases including early alcohol-induced liver injury.

Phthalates rapidly increase production of reactive oxygen species in vivo: role of Kupffer cells.

The role of oxidants in the mechanism of tumor promotion by peroxisome proliferators remains controversial. The idea that induction of acyl-coenzyme A oxidase leads to increased production of H(2)O(2), which damages DNA, seems unlikely; still, free radicals might be important in signaling in specialized cell types such as Kupffer cells, which produce mitogens. Because hard evidence for increased oxidant production in vivo after treatment with peroxisome proliferators is lacking, the spin-trapping technique and electron spin resonance spectroscopy were used. Rats were given di(2-ethylhexyl) phthalate (DEHP) acutely. The spin trapping agent alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone was also given and bile samples were collected for 4 h. Under these conditions, the intensity of the six-line radical adduct signal increased to a maximum value of 2.5-fold 2 h after administration of DEHP, before peroxisomal oxidases were induced. Furthermore, DEHP given with [(13)C(2)]dimethyl sulfoxide produced a 12-line electron spin resonance spectrum, providing evidence that DEHP stimulates (*)OH radical formation in vivo. Furthermore, when rats were pretreated with dietary glycine, which inactivates Kupffer cells, DEHP did not increase radical signals. Moreover, similar treatments were performed in knockout mice deficient in NADPH oxidase (p47(phox) subunit). Importantly, DEHP increased oxidant production in wild-type but not in NADPH oxidase-deficient mice. These data provide evidence for the hypothesis that the molecular source of free radicals induced by peroxisome proliferators is NADPH oxidase in Kupffer cells. On the contrary, radical adduct formation was not affected in peroxisome proliferator-activated receptor alpha knockout mice. These observations represent the first direct, in vivo evidence that phthalates increase free radicals in liver before peroxisomal oxidases are induced.

Delivery of the Cu/Zn-superoxide dismutase gene with adenovirus reduces early alcohol-induced liver injury in rats.

BACKGROUND AND AIMS: Alcohol-induced liver injury is associated with an increase in oxidants from a variety of possible sources. Therefore, it was hypothesized that increased and stable expression of the antioxidant enzyme Cu/Zn-superoxide dismutase (SOD1) would diminish oxygen free radicals and reduce alcohol-induced liver injury. METHODS: To test this hypothesis, rats were given recombinant adenovirus containing Cu/Zn-superoxide dismutase (Ad.SOD1) or beta-galactosidase (Ad.lacZ) and fed ethanol enterally for 3 weeks. RESULTS: SOD was increased significantly 3-5-fold over endogenous levels in both hepatocytes as well as Kupffer cells 3 weeks after infection. Serum transaminase levels and pathology were elevated significantly in Ad.lacZ-treated animals by using an intragastric feeding model. This effect was blunted significantly in Ad.SOD1-infected animals. Importantly, electron spin resonance-detectable free-radical adducts caused by ethanol were also decreased by SOD1 overexpression. Moreover, the increase in nuclear factor kappaB (NFkappaB), tumor necrosis factor alpha (TNF-alpha), and interleukin 1 messenger RNA (mRNA) caused by ethanol was blunted in animals treated with Ad.SOD1. CONCLUSIONS: These data support the hypothesis that oxidant production is critical in early alcohol-induced liver injury and that gene delivery of antioxidant enzymes may be useful in prevention and treatment.

Ebselen prevents early alcohol-induced liver injury in rats.

Oxidants have been shown to be involved in alcohol-induced liver injury. Moreover, 2-phenyl-1,2-benzisoselenazole-3(2H)-one (ebselen), an organoselenium compound and glutathione peroxidase mimic, decreases oxidative stress and protects against stroke clinically. This study was designed to test the hypothesis that ebselen protects against early alcohol-induced liver injury in rats. Male Wistar rats were fed high-fat liquid diets with or without ethanol (10-16 g/kg/d) continuously for up to 4 weeks using the intragastric enteral feeding protocol developed by Tsukamoto and French. Ebselen (50 mg/kg twice daily, intragastrically) or vehicle (1% tylose) was administered throughout the experiment. Mean urine ethanol concentrations were not significantly different between treatment groups, and ebselen did not affect body weight gains or cyclic patterns of ethanol concentrations in urine. After 4 weeks, serum ALT levels were increased significantly about 4-fold over control values (37 +/- 5 IU/l) by enteral ethanol (112 +/- 7 IU/l); ebselen blunted this increase significantly (61 +/- 8 IU/l). Enteral ethanol also caused severe fatty accumulation, mild inflammation, and necrosis in the liver (pathology score: 4.3 +/- 0.3). In contrast, these pathological changes were blunted significantly by ebselen (pathology score: 2.5 +/- 0.4). While there were no significant effects of either ethanol or ebselen on glutathione peroxidase activity in serum or liver tissue, ebselen blocked the increase in serum nitrate/nitrite caused by ethanol. Furthermore, ethanol increased the activity of NF-kappaB over 5-fold, the number of infiltrating neutrophils 4-fold, and the accumulation of 4-hydroxynonenal over 5-fold. Ebselen blunted all of these effects significantly. These results indicate that ebselen prevents early alcohol-induced liver injury, most likely by preventing oxidative stress, which decreases inflammation.

Endothelial cells contain a glycine-gated chloride channel.

Glycine inhibited growth of B16 melanoma tumors in vivo most likely because of the inhibition of angiogenesis. Here, the hypothesis that the anticancer effect of glycine in vivo is due to expression of a glycine-gated Cl- channel in endothelial cells was tested. First, the effects of glycine on vascular endothelial growth factor-induced increases in intracellular Ca2+ concentration in a bovine endothelial (CPA) cell line were studied. Vascular endothelial growth factor (1 ng/ml) increased intracellular Ca2+ concentration, with peak values reaching 141 +/- 11 nM. Glycine blunted this increase dose dependently. Furthermore, the inhibitory effects of glycine were prevented by 1 microM strychnine, a glycine receptor antagonist, or when cells were incubated in Cl(-)-free buffer. Moreover, glycine increased influx of 36Cl into CPA cells approximately 10-fold; this reaction was also strychnine sensitive. Furthermore, mRNA similar to the beta-subunit of the glycine-gated Cl- channel from spinal cord was identified in endothelial cells by reverse transcription-polymerase chain reaction. In addition, Western analysis using antibody for the glycine receptor demonstrated expression of the beta-subunit of the glycine receptor. Importantly, glycine diminished serum-stimulated proliferation and migration of endothelial cells. Collectively, these data indicate that the inhibitory effect of glycine on growth and migration of endothelial cells is due to activation of a glycine-gated Cl- channel. This hyperpolarizes the cell membrane and blocks influx of Ca2+, thereby minimizing growth factor-mediated signaling.

Tolerance and sensitization to endotoxin in Kupffer cells caused by acute ethanol involve interleukin-1 receptor-associated kinase.

Ethanol changes sensitivity of Kupffer cells to endotoxin. Here, the hypothesis that interleukin-1 receptor-associated kinase (IRAK), a downstream signaling molecule of toll-like receptors, regulates the response to LPS in Kupffer cells after ethanol treatment was evaluated. C57BL/6 mice were given ethanol intragastrically, and LPS was injected 1 or 21 h later. One hour after ethanol treatment, serum transaminases after LPS were 60% of control, while ethanol increased these parameters about 3-fold 21 h after ethanol. Pretreatment with antibiotics blocked these effects of ethanol. In Kupffer cells from mice treated with ethanol 1 h earlier, LPS-induced TNFalpha production, and IRAK expression and activity and NFkappaB were decreased 50-60% of control. In contrast, in Kupffer cells from mice treated with ethanol 21 h earlier, LPS-induced TNFalpha production, expression and activity of IRAK were increased 1.5-fold over controls, while NFkappaB was elevated 3-fold. These data indicate that ethanol-induced tolerance and sensitization of Kupffer cells to endotoxin in mice involve IRAK.

NADPH oxidase-derived free radicals are key oxidants in alcohol-induced liver disease.

In North America, liver disease due to alcohol consumption is an important cause of death in adults, although its pathogenesis remains obscure. Despite the fact that resident hepatic macrophages are known to contribute to early alcohol-induced liver injury via oxidative stress, the exact source of free radicals has remained a mystery. To test the hypothesis that NADPH oxidase is the major source of oxidants due to ethanol, we used p47(phox) knockout mice, which lack a critical subunit of this major source of reactive oxygen species in activated phagocytes. Mice were treated with ethanol chronically, using a Tsukamoto-French protocol, for 4 weeks. In wild-type mice, ethanol caused severe liver injury via a mechanism involving gut-derived endotoxin, CD14 receptor, production of electron spin resonance-detectable free radicals, activation of the transcription factor NF-kappaB, and release of cytotoxic TNF-alpha from activated Kupffer cells. In NADPH oxidase-deficient mice, neither an increase in free radical production, activation of NF-kappaB, an increase in TNF-alpha mRNA, nor liver pathology was observed. These data strongly support the hypothesis that free radicals from NADPH oxidase in hepatic Kupffer cells play a predominant role in the pathogenesis of early alcohol-induced hepatitis by activating NF-kappaB, which activates production of cytotoxic TNF-alpha.

Chronic ethanol increases adeno-associated viral transgene expression in rat liver via oxidant and NFkappaB-dependent mechanisms.

Recombinant adeno-associated virus (rAAV) transduction is limited in vivo, yet can be enhanced by hydroxyurea, ultraviolet-irradiation, or adenovirus coinfection, possibly via mechanisms involving stress in the host cell. Because chronic ethanol induces oxidative stress, it was hypothesized that chronic ethanol would increase rAAV transduction in vivo. To test this hypothesis, rAAV encoding beta-galactosidase was given to Wistar rats that later received either ethanol diet or high-fat control diet via an enteral-feeding protocol for 3 weeks. Expression and activity of beta-galactosidase in the liver were increased nearly 5-fold by ethanol. The increase in transgene expression was inhibited by antioxidant diphenylene iodonium (DPI), which is consistent with the hypothesis that ethanol causes an increase in rAAV transduction via oxidative stress. Ethanol increased DNA synthesis only slightly; however, it increased the nuclear transcription factor kappaB (NFkappaB) 4-fold, a phenomenon also sensitive to DPI. Moreover, a 6-fold increase in rAAV transgene expression was observed in an acute ischemia-reperfusion model of oxidative stress. Transgene expression was transiently increased 24 hours after ischemia-reperfusion 3 days and 3 weeks after rAAV infection. Further, adenoviral expression of superoxide dismutase or IkappaBalpha superrepressor inhibited rAAV transgene expression caused by ischemia-reperfusion. Therefore, it is concluded that ethanol increases rAAV transgene expression via mechanisms dependent on oxidative stress, and NFkappaB likely through enhancement of cytomegaloviral (CMV) promoter elements. Alcoholic liver disease is an attractive target for gene therapy because consumption of ethanol could theoretically increase expression of therapeutic genes (e.g., superoxide dismutase). Moreover, this study has important implications for rAAV gene therapy and potential enhancement and regulation of transgene expression in liver.

Oxidants from nicotinamide adenine dinucleotide phosphate oxidase are involved in triggering cell proliferation in the liver due to peroxisome proliferators.

It was shown that 4-chloro-6-(2,3-xylidino)-2-pyrimidinylthio acetic acid (Wy-14,643), a potent peroxisome proliferator, caused rapid oxidant-dependent activation of nuclear factor kappaB (NF-kappaB) in Kupffer cells in vivo and activated superoxide production by isolated Kupffer cells. Here, we tested the hypothesis that NADPH oxidase (NADPH OX) is the source of oxidants increased by Wy-14,643. Indeed, both activation of NF-kappaB and increases in cell proliferation due to a single dose of Wy-14,643 (100 mg/kg) were prevented completely when rats were pretreated with diphenyleneiodonium (1 mg/kg), an inhibitor of NADPH OX. p47phox is a critical subunit of NADPH OX; therefore, p47phox knockout mice were used to specifically address the hypothesis of NADPH OX involvement. In livers of wild-type mice, Wy-14,643 activated NF-kappaB, followed by an increase in mRNA for tumor necrosis factor a. Importantly, these changes did not occur in p47phox knockouts. Moreover, when Kupffer cells were treated with Wy-14,643 in vitro, superoxide production was increased in cells from wild-type but not p47phox-null mice. Finally, when mice were fed a Wy-14,643-containing (0.1%) diet for 7 days, the increase in liver weight and cell proliferation caused by Wy-14,643 in wild-type mice was blocked in p47phox-null mice. Combined, these results are consistent with the hypothesis that Wy-14,643 activates NADPH OX, which leads to NF-kappaB-mediated production of mitogens that causes hepatocellular proliferation characteristic of this class of nongenotoxic carcinogens.

Peroxisome proliferator-activated receptor alpha is restricted to hepatic parenchymal cells, not Kupffer cells: implications for the mechanism of action of peroxisome proliferators in hepatocarcinogenesis.

Peroxisome proliferators increase hepatocyte proliferation and cause liver tumors in rodents, yet the mechanism of action is not understood. Based on studies with null mice it is known that peroxisome proliferator-activated receptor-alpha (PPARalpha) is involved. There is also evidence that Kupffer cells play a central role in peroxisome proliferator-induced carcinogenesis, most likely via mechanisms involving increases in superoxide, activation of nuclear factor kappaB and production of tumor necrosis factor-alpha (TNFalpha). However, it is not known whether PPARalpha is constitutively expressed in Kupffer cells. Therefore, the expression of PPAR isoforms in rat Kupffer and parenchymal cells was examined. Kupffer cells and hepatocytes of >99% purity were isolated from rats fed either a control diet or one containing 0.1% WY-14,643 for 1 week. Protein and RNA were obtained and PPAR expression was analyzed using northern and western blots. PPARalpha, PPARbeta and PPARgamma mRNA was detected in purified hepatocytes. In Kupffer cells, mRNA encoding PPARgamma was present while transcripts for PPARalpha and PPARbeta were not detected. Immunoblots were consistent with the results found by northern analysis. Moreover, when Kupffer cells from wild-type or PPARalpha-null mice were treated with WY-14,643 in vitro, superoxide production was similar. Combined, these results show that PPARalpha is expressed in rat parenchymal cells but not in Kupffer cells. These data are consistent with the hypothesis that parenchymal cells respond to Kupffer cell-derived TNFalpha via mechanisms dependent on PPARalpha within the parenchymal cells.