NO plays a role in LPS-induced decreases in circulating IGF-I and IGFBP-3 and their gene expression in the liver

In this study, we administered aminoguanidine, a relatively selective inducible nitric oxide synthase (iNOS) inhibitor, to study the role of nitric oxide (NO) in LPS-induced decrease in IGF-I and IGFBP-3. Adult male Wistar rats were injected intraperitoneally with LPS (100 microg/kg), aminoguanidine (100 mg/kg), LPS plus aminoguanidine, or saline. Rats were injected at 1730 and 0830 the next day and killed 4 h after the last injection. LPS administration induced an increase in serum concentrations of nitrite/nitrate (P < 0.01) and a decrease in serum concentrations of growth hormone (GH; P < 0.05) and IGF-I (P < 0.01) as well as in liver IGF-I mRNA levels (P < 0.05). The LPS-induced decrease in serum concentrations of IGF-I and liver IGF-I gene expression seems to be secondary to iNOS activation, since aminoguanidine administration prevented the effect of LPS on circulating IGF-I and its gene expression in the liver. In contrast, LPS-induced decrease in serum GH was not prevented by aminoguanidine administration. LPS injection decreased IGFBP-3 circulating levels (P < 0.05) and its hepatic gene expression (P < 0.01), but endotoxin did not modify the serum IGFBP-3 proteolysis rate. Aminoguanidine administration blocked the inhibitory effect of LPS on both IGFBP-3 serum levels and its hepatic mRNA levels. When aminoguanidine was administered alone, IGFBP-3 serum levels were increased (P < 0.05), whereas its hepatic mRNA levels were decreased. This contrast can be explained by the decrease (P < 0.05) in serum proteolysis of this binding protein caused by aminoguanidine. These data suggest that iNOS plays an important role in LPS-induced decrease in circulating IGF-I and IGFBP-3 by reducing IGF-I and IGFBP-3 gene expression in the liver.     

Role of nitric oxide in tolerance to lipopolysaccharide in mice.

The injection of repeated doses of lipopolysaccharide (LPS) results in attenuation of the febrile response, which is called endotoxin tolerance. We tested the hypothesis that nitric oxide (NO) arising from inducible NO synthase (iNOS) plays a role in endotoxin tolerance, using not only pharmacological trials but also genetically engineered mice. Body core temperature was measured by biotelemetry in mice treated with NG-monomethyl-L-arginine (L-NMMA, 40 mg/kg; a nonselective NO synthase inhibitor) or aminoguanidine (AG, 10 mg/kg; a selective iNOS inhibitor) and in mice deficient in the iNOS gene (iNOS KO) mice. Tolerance to LPS was induced by means of three consecutive LPS (100 microg/kg) intraperitoneal injections at 24-h intervals. In wild-type mice, we observed a significant reduction of the febrile response to repeated administration of LPS. Injection of L-NMMA and AG markedly enhanced the febrile response to LPS in tolerant animals. Conversely, iNOS-KO mice repeatedly injected with LPS did not become tolerant to the pyrogenic effect of LPS. These data are consistent with the notion that NO modulates LPS tolerance in mice and that iNOS isoform is involved in NO synthesis during LPS tolerance.     

Reduced lipopolysaccharide (LPS)-induced nitric oxide production in peritoneal macrophages and inhibited LPS-induced lethal shock in mice by a sugar cane (Saccharum officinarum L.) extract

A sugar cane extract (SCE) has been found to have an immunostimulating effect in several animals. Lipopolysaccharide (LPS) is known to induce endotoxin shock via the production of inflammatory modulators such as tumor necrosis factor (TNF)-alpha and nitric oxide (NO). We examined in the present study the effects of SCE on the TNF-alpha and NO production in LPS-stimulated mice peritoneal cells and the endotoxin shock in mice. The supplementation of SCE to peritoneal macrophages cultured with LPS resulted in a significant decrease in NO production. All the mice injected intraperitoneally with LPS and D-galactosamine (LPS+GalN) died within 24 h. However, a peritoneal injection, but no intravenous or oral administration, of SCE (500-1,000 mg/kg) at 3 to 48 h before the LPS+GalN-challenge resulted in a significantly improved survival rate. These results suggest that SCE had a protective effect on LPS-induced endotoxin shock via one of possible mechanisms involving the suppression of NO production in the mouse peritoneal cavity.     

Intracerebroventricular injection of neuronal and inducible nitric oxide synthase inhibitors attenuates fever due to LPS in rats.

Nitric oxide (NO) has been shown to be an important mediator of febrile response to lipopolisaccharide (LPS). To clarify the role of different isoforms of NO synthase (NOS) in febrile response to immune challenge, effects of selective iNOS and nNOS inhibitors on fever to LPS were examined in freely moving biotelemetered rats. Vinyl-L-NIO (N(5) - (1-Imino-3-butenyl) - ornithine (vL-NIO), a neuronal nitric oxide synthase (nNOS) inhibitor, and aminoguanidine hydrochloride, an inducible nitric oxide synthase (iNOS) inhibitor, were injected intracerebroventricularly at a dose of 10 microg/rat just before intraperitoneal injection of LPS at a dose of 50 microg/kg. Both inhibitors injected at a selected doses had no effect on normal day-time body temperature (T(b)) and normal night-time T(b). vinyl-L-NIO and aminoguanidine injected intracerebroventricularly at a dose of 10 microg/animal suppressed the LPS-induced fever in rats. The fever index calculated for rats pretreated with v-LNIO or with aminoguanidine and injected with LPS was reduced by 43% and 72%, respectively, compared to that calculated for water-pretreated and LPS-injected rats. Whereas vL-NIO partly attenuated both phases of febrile rise in T(b), administration of aminoguanidine into the brain completely prevented fever induced by LPS. These data indicate that activation of iNOS inside the brain is not only responsible for triggering but also for maintaining of LPS-induced fever in rats. It is, therefore, reasonable to hypothesize that, activation of iNOS inside the brain is more important in fever development than activation of nNOS.     

Reduced Lipopolysaccharide (LPS)-Induced Nitric Oxide Production in Peritoneal Macrophages and Inhibited LPS-Induced Lethal Shock in Mice by a Sugar Cane (Saccharum officinarum L.) Extract

A sugar cane extract (SCE) has been found to have an immunostimulating effect in several animals. Lipopolysaccharide (LPS) is known to induce endotoxin shock via the production of inflammatory modulators such as tumor necrosis factor (TNF)-α and nitric oxide (NO). We examined in the present study the effects of SCE on the TNF-α and NO production in LPS-stimulated mice peritoneal cells and the endotoxin shock in mice. The supplementation of SCE to peritoneal macrophages cultured with LPS resulted in a significant decrease in NO production. All the mice injected intraperitoneally with LPS and D-galactosamine (LPS+GalN) died within 24 h. However, a peritoneal injection, but no intravenous or oral administration, of SCE (500–1,000 mg/kg) at 3 to 48 h before the LPS+GalN-challenge resulted in a significantly improved survival rate. These results suggest that SCE had a protective effect on LPS-induced endotoxin shock via one of possible mechanisms involving the suppression of NO production in the mouse peritoneal cavity.     

Inhibitory effect of vitamin E on proinflammatory cytokines-and endotoxin-induced nitric oxide release in alveolar macrophages

Nitric oxide is thought to be an important modulator of various functions in normal and inflamed airways. In the present study, we evaluated the effects of high vitamin E (250 mg and 1250 mg alpha-tocopheryl acetate (TA)/kg diet/10 days) on nitric oxide (NO(.)) release by alveolar macrophages (AMs) in response to lipopolysaccharide (LPS), interleukin-1beta (IL-1beta) and tumor necrosis factor (TNF-alpha). LPS and IL-1beta treatment (1-10 microg/ml) enhanced NO(.) release in AMs from control animals fed on 50 mg vitamin E/kg diet in a concentration dependent manner. However, this enhancement of NO(.) was attenuated in the AMs of animals fed with 250 mg or 1250 mg vitamin E/kg diet. TNF-alpha had no effect in eliciting the release of NO(.) in AMs obtained either from control or from hyper vitamin E fed animals. Further, LPS (1-10 microg/ml) enhanced the inducible nitric oxide synthase (iNOS) activity of AMs of control group and TA-fed animals almost to equal extent. Similarly, LPS-induced formation of N-nitrosamine (N-nitroso-L-[(14)C]-proline) in AMs of control and TA-supplemented animals were not different statistically. On the other hand, in vitro addition of vitamin E (200 microM) in AMs of control animals, when triggered with 10 microg LPS/ml, caused a significant decrease in N-nitroso-L-[(14)C]-proline formation. It seems that high doses of TA in diet may play a role in reducing the lipopolysaccharide and proinflammatory cytokines-induced NO(.)-mediated damage by AMs.     

Effect of 5-lipoxygenase inhibitor against lipopolysaccharide-induced hypothermia in mice

Bacterial endotoxin produces sepsis associated with alterations in body temperature (fever or hypothermia). The intraperitoneal administration of bacterial endotoxin, lipopolysaccharide (LPS; 50 microg/mouse) led to a decrease in colonic temperature starting 1 hr after the injection. The hypothermic effect was accompanied by a significant increase in hypothalamic leukotriene B4 (LTB4) and prostaglandin E2 (PGE2) levels. 5-lipoxygenase inhibitor, zileuton (200 and 400 mg/kg, po) administered 30 min before LPS challenge significantly prevented hypothermia. However, non-selective cyclooxygenase inhibitor, indomethacin (10, 20 mg/kg, po) did not reverse the hypothermic response. Further, pretreatment of mice with zileuton prevented LPS-stimulated increase in hypothalamic LTB4 levels and caused a relatively small increase in PGE2 levels. Indomethacin had no effect on LTB4 levels but it reduced PGE2 levels. These results suggest a possible involvement of leukotrienes in LPS-induced hypothermia and the potential protective role of 5-lipoxygenase inhibitors in endotoxemia.     

Up-regulation of rat adrenomedullin gene expression by endotoxin: Relation to nitric oxide synthesis

The effect of bacterial endotoxin (LPS) on adrenomedullin (AM) gene expression was investigated in cultured rat aortic vascular smooth muscle (VSM) cells and in tissues from anesthetized rats. The addition of LPS together with interferon-y to VSM cells resulted in a marked increase in the abundance of AM mRNA as well as the appearance of mRNA for the inducible isoform of nitric oxide (NO) synthase (iNOS). Intravenous injection of LPS into rats also increased AM mRNA abundance and induced iNOS mRNA in lung, heart, liver, and kidney. AM significantly enhanced NO synthesis evoked by LPS and interferon-v in cultured VSM cells. These data suggest that AM may contribute to circulatory failure during endotoxin shock, in part, by modulating NO synthesis.     

Inhibitory effect of FK 506 and cyclosporin A on nitric oxide production by LPS-treated cultured rat macrophages

We analyzed the effect of FK 506 on the production of nitric oxide by macrophages. Isolated rat peritoneal macrophages were cultured for 24 h with or without lipopolysaccharide (LPS) (5 microg/ml) and in the absence or presence of FK 506 (0.1 and 1 microg/ml). The concentration of NO2- in culture supernatants was taken as a measure of nitric oxide production. FK 506 (0.1 and 1 microg/ml) reduced the LPS-induced increase of NO2- levels by 68% and 81%, respectively. The impact of cyclosporin A (CsA) was studied in order to compare their effects. CsA (0.1 and 1 microg/ml) decreased the levels of nitrites by 39% and 69%, respectively. The results obtained suggest that both immunosuppressive drugs exhibit a dose-dependent inhibitory effect on nitric oxide production and that FK 506 is a more potent agent than CsA in this respect.     

Identification of a novel compound that inhibits iNOS and COX-2 expression in LPS-stimulated macrophages from Schisandra chinensis

A novel α-iso-cubebenol, which has anti-inflammatory effects in lipopolysaccharide (LPS)-treated RAW 264.7 macrophages, was isolated from the fruits of Schisandra chinensis. α-iso-cubebenol inhibited LPS-induced nitric oxide (NO) and prostaglandin E₂ (PGE₂) production. Consistent with these findings, α-iso-cubebenol also reduced the LPS-induced expression of inducible nitric oxide synthase and cyclooxygenase-2 at the protein and mRNA levels in a concentration-dependent manner. α-iso-cubebenol also inhibited LPS-induced nuclear translocation of the NF-κB p65 subunit. Furthermore, α-iso-cubebenol suppressed the phosphorylation of ERK, JNK, and p38 kinase induced by LPS. Since the novel α-iso-cubebenol blocked the production of several pro-inflammatory mediators induced by LPS in macrophages, the molecule can be useful material for the development of anti-inflammatory agents against bacterial infections or endotoxin.     

Lipopolysaccharide (LPS)-induced intra-uterine fetal death (IUFD) in mice is principally due to maternal cause but not fetal sensitivity to LPS

The present study deals with whether lipopolysaccharide (LPS)-induced intra-uterine fetal death (IUFD) is related to LPS-susceptibility of either mother or fetus and how LPS or LPS-induced TNF causes IUFD. LPS-susceptible C3H/HeN or -hypo-susceptible C3H/HeJ pregnant mice and the mice mated reciprocally with these mice were used on days 14 to 16 of gestation for experiments. All of fetuses in pregnant C3H/HeN mice mated with either C3H/HeN males [HeN(HeN)] or C3H/HeJ males [HeN(HeJ)] were killed within 24 hr when injected intravenously (i.v.) with 50 or 100 microg of LPS. On the other hand, the majority of fetuses in C3H/HeJ females mated with either C3H/HeJ males [HeJ(HeJ)] or C3H/HeN males [HeJ(HeN)] survived when injected i.v. with even 400 microg of LPS. These findings indicate that LPS-induced IUFD depends on the maternal LPS-responsiveness. LPS injected into mothers could pass through placenta to fetuses, since an injection with 125I-labeled LPS or IgG into pregnant mice resulted in considerable levels of radioactivity in fetuses as well as placenta. Cultured peritoneal macrophages derived from F1 mice of HeJ(HeN) or HeN(HeJ) mice, produced nitric oxide (NO) and tumor necrosis factor (TNF) in response to LPS, although the levels of NO and TNF were lower in comparison with those of C3H/HeN macrophage cultures, suggesting a possibility that the fetus as well as F1 cells might be responsible to LPS. LPS-induced IUFD was not blocked by treatment with anti-TNF antibody which inhibited LPS-induced TNF production in pregnant females, although an injection of recombinant TNFalpha instead of LPS could induce IUFD, suggesting that the cause of IUFD cannot be attributed to mother-derived TNF alone. The roles of LPS passed through placenta and LPS-induced mediators on IUFD were discussed.     

Melatonin inhibits expression of the inducible NO synthase II in liver and lung and prevents endotoxemia in lipopolysaccharide-induced multiple organ dysfunction syndrome in rats.

We evaluated the role of melatonin in endotoxemia caused by lipopolysaccharide (LPS) in unanesthetized rats. The expression of inducible isoform of nitric oxide synthase (iNOS) and the increase in the oxidative stress seem to be responsible for the failure of lungs, liver, and kidneys in endotoxemia. Bacterial LPS (10 mg/kg b. w) was i.v. injected 6 h before rats were killed and melatonin (10-60 mg/kg b.w.) was i.p. injected before and/or after LPS. Endotoxemia was associated with a significant rise in the serum levels of aspartate and alanine aminotransferases, gamma-glutamyl-transferase, alkaline phosphatase, creatinine, urea, and uric acid, and hence liver and renal dysfunction. LPS also increased serum levels of cholesterol and triglycerides and reduced glucose levels. Melatonin administration counteracted these organ and metabolic alterations at doses ranging between 20 and 60 mg/kg b. w. Melatonin significantly decreased lung lipid peroxidation and counteracted the LPS-induced NO levels in lungs and liver. Our results also show an inhibition of iNOS activity in rat lungs by melatonin in a dose-dependent manner. Expression of iNOS mRNA in lungs and liver was significantly decreased by melatonin (60 mg/kg b. w., 58-65%). We conclude that melatonin inhibits NO production mainly by inhibition of iNOS expression. The inhibition of NO levels may account for the protection of the indoleamine against LPS-induced endotoxemia in rats.     

Inhibition of LPS-induced iNOS and NO synthesis in primary rat microglial cells

Nitric oxide (NO) has been implicated in the etiopathology of multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE), and inhibition of NO synthesis has been proposed to be a possible mechanism of action of drugs to treat MS. In the present study, we investigated the inhibitory effect on NO synthesis of various steroids, cytokines and drugs used or proposed for the treatment of MS. As a model system, we used primary rat microglial cells which produce NO synthase and subsequently release NO upon stimulation with lipopolysaccharide (LPS). Among the substances tested, the glucocorticoids prednisone, hydrocortisone, dexamethasone and progesterone as well as transforming growth factor-beta (TGF-beta) dose-dependently inhibited LPS-induced nitric oxide synthase (iNOS) and NO synthesis. In contrast, COP-1, the phosphodiesterase inhibitors rolipram and pentoxifylline, the cytokines interleukin-10 (IL-10) and interferon-beta (IFN-beta) as well as the steroids beta-estradiol, testosterone, and dehydroepiandrosterone (DHEA) showed no inhibitory effect. Cholesterol slightly, but not significantly, increased LPS-induced nitric oxide synthesis. We conclude from the present study that with respect to treatment of MS, inhibition of NO synthesis may be an important mechanism of action of glucocorticoids and transforming growth factor-beta, but not of other drugs used or proposed to treat MS.     

N-acetylcysteine inhibits in vivo nitric oxide production by inducible nitric oxide synthase.

This in vivo study evaluates the effect of N-acetylcysteine (NAC) administration on nitric oxide (NO) production by the inducible form of nitric oxide synthase (iNOS). NO production was induced in the rat by the ip administration of 2 mg/100 g lipopolysaccharide (LPS). This treatment caused: (1) a decrease in body temperature within 90 min, followed by a slow return to normal levels; (2) an increase in plasma levels of urea, nitrite/nitrate, and citrulline; (3) the appearance in blood of nitrosyl-hemoglobin (NO-Hb) and in liver of dinitrosyl-iron-dithiolate complexes (DNIC); and (4) increased expression of iNOS mRNA in peripheral blood mononuclear cells (PBMC). Rat treatment with 15 mg/100 g NAC ip, 30 min before LPS, resulted in a significant decrease in blood NO-Hb levels, plasma nitrite/nitrate and citrulline concentrations, and liver DNIC complexes. PBMC also showed a decreased expression of iNOS mRNA. NAC pretreatment did not modify the increased levels of plasma urea or the hypothermic effect induced by the endotoxin. The administration of NAC following LPS intoxication (15 min prior to sacrifice) did not affect NO-Hb levels. These results demonstrate that NAC administration can modulate the massive NO production induced by LPS. This can be attributed mostly to the inhibitory effect of NAC on one of the events leading to iNOS protein expression. This hypothesis is also supported by the lack of effect of late NAC administration.     

Psychosocial stress affects the involvement of prostaglandins and nitric oxide in the lipopolysaccharide-induced hypothalamic-pituitary-adrenal response.

The role of prostaglandins and nitric oxide (NO), generated after peripheral lipopolysaccharide (LPS) administration, in the adaptation of hypothalamic-pituitary-adrenal (HPA) axis under stressful circumstances remains to be elucidated. The aim of the present study was to assess the effect of chronic repetitive restraint or social crowding stress on the involvement of nitric oxide and prostaglandins in the LPS-induced pituitary-adrenocortical response. Male Wistar rats were restrained in metal tubes 2 x 10 min/day or crowded in cages for 7 days prior to treatment. All compounds were injected i.p., cyclooxygenase (COX) and nitric oxide synthase (NOS) inhibitors 15 min before LPS. Two hrs after injection LPS induced a significant increase in ACTH and corticosterone secretion. Repeated restraint impaired more potently than crowding stress the LPS-induced HPA-response. Indomethacin, a non-selective COX inhibitor, considerably reduced the LPS-induced HPA response in non-stressed rats and to a lesser extent diminished this response in repeatedly restrained or crowded rats. Neuronal NOS inhibitor, Nomega-nitro-L-arginine decreased the LPS-induced HPA response, more potently in control than crowded rats. Aminoguanidine, an iNOS inhibitor, diminished the LPS-elicited ACTH response in crowded rats. These results indicate that prostaglandins and NO generated by neuronal and inducible NOS are involved in the LPS-induced HPA axis response under basal conditions and during its adaptation to chronic social stress circumstances.     

Investigation of the mechanisms involved in the central effects of glucagon-like peptide-1 on ethanol-induced gastric mucosal lesions

The aim of this study was to investigate the effects of intracerebroventricularly injected glucagon-like peptide-1 (GLP-1) on ethanol-induced gastric mucosal damage and to elucidate the mechanisms involved. Absolute ethanol was administered through an orogastric cannula 5 min before GLP-1 (1 microg/10 microl) injection. One hour later, the rats were decapitated, their stomachs were removed and scored for mucosal damage. GLP-1 inhibited the ethanol-induced gastric mucosal damage by 92%. Centrally injected atropine sulphate, a muscarinic receptor antagonist (5 microg/10 microl), prevented the gastroprotective effect of GLP-1, while mecamylamine, a nicotinic receptor antagonist (25 microg/10 microl), was ineffective. Peripherally injected atropine methyl nitrate (1 mg/kg) did not change the effect of GLP-1, but mecamylamine (5 mg/kg) blocked it. Cysteamine, a somatostatin depletor (280 mg/kg, s.c.), did not affect the protective activity of GLP-1, while inhibition of nitric oxide (NO) synthesis by L-NAME (3 mg/kg, i.v.) significantly abolished the protective effect of GLP-1 on ethanol-induced gastric mucosal lesions. We conclude that central muscarinic and peripheral nicotinic cholinergic receptors and NO, but not somatostatin, contribute to the protective effect of intracerebroventricularly injected GLP-1 on ethanol-induced gastric mucosal damage.     

Expression of glucose transporter GLUT3 by endotoxin in cultured rat astrocytes: the role of nitric oxide

The induction of nitric oxide (NO) synthase in astrocytes by endotoxin and/or cytokine treatment is associated with increased glucose consumption and glycolysis, but the mechanism whereby this phenomenon occurs remains obscure. In this work, we have addressed this issue and found that incubation of cultured rat astrocytes with lipopolysaccharide (LPS; 1 microg/mL) for 24 h increased the level of constitutively expressed GLUT1 glucose transporter mRNA, and triggered GLUT3 mRNA expression, which was absent in normal astrocytes. The occurrence of GLUT3 protein after LPS treatment was corroborated by western blotting and immunocytochemistry. A 4-h incubation of astrocytes in the absence of glucose, or under an oxygen-poor (3%) atmosphere also resulted in GLUT3 mRNA overexpression. Experiments performed with 2-deoxy-D-[U-14C]glucose (at 0.1 mM of D-glucose) confirmed that LPS (0.1-10 microg/mL) dose-dependently increased the rate of glucose uptake (by a factor of 1.6 at 1 microg/mL of LPS), which was paralleled with the increase in NO synthesis. Furthermore, blockade of NO synthase with 2-amino-5,6-dihydro-6-methyl-(4H)-1,3-thiazine (AMT; 50 microM) partially (by 45%) prevented the LPS-mediated increase in glucose uptake. Finally, incubation of astrocytes with the NO donor 1-[2-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (DETA; 100 microM) increased by a factor of 1.4 the rate of glucose uptake. We conclude that the increase in GLUT3-driven glucose uptake in astrocytes would have a neuroprotective role under conditions in which NO formation is combined with hypoglycaemia, such as in brain ischemia.     

Effect of a potent iNOS inhibitor (ONO-1714) on acetaminophen-induced hepatotoxicity in the rat

Overproduction of nitric oxide (NO) in the liver has been implicated as an important event in endotoxin shock and in other models of hepatic inflammation and injury. The present study was undertaken to evaluate the effect of ONO-1714, a potent and specific inhibitor of inducible NO synthase (iNOS), on acetaminophen-induced hepatotoxicity in the rats. Oral administration of ONO-1714 dose-dependently inhibited NOx (NO2- and NO3-) accumulation in rat plasma after lipopolysaccharide (LPS) treatment. Intraperitoneal acetaminophen at 1 g/kg caused damage to the centrilobular regions of the liver and increase in serum alanine and aspartate transaminase (ALT and AST, respectively) levels accompanied by elevated plasma NOx levels after 24 h. Oral administration of ONO-1714 at 10 and 100 microg/kg dose-dependently reduced the acetaminophen-induced hepatic tissue damage and the increases in serum ALT and AST levels. ONO-1714 also blocked the increase in plasma NOx concentrations. These findings demonstrate that oral ONO-1714, an iNOS inhibitor, protects against acetaminophen-evoked hepatic inflammation/injury, strongly suggesting that NO produced by iNOS plays a key role in the pathogenesis of this drug-induced hepatotoxicity.