Department of Pharmacology and Toxicology, National Food Safety and Toxicology Center and Institute for Environmental Toxicology, Michigan State University, East Lansing, Michigan 48824
Received July 30, 2002; accepted October 10, 2002
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key Words: liver; endotoxin; monocrotaline; Kupffer cell; TNF-; inflammation; gadolinium chloride; pentoxifylline; COX-2; NS-398.
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Bacterial lipopolysaccharide (LPS), a constituent of the outer cell wall of Gram-negative bacteria, is a potent inflammagen (Holst et al., 1996). In rats, liver injury from a large, toxic dose of LPS is characterized by neutrophil infiltration with hepatocellular degeneration and coagulative necrosis in midzonal regions of liver lobules (Hewett et al., 1992
; Yee et al., 2000
). The mechanism for LPS-induced liver injury is complex and involves the interaction of numerous inflammatory cells and soluble mediators, including Kupffer cells (KCs) and tumor necrosis factor-
(TNF-
) (Brouwer et al., 1995
; Brown et al., 1997
; Ganey and Roth, 2001
; Hewett et al., 1992
, 1993
).
Exposure to a smaller LPS dose results in a modest, noninjurious inflammatory response (Ganey and Roth, 2001; Michie, 1988
; Spitzer and Mayer, 1993
). It has been hypothesized that the release of inflammatory mediators has the potential to alter tissue homeostasis and increase the susceptibility of tissues to chemical-induced injury (Ganey and Roth, 2001
). Yee et al. (2000)
recently demonstrated that synergistic liver injury resulted when a noninjurious dose of LPS was administered 4 h after a small, nonhepatotoxic dose of MCT. The resulting liver lesions were both centrilobular and midzonal, exhibiting characteristics similar to lesions associated with larger, toxic doses of MCT or LPS, respectively. Failure to reproduce synergistic injury in isolated hepatic parenchymal cells (HPCs) in vitro suggested that the enhanced toxicity resulted not from direct interaction of MCT and LPS with HPCs, but rather from an indirect mechanism (Yee et al., 2000
). Inasmuch as LPS is an inflammagen, the present study was designed to identify inflammatory factors critical to the synergistic hepatotoxicity in this model.
KCs are the resident macrophages of the liver. When activated by LPS, they produce and release numerous mediators, including cytokines (TNF-, interleukin-1, and interleukin-6) and reactive oxygen species (Holst et al., 1996
; Michie et al., 1988
). Cyclooxygenase-2 (COX-2) is also induced, leading to the enhanced synthesis of prostaglandins (PGs) that have proinflammatory actions (Brouwer et al., 1995
; Dieter et al., 1999
). Many of these mediators further activate or modulate the effects of nearby cells involved in the inflammatory process and can contribute to the development of injury (Hewett et al., 1993
; Holst et al., 1996
). Inactivation of KCs with gadolinium chloride (GdCl3) in rats given a large, hepatotoxic dose of LPS significantly attenuates hepatocellular necrosis, suggesting that KCs are critical to the pathogenesis of LPS-induced injury (Brown et al., 1997
).
TNF- is a potent inflammatory cytokine produced by activated macrophages and to a lesser degree by other cell types (Bradham et al., 1998
; Vassalli, 1992
). This cytokine can exert a variety of effects on cells ranging from mitochondrial damage and oncotic or apoptotic necrosis to cell proliferation (Bradham et al., 1998
). TNF-
may also prompt the accumulation of neutrophils (PMNs) by activating endothelial cells (Bradham et al., 1998
; Vassalli, 1992
). It can indirectly promote toxicity by priming PMNs to release reactive oxygen and nitrogen species and proteases that damage nearby cells (Nagaki et al., 1991
; Vasselli, 1992). Inhibition of TNF-
synthesis or activity attenuates injury caused by a hepatotoxic dose of LPS, indicating that TNF-
is a critical factor in the pathogenesis (Hewett et al., 1993
).
The purpose of this study was to investigate the role of inflammatory factors in the synergistic liver injury that occurs following coexposure to small doses of MCT and LPS. Specifically, the hypothesis that KCs and TNF- play causal roles in the hepatotoxicity was tested. In addition, the importance of COX-2 products for the development of injury was evaluated.
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Animals.
Male Sprague-Dawley rats (Crl:CD (SD)IGS BR, Charles River, Portage, MI) weighing 200300 g were used in all studies. Animals were allowed food (Rodent Chow/Tek 8640, Harlan Teklad, Madison, WI) and water ad libitum. They were housed no more than three to a cage on Aspen chip bedding (Northeastern Products Company, Warrenburg, NY) and were maintained on a 12-h light/dark cycle in a controlled temperature (1821°C) and humidity (55 ± 5%) environment for a period of 1 week before use. All procedures on animals followed the guidelines for humane treatment set by the American Association of Laboratory Animal Sciences and the University Laboratory Animal Research Unit at Michigan State University.
Treatment protocol.
MCT was dissolved in sterile saline minimally acidified by 0.2 M HCl. The pH was brought to 7 by addition of 2 M NaOH, and the volume adjusted with sterile saline to the appropriate final concentration. Rats were given MCT (100 mg/kg) or an equivalent volume of sterile saline, intraperitoneally, followed 4 h later by LPS (7.4 x 106 EU/kg) or its saline vehicle via tail vein injection. LPS was administered 4 h after MCT to minimize interference with MCT bioactivation (Allen et al., 1972). As previously reported, these doses of MCT and LPS resulted in minimal to no injury by themselves. However, when they were administered together, synergistic liver injury resulted, with maximal injury occurring by 18 h (Yee et al., 2000
).
At the times indicated in figure and table legends, rats were anesthetized with sodium pentobarbital (50 mg/kg, ip). A midline abdominal incision was made, blood was collected from the inferior vena cava into a syringe containing sodium citrate (0.38% final concentration), and animals were euthanized by exsanguination. Livers were removed intact and fixed in 10% neutral buffered formalin for at least 3 days before being processed for histologic analysis.
Treatment with gadolinium chloride.
Rats were treated with 10 mg GdCl3-6H2O/kg (pH 3.5) or saline vehicle intravenously 24 h before the administration of LPS or its vehicle. This treatment protocol has been shown to inactivate KC function (Brown et al., 1997).
Treatment with pentoxifylline.
Rats received 100 mg PTX/kg or its saline vehicle intravenously 1 h before LPS treatment. This treatment regimen prevents the LPS-induced increase in plasma TNF- activity (Barton et al., 2001
).
Treatment with anti-TNF- serum.
Rats were treated intravenously with rabbit ATS (1 ml) or control rabbit serum (CS; 1 ml) 1 h before LPS administration. This treatment protocol has been shown to prevent LPS-induced increase in plasma TNF- activity (Barton et al., 2001
; Hewett et al., 1993
).
Treatment with NS-398.
NS-398 is a selective inhibitor of COX-2 (Futaki et al., 1994, 1997
). Rats were given 5 mg NS-398/kg or olive oil intraperitoneally 1 h before LPS administration.
Assessment of liver injury, plasma TNF-, and plasma hyaluronic acid.
HPC injury was estimated by increases in the activities of plasma ALT and AST. An ELISA was used to measure plasma hyaluronic acid (HA), a marker of hepatic SEC injury (Copple et al., 2002; Deaciuc et al., 1993
, 1994
). Plasma TNF-
concentration was determined via a rat TNF-
ELISA kit.
Histopathologic evaluation and morphometry.
Serial transverse sections from the left lateral liver lobe were processed for light microscopy. Paraffin-embedded sections were cut at 4 µm, stained with hematoxylin and eosin, and evaluated for lesion size and severity. Tissue sections were analyzed histologically without knowledge of the treatment group.
Digitized color images of hematoxylin- and eosin-stained liver sections were visualized with an Olympus AX-80T light microscope (Olympus Corp., Lake Success, NY) interfaced with a high-resolution CCD color camera (OLY-750, Olympus-America, Inc., Melville, NY) to quantify treatment-induced changes in liver morphology. Images were evaluated with Scion Image software (Scion Corporation, Frederick, MD) employing a 64-point lattice grid to determine (1) the total area of liver analyzed, (2) the area of centrilobular lesion, (3) the area of midzonal lesion, (4) the area of normal parenchyma, and (5) the area of nonparenchymal space. A lesion was defined as hepatic parenchymal cells with either swollen, eosinophilic cytoplasm and karyolytic or pyknotic nuclei (i.e., oncosis), or cells with shrunken cytoplasm and karyorrhexic nuclei or apoptotic bodies (i.e., apoptosis; Levin et al., 1999; Majno and Joris, 1995
). Nonparenchymal space was defined as nonparenchymal tissue, vessel lumen, and regions outside the perimeter of the liver section. The area of each object (category) of interest (i.e., lesion) was calculated from the following expression (Cruz-Orive, 1982
):
![]() |
![]() |
Distance between points was 55 µm. Accordingly, the area represented by each point was 3025 µm2. One section from the liver of each animal in a treatment group was systematically scanned using adjacent, nonoverlapping microscopic fields. The first image field analyzed in each section was chosen using a random number table (i.e., any image field between 1 and 10). Thereafter, every 10th field containing hepatic parenchymal cells was evaluated (minimum of 20 fields measured/section). The measured fields represented approximately 10% of the total area of each liver section. Eight animals per group were analyzed. Percentage lesion area was estimated based on the following formula:
![]() |
Statistical analysis.
Results are expressed as mean ± SEM. Data expressed as percentages were transformed by arc sine square root prior to analysis. Data for single comparisons were analyzed by Student's t-test or, when appropriate, Fisher's exact test (Steele et al., 1997). Homogeneous data were analyzed by one- or two-way analysis of variance (ANOVA), as appropriate, and group means were compared using Student-Newman-Keuls post hoc test (Steele et al., 1997
). When variances were not homogeneous, data were analyzed using the Kruskal-Wallis nonparametric ANOVA, and Dunn's Multiple Comparison test was used to compare group means (Steele et al., 1997
). The criterion for significance was p
0.05 for all comparisons.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
MCT/LPS-cotreated control animals exhibited centrilobular and midzonal liver lesions as previously described by Yee et al. (2000). Centrilobular lesions consisted of moderate to marked hepatocellular apoptotic and oncotic necrosis, degeneration, hemorrhage, and vascular injury. Midzonal lesions in the MCT/LPS-cotreated group comprised marked but more frequent and well-defined areas of hepatocellular coagulative necrosis accompanied by neutrophil infiltration with congestion and hemorrhage. Liver lesions from animals treated with GdCl3/MCT/LPS exhibited qualitatively similar centrilobular and midzonal lesions; however, these lesions were both smaller in size and less frequent. The GdCl3-induced reduction in lesion size and frequency was more pronounced in the midzonal regions. Morphometric analysis of liver lesions supported the observation that GdCl3 pretreatment reduced the liver area affected by centrilobular and midzonal lesions (Table 1
). Furthermore, the number of hepatic PMNs was smaller in MCT/LPS-treated animals that received GdCl3 pretreatment compared to MCT/LPS-treated animals that did not. Histologically, no evidence of liver injury was observed in animals treated with vehicles or GdCl3 alone.
|
|
|
Histological examination of the livers from animals that received the MCT/PTX/LPS treatment revealed lesions qualitatively similar to those observed in animals that received MCT/LPS alone, but they were smaller in size and less frequent. Midzonal lesions exhibited the greatest reduction in size and frequency and demonstrated less congestion and hemorrhage. These observations were consistent with the mophometric analysis of lesions in this study (Table 1). Hepatic PMN accumulation was also reduced by PTX treatment. No lesions were observed in livers from animals that were given vehicles or PTX alone.
Effect of Anti-TNF- Serum on Liver Injury from MCT/LPS Cotreatment
MCT/LPS-cotreated animals were given ATS before LPS treatment to confirm the causal relationship between TNF- and MCT/LPS-induced liver injury. ATS given to MCT/LPS-treated animals prevented the increase in plasma TNF-
concentration at 18 h (Fig. 3A
).
|
Animals treated with MCT/ATS/LPS had liver lesions qualitatively similar to those in rats given MCT/CS/LPS, although the lesions were reduced in size and frequency. Midzonal liver lesions from MCT/ATS/LPS-treated animals exhibited less congestion and hemorrhage than livers from MCT/CS/LPS animals. Morphometric analysis of liver lesions supports the observation that ATS treatment lessened the centrilobular, MCT-like and midzonal, LPS-like lesions (Table 1). Furthermore, hepatic PMN accumulation was reduced in MCT/ATS/LPS-treated animals compared to animals given MCT/CS/LPS. No injury was observed histologically in the livers of animals that received treatment with vehicles or ATS alone.
Effect of NS-398 on MCT/LPS-Induced Liver Injury
Among the various mediators released by LPS-activated KCs are metabolites of arachidonic acid, including PGE2 and PGD2 (Decker, 1990; Ganey et al., 2001
). These PGs are produced via the inflammation-inducible COX-2 enzyme and have a variety of effects on other inflammatory cells and HPCs that can contribute to pathophysiological alterations in tissues (Casteleijn et al., 1988
; Decker, 1990
; Ganey et al., 2001
).
NS-398, a selective COX-2 inhibitor, was administered one hour before LPS at a dose that reduces PG production in vivo (Ganey et al., 2001). NS-398 did not attenuate plasma ALT (Fig. 4A
) or AST (Fig. 4B
) activity in MCT/LPS-treated animals. Plasma ALT or AST activity was unaffected in animals receiving treatment with NS-398 alone. Histologically, liver lesions appeared the same in the MCT/LPS-cotreated animals given NS-398 or its vehicle. No liver lesions were observed in control animals that received vehicles or NS-398 alone.
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
In this model of liver injury, plasma TNF- concentration is elevated before the onset of HPC injury, raising the possibility that this cytokine might have a causal role in injury development (Table 2
). Either inhibition of TNF-
synthesis by PTX (Figs. 2B and 2C
) or neutralization of TNF-
activity by ATS (Figs. 3B and 3C
) resulted in the attenuation of HPC injury. Both PTX and ATS decreased plasma TNF-
concentration in MCT/LPS cotreated animals, confirming that the agents were effective in reducing TNF-
(Figs. 2A and 3A
). These results suggest that TNF-
is causally involved in the hepatocellular injury.
In rats treated with a small, noninjurious dose of LPS 4 h after a small, nontoxic dose of MCT, morphologic evidence of liver lesions develop within 1218 h after MCT administration, consistent with elevated plasma ALT and AST activities (Yee et al., 2000). Both centrilobular and midzonal liver lesions result and are qualitatively similar to lesions that characterize larger, toxic doses of MCT and LPS, respectively. Centrilobular liver lesions comprise moderate to marked hepatocellular apoptotic and oncotic necrosis, degeneration, congestion, hemorrhage and vascular injury. Midzonal lesions consist of frequent, well-defined areas of marked hepatocellular coagulative necrosis, accompanied by neutrophil infiltration, congestion and hemorrhage (Yee et al., 2000
). SEC injury is apparent in both lesions (Yee et al., 2002
). The observation that both MCT-like and LPS-like lesions develop suggests that each agent enhances the effect of the other in this model (Yee et al., 2000
).
Administration of GdCl3, PTX or ATS to MCT/LPS-treated animals reduced the size and frequency of liver lesions. Morphometric analysis of these lesions demonstrated that Kupffer cell inactivation or TNF- depletion in MCT/LPS-cotreated animals significantly reduced the area of the centrilobular, MCT-like and midzonal, LPS-like lesions (Table 1
). This suggests that pharmacologic manipulation ameliorated the interactive effect of these agents.
In this model of synergistic liver injury, LPS caused an expected, early rise in plasma TNF- concentration. As seen previously (Barton et al., 2001
), this increase was transient in rats treated with LPS alone. Unexpectedly, however, plasma TNF-
concentration remained significantly elevated for at least 12 h in rats treated with MCT/LPS (Table 2
). This prolonged elevation contrasts with another model of LPS potentiation of hepatotoxicity in which the LPS-induced increase was not prolonged by aflatoxin B1 cotreatment (Barton et al., 2001
). Even at a larger, hepatotoxic dose of LPS, the elevation in plasma TNF-
concentration remains transient (Iimuro et al., 1994
; Pearson et al., 1996
). It is noteworthy that MCT given by itself to rats, either at the nontoxic dose used in this study (Table 2
) or at a larger, hepatotoxic dose (300 mg/kg; unpublished observation), does not increase plasma TNF-
concentration. Thus, the interaction of these two agents is required for this prolonged effect in MCT/LPS-treated rats.
Under normal conditions, approximately 90% of HA circulating in the blood is removed and degraded by SECs in the liver (Kobayashi et al., 1999). SEC injury impairs the ability of these cells to clear HA from the circulation, and this results in increased plasma HA concentration. Such increases have been used as a biomarker of SEC injury in vivo after exposure of animals to toxicants. An increase in plasma HA concentration correlates with histopathologic evidence of SEC injury and destruction in rats given a hepatotoxic dose of either MCT (Copple et al., 2002
) or LPS (Deaciuc et al., 1994
; Spapen et al., 1999
). Similarly, MCT/LPS coexposure results in increased plasma HA concentration in rats, and this correlates with SEC injury (Yee et al., 2002
). The elevation in plasma HA concentration was slightly but significantly smaller after coadministration of GdCl3 or ATS to MCT/LPS-treated animals (Figs. 1D and 3D
). Administration of PTX to MCT/LPS-treated animals resulted in a trend toward a decrease that was not statistically significant (Fig. 2D
). These results suggest that KC inactivation or TNF-
neutralization in MCT/LPS-treated animals significantly but incompletely attenuated SEC injury.
The attenuation of HPC and SEC injury in this model was likely not the result of GdCl3, PTX, or ATS administration interfering with MCT bioactivation. In animals given a large, hepatotoxic dose of MCT (300 mg/kg), neither GdCl3 nor PTX pretreatment of rats altered liver injury (unpublished observation), suggesting that these agents do not interfere with MCT bioactivation. Badger et al. (1997) demonstrated that GdCl3 pretreatment of rats caused a modest decrease in hepatic cytochrome P450 (CYP). However, this reduction could be explained by a decrease in the CYP 2E1 isoform (Badger et al., 1997
), and since MCT is bioactivated by a different isoform (i.e., CYP 3A family; Kasahara et al., 1997
), it is unlikely that GdCl3 reduced injury in this model by decreasing CYP concentration. In the present study, PTX and ATS were given to MCT/LPS-cotreated animals at a time when most of the administered MCT has been metabolized (Allen et al., 1972
). Neither GdCl3 (Mimura et al., 1995
; Vollmar et al., 1996
) nor PTX (Heller et al., 1999
) interfere with LPS clearance. Accordingly, it is unlikely that the pharmacological agents used in this study reduced injury by interfering with MCT bioactivation or affecting LPS metabolism.
Rose et al. (2001) demonstrated that GdCl3 caused a transient activation of NF
B and enhanced hepatocellular proliferation in the liver. However, NF
B activation returned to baseline within 24 h after GdCl3 administration; therefore, it is unlikely that NF
B activation influenced the results in this study, since MCT/LPS-cotreatment commenced at this time. The possibility that GdCl3 had effects independent of its ability to inactivate Kupffer cells cannot be excluded; however, a similar degree of hepatotoxicity from a large, toxic dose of MCT (300 mg/kg) was unaffected by GdCl3 (unpublished observation), suggesting that it is unlikely to protect through nonselective modes of action such as enhancing cell proliferation.
TNF- can promote liver injury in a number of ways. For example, in vitro TNF-
renders hepatocytes more susceptible to toxicity (Adamson and Billings, 1992
; El-Sisi et al., 1993
; Hoek and Pastorino, 2002
). Likewise, HPCs altered homeostatically by the actions of hepatoxicants may be sensitive to TNF-
-induced cell killing (Jaeschke et al., 1998
; Lawson et al., 1998
). In addition, TNF-
can prime PMNs to release toxic products (i.e., ROS and proteases) that can damage nearby cells (Kushimoto et al., 1996
; Nagaki et al., 1991
; Vassalli, 1992
). Further study will be required to understand how TNF-
acts to promote hepatotoxicity in this model.
In addition to TNF-, other inflammatory mediators are released by KCs that may have deleterious effects on liver (Holst et al., 1996
). For example, COX-2 products mediate liver injury in other models. NS-398, a selective COX-2 inhibitor, was administered to rats using a treatment protocol that inhibits COX-2 in vivo and affords protection from liver injury in a PG-dependent model (Ganey et al., 2001
). The increases in plasma ALT and AST activities in MCT/LPS-cotreated animals were not attenuated by NS-398, which suggests that COX-2 products are not needed for HPC injury in this model (Figs. 4A and 4B
). A similar result was observed in a model of LPS-potentiated aflatoxin B1 hepatotoxicity (Barton et al., 2001
); in contrast, a significant protective effect occurred in rats treated with LPS and allyl alcohol (Ganey et al., 2001
). These contrasting results suggest that the critical mediators of LPS-potentiated hepatotoxic responses differ with different hepatotoxicants.
In summary, GdCl3 administered to MCT/LPS-treated rats at a dose that inhibits KC function reduced HPC and SEC injury. Moreover, the administration of TNF- depleting agents to MCT/LPS-cotreated rats also protected against HPC injury and caused a modest attenuation of SEC injury. Accordingly, KCs and TNF-
appear to play important roles in the synergistic hepatotoxicity from MCT/LPS exposure in rats.
![]() |
ACKNOWLEDGMENTS |
---|
![]() |
NOTES |
---|
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Adamson, G. M., and Billings, R. E. (1992). Tumor necrosis factor induced oxidative stress in isolated mouse hepatocytes. Arch. Biochem. Biophys. 294, 223229.[ISI][Medline]
Allen, J. R., Chesney, C. F., and Frazee, W. J. (1972). Modifications of pyrrolizidine alkaloid intoxication resulting from altered hepatic microsomal enzymes. Toxicol. Appl. Pharmacol. 23, 470479.[ISI][Medline]
Badger, D. A., Kuester, R. K., Sauer, J.-M., and Sipes, I. G. (1997). Gadolinium chloride reduces cytochrome P450: Relevance to chemical induced hepatotoxicity. Toxicology 121, 143153.[ISI][Medline]
Barton, C. C., Barton E. X., Ganey, P. E., Kunkel, S. L., and Roth, R. A. (2001). Bacterial lipopolysaccharide enhances aflatoxin B1 hepatotoxicity in rats by a mechanism that is dependent on tumor necrosis factor . Hepatology 33, 6673.[ISI][Medline]
Bradham, C. A., Plümpe, J., Manns, M. P., Brenner, D. A., and Trautwein, C. (1998). Mechanisms of hepatic toxicity. I. TNF-induced liver injury. Am. J. Physiol. 275, G387392.
Brouwer, A., Parker, S. G., Hendriks, H. F. J., Gibbons, L., and Horan, M. A. (1995). Production of eicosanoids and cytokines by Kupffer cells from young and old rats stimulated by endotoxin. Clin. Sci. 88, 211217.[ISI][Medline]
Brown, A. P., Harkema, J. R., Schultze, A. E., Roth, R. A., and Ganey, P. E. (1997). Gadolinium chloride pretreatment protects against hepatic injury but predisposes the lung to alveolitis after lipopolysaccharide administration. Shock 7, 186192.[ISI][Medline]
Casteleijn, E., Kuiper, J., Van Rooij, H. C., Kamps, J. A., Koster, J. F., and Van Berkel, T. J. (1988). Endotoxin stimulates glycogenolysis in the liver by means of intercellular communication. J. Biol. Chem. 263, 69536955.
Copple, B. L., Banes, A., Ganey, P. E., and Roth, R. A. (2002). Endothelial cell injury and fibrin deposition in rat liver after monocrotaline exposure. Toxicol. Sci. 65, 309318.
Cruz-Orive, L. M. (1982). The use of quadrats and test systems in stereology, including magnification corrections. J. Microsc. 125, 89102.[ISI]
Deaciuc, I. V., Bagby, G. J., Neisman, M. R., Skrepnik, M., and Spitzer J. J. (1994). Modulation of hepatic sinusoidal endothelial cell function by Kupffer cells: An example of intracellular communication in the liver. Hepatology 19, 464470.[ISI][Medline]
Deaciuc, I. V., McDonough, K. H., Bagby, G. J., and Spitzer, J. J. (1993). Alcohol consumption in rats potentiates the deleterious effects of Gram-negative sepsis on hepatic hyaluronan uptake. Alcohol Clin. Exp. Res. 17, 10021008.[ISI][Medline]
Decker, K. (1990). Biologically active products of stimulated liver macrophages (Kupffer cells). Eur. J. Biochem. 192, 245261.[ISI][Medline]
DeLeve, L. D., McCuskey, R. S., Wang, X., Hu, L., McCuskey, M. K., Epstein, R. B., and Kanel, G. C. (1999). Characterization of a reproducible rat model of hepatic veno-occlusive disease. Hepatology 29, 17791791.[ISI][Medline]
Dezube, B. J., Sherman, M. L., Fridovich-Keil, J. L., Allen-Ryan, J., and Pardee, A. B. (1993). Down-regulation of tumor necrosis factor expression by pentoxifylline in cancer patients: A pilot study. Cancer Immunol. Immunother. 36, 5760.[ISI][Medline]
Dieter, P., Hempel, U., Kamionka, S., Kolada, A., Malessa, B., Fitzke, E., and Tran-Thi, T. A. (1999). Prostaglandin E2 affects differently the release of inflammatory mediators from resident macrophages by LPS and muramyl tripeptides. Mediators Inflamm. 8, 295303.[ISI][Medline]
El-Sisi, A. E. D., Earnest, D. L., and Sipes, I. G. (1993). Vitamin A potentiation of carbon tetrachloride-induced liver injury: Role of liver macrophages and active oxygen species. Toxicol. Appl. Pharmacol. 119, 295301.[ISI][Medline]
Futaki, N., Takahashi, S., Kitigawa, T., Yamakawa, Y., Tanaka, M., and Higuchi, S. (1997). Selective inhibition of cyclooxygenase-2 by NS-398 in endotoxin shock rats in vivo. Inflamm. Res. 46, 496502.[ISI][Medline]
Futaki, N., Takahashi, S., Yokoyama, M., Arai, I., Higuchi, S., and Otomo, S. (1994). NS-398, a new anti-inflammatory agent, selectively inhibits prostaglandin G/H synthase/cyclooxygenase (COX-2) activity in vitro. Prostaglandins 47, 5559.[Medline]
Ganey, P. E., Barton, Y.-W., Kinser, S., Sneed, R. A., Barton, C. C., and Roth, R. A. (2001). Involvement of cyclooxygenase-2 in the potentiation of allyl alcohol-induced liver injury by bacterial lipopolysaccharide. Toxicol. Appl. Pharmacol. 174, 113121.[ISI][Medline]
Ganey, P. E., and Roth, R. A. (2001). Concurrent inflammation as a determinant of susceptibility to toxicity from xenobiotic agents. Toxicology 169, 195208.[ISI][Medline]
Heller, S., Weber, K., Heller, A., Urbaschek, R., and Koch, T. (1999). Pentoxifylline improves bacterial clearance during hemorrhage and endotoxemia. Crit. Care Med. 27, 756763.[ISI][Medline]
Hewett, J. A., Schultze, A. E., Van Cise, S., and Roth, R. A. (1992). Neutrophil depletion protects against liver injury from bacterial endotoxin. Lab. Invest. 66, 347361.[ISI][Medline]
Hewett, J. A., Jean, P. A., Kunkel, S. L., and Roth, R. A. (1993). Relationship between tumor necrosis factor-alpha and neutrophils in endotoxin-induced liver injury. Am. J. Physiol. 265, G10111015.
Hoek, J. B., and Pastorino, J. G. (2002). Ethanol, oxidative stress, and cytokine-induced liver cell injury. Alcohol 27, 6368.[ISI][Medline]
Holst, O., Ulmer, A. J., Brade, H., Flad, H.-D., and Rietschel, E. T. (1996). Biochemistry and cell biology of bacterial endotoxins. FEMS Immunol. Med. Microbiol. 16, 83104.[ISI][Medline]
Iimuro, Y., Yamamoto, M., Kohno, H., Itakura, J., Fujii, H., and Matsumoto, Y. (1994). Blockade of liver macrophages by gadolinium chloride reduces lethality in endotoxemic ratesanalysis of mechanisms of lethality in endotoxemia. J. Leukoc. Biol. 55, 723728.[Abstract]
Jaeschke, H., Fisher, M. A., Lawson, J. A., Simmons, C. A., Farhood, A., and Jones, D. A. (1998). Activation of caspase 3 (CPP32)-like proteases is essential for TNF--induced hepatic parenchymal cell apoptosis and neutrophil-mediated necrosis in a murine endotoxin shock model. J. Immunol. 160, 34803486.
Kasahara, Y., Kiyatake, K., Ztatsumi, K., Sugito, K., Kakusaka, I., Yamagata, S.-I., Ohmori, S, Kitada, M., and Kuriyama, T. (1997). Bioactivation of monocrotaline by P-450 3A in rat liver. J. Cardiovasc. Pharmacol. 30, 124129.[ISI][Medline]
Kobayashi, H., Horikoshi, K., Yamataka, A., Yamataka, T., Okazaki, T., Lane, G. J., and Miyano, T. (1999). Hyaluronic acid: A specific prognostic indicator of hepatic damage in biliary atresia. J. Pediatr. Surg. 34, 17911794.[ISI][Medline]
Kushimoto, S., Okajima, K., Uchiba, M., Murakami, K., Harada, N., Okabe, H., and Takatsuki, K. (1996). Role of granulocyte elastase in ischemia/reperfusion injury of rat liver. Crit. Care Med. 24, 19081912.[ISI][Medline]
Lawson, J. A., Fisher, M. A., Simmons, C. A., Farhood, A., and Jaeschke, H. (1998). Parenchymal cell apoptosis as a signal for sinusoidal sequestration and transendothelial migration of neutrophils in murine models of endotoxin and Fas-antibody-induced liver injury. Hepatology 28, 761767.[ISI][Medline]
Levin, S., Bucci, T. J., Cohen, S. M., Fix, A. S., Hardisty, J. F., LeGrand, E. K., Maronpot, R. R., and Trump, B. F. (1999). The nomenclature of cell death: Recommendation of an ad hoc committee of the society of toxicologic pathologists. Toxicol. Pathol. 27, 484490.[ISI][Medline]
Majno, G., and Joris, I. (1995). Apoptosis, oncosis, and necrosis. An overview of cell death. Am. J. Pathol.146, 315.[Abstract]
Mattocks, A. R. (1986). Chemistry and Toxicology of Pyrrolizidine Alkaloids. Academic Press, London.
Michie, H. R., Manogue, K. B., Spriggs, D. R., Revhaug, A., O'Dwyer, S., Dinarello, C. A., Cerami, A., Wolff, S. M., and Wilmore, D. W. (1988). Detection of circulating tumor necrosis factor after endotoxin administration. N. Eng. J. Med. 318, 14811486.[Abstract]
Mimura, Y., Sakisaka, S., Harada, M., Sata, M., and Tanikawa, K. (1995). Role of hepatocytes in direct clearance of lipopolysaccharide in rats. Gastroenterology 109, 19691976.[ISI][Medline]
Nagaki, M., Muto, Y., Ohnishi, H., and Moriwaki, H. (1991). Significance of tumor necrosis factor (TNF) and interleukin-1 (IL-1) in the pathogenesis of fulminant hepatitis: Possible involvement of serine protease in TNF-mediated liver injury. Gastroenterology Jpn. 26, 448455.
Pearson, J. M., Brown, A. P., Schultze, A. E., Ganey, P. E., and Roth, R. A. (1996). Gadolinium chloride treatment attenuates hepatic platelet accumulation after lipopolysaccharide administration. Shock 5, 408415.[ISI][Medline]
Rose, M. L., Bradford, B. U., Germolec, D. R., Lin, M., Tsukamoto, H., and Thurman, R. G. (2001). Gadolinium chloride-induced hepatocyte proliferation is prevented by antibodies to tumor necrosis factor . Toxicol. Appl. Pharmacol. 170, 3945.[ISI][Medline]
Schoental, R., and Head, M. A. (1955). Pathological changes in rats as a result of treatment with monocrotaline. Br. J. Cancer 9, 229237.[ISI][Medline]
Spapen, H., Zhang, H., Wisse, E., Baekeland, M., Seynaeve, C., Eddouks, M, and Vincent, J. L. (1999). The 21-aminosteroid U74389G enhanced hepatic blood flow and preserved sinusoidal endothelial cell function and structure in endotoxin-shocked dogs. J. Surg. Res. 86, 183191.[ISI][Medline]
Spitzer, J. A., and Mayer, A. M. S. (1993). Hepatic neutrophil influx: Eicosanoid and superoxide formation in endotoxemia. J. Surg. Res. 55, 6067.[ISI][Medline]
Stachlewitz, R. F., Seabra, V., Bradford, B., Bradham, C. A., Rusyn, I., Germolec, D., and Thurman, R. G. (1999). Glycine and uridine prevent D-galactosamine hepatotoxicity in the rat: Role of Kupffer cells. Hepatology 29, 737745.[ISI][Medline]
Steele, R. G. D., Torrie, J. H., and Dickey, D. A. (1997). Principles and Procedures in Statistics: A Biometric Approach, 3rd ed. McGraw-Hill, New York.
Stegelmeier, B. L., Edgar, J. A., Colegate, S. M., Gardner, D. R., Scloch, T. K., Coulombe, R. A., and Molyneux, R. J. (1999). Pyrrolizidine alkaloid plants, metabolism and toxicity. J. Nat. Toxins 8, 95116.[ISI][Medline]
Vassalli, P. (1992). The pathophysiology of tumor necrosis factor. Annu. Rev. Immunol. 10, 411452.[ISI][Medline]
Vollmar, B., Rüttinger, D., Wanner, G. A, Leiderer, R., and Menger, M. D. (1996). Modulation of Kupffer cell activity by gadolinium chloride in endotoxemic rats. Shock 6, 434441.[ISI][Medline]
White, I. N. H., and Mattocks, A. R. (1972). Reaction of dihydropyrrolizidines with deoxyribonucleic acids in vitro. Biochem. J. 128, 291297.[ISI][Medline]
Yee, S. B., Kinser, S., Hill, D. A., Barton, C. C., Hotchkiss, J. A., Harkema, J. R., Ganey, P. E., and Roth, R. A. (2000). Synergistic hepatotoxicity from coexposure to bacterial endotoxin and the pyrrolizidine alkaloid monocrotaline. Toxicol. Appl. Pharmacol. 166, 173185.[ISI][Medline]
Yee, S. B., Copple, B. L., Ganey, P. E., and Roth, R. A. (2002). Sinusoidal endothelial cell injury during hepatotoxicity from monocrotaline and bacterial lipopolysaccharide coexposure. Toxicol. Sci. 66(Suppl. 1), 352.