Complication Of Acute Liver Failure Research Articles

Microglia contribute to ammonia-induced astrocyte swelling in culture

Brain edema, a lethal complication of acute liver failure (ALF), is believed to be largely cytotoxic due to the swelling of astrocytes. Ammonia, a principal neurotoxin in ALF, has been strongly implicated in the development of the brain edema. It was previously shown that treatment of cultured astrocytes with ammonia (5 mM NH₄Cl) results in cell swelling. While ammonia continues to exert a direct effect on astrocytes, it is possible that ammonia can affect other neural cells, particularly microglia. Microglia are capable of evoking an inflammatory response, a process known to contribute to the brain edema. We therefore examined the potential role of microglia in the mechanism of ammonia-induced astrocyte swelling. Conditioned media (CM) derived from ammonia-treated cultured microglia when added to cultured astrocytes resulted in significant cell swelling. Such swelling was synergistically increased when astrocytes were additionally treated with 5 mM ammonia. CM from ammonia-treated microglia also showed significant release of oxy-radicals and nitric oxide into the CM. CM from ammonia-treated microglia containing Tempol (a superoxide scavenger) or uric acid (a peroxynitrite scavenger) when added to astrocytes resulted in marked reduction in the cell swelling. Together, these studies indicate that microglia contribute to the ammonia-induced astrocyte swelling by a mechanism involving oxidative/nitrosative stress.

Real-time analysis of microglial activation and motility in hepatic and hyperammonemic encephalopathy

Hepatic encephalopathy (HE) is a potentially fatal complication of acute liver failure, associated with severe neurological dysfunction and coma. The brain’s innate immune cells, microglia, have recently been implicated in the pathophysiology of HE. To date, however, only ex vivo studies have been used to characterize microglial involvement. Our study uses in vivo two-photon imaging of awake-behaving mice expressing enhanced green fluorescent protein (eGFP) under the Cx3cr1 promoter to examine microglial involvement in two different models of encephalopathy – a slower, fatal model of azoxymethane-induced HE and a rapid, reversible acute hyperammonemic encephalopathy (AHE) induced by an ammonia load. To investigate the potential contribution of microglia to the neurological deterioration seen in these two models, we developed a software to analyze microglial activation and motility in vivo. In HE, we found that microglia do not become activated prior to the onset of neurological dysfunction, but undergo activation with mildly impaired motility during the terminal stage IV. We demonstrate that this microglial activation coincides with blood–brain barrier (BBB) opening and brain edema. Conversely, both microglial activation and motility are unchanged during AHE, despite the mice developing pathologically increased plasma ammonia and severe neurological dysfunction. Our study indicates that microglial activation does not contribute to the early neurological deterioration observed in either HE or AHE. The late microglial activation in HE may therefore be associated with terminal BBB opening and brain edema, thus exacerbating the progression to coma and increasing mortality.

Pathogenesis of Hepatic Encephalopathy

Hepatic encephalopathy can be a serious complication of acute liver failure and chronic liver diseases, predominantly liver cirrhosis. Hyperammonemia plays the most important role in the pathogenesis of hepatic encephalopathy. The brain-blood barrier disturbances, changes in neurotransmission, neuroinflammation, oxidative stress, GABA-ergic or benzodiazepine pathway abnormalities, manganese neurotoxicity, brain energetic disturbances, and brain blood flow abnormalities are considered to be involved in the development of hepatic encephalopathy. The influence of small intestine bacterial overgrowth (SIBO) on the induction of minimal hepatic encephalopathy is recently emphasized. The aim of this paper is to present the current views on the pathogenesis of hepatic encephalopathy.

1014 Critical Care Management of Fulminant Hepatitis

Background:Fulminant hepatitis in pediatrics is the result of acute liver failure, and it may have multiple causes. Without support therapy and/or liver transplantation, mortality may reach 70% or more. The developments of fulminant hepatitis in adults, including liver necrosis and encephalopathy within 8 wees, are different than those in pediatrics, especially if fulminant hepatitis is secondary to a metabolic or autoimmune disease. Congenital metabolic defects and infections are common in infants, whereas viral hepatitis and drug-induced liver failure are more common in older children. Goals: Screening for infections and drugs is important to determine the cause of liver failure. The biochemical features evidence significant conjugated hyperbilirubinemia, high aminotransferase, high blood ammonia levels and coagulopathy. Methodology: Management of acute liver failure includes: Assessment of prognosis for liver transplantation Prevention of complications while awaiting hepatic regeneration or liver transplantation. Hepatic support Many are the complications of acute liver failure. Management includes fluid restriction to 75% of maintenance using dextrose or high-calorie enteral nutrition. It is extremely important to maintain proper glucose levels. In the early stages of establishing the prognosis and requirements for liver transplantation, coagulation support is not recommended. If coagulation is severe (prothrombin time > 60”) and/ or the decision for liver transplantation has been made. Conclusion: Acute liver failure in pediatrics is rare but fatal disease. The development of effective supportive medical therapies and outcome of liver transplantation improve the prognosis, however also depends of the level of nursing care.

Marked potentiation of cell swelling by cytokines in ammonia-sensitized cultured astrocytes

BackgroundBrain edema leading to high intracranial pressure is a lethal complication of acute liver failure (ALF), which is believed to be cytotoxic due to swelling of astrocytes. In addition to the traditional view that elevated levels of blood and brain ammonia are involved in the mechanism of brain edema in ALF, emerging evidence suggests that inflammatory cytokines also contribute to this process. We earlier reported that treatment of astrocyte cultures with a pathophysiological concentration of ammonia (5 mM NH4Cl) resulted in the activation of nuclear factor-kappaB (NF-κB) and that inhibition of such activation diminished astrocyte swelling, suggesting a key role of NF-κB in the mechanism of ammonia-induced astrocyte swelling. Since cytokines are also well-known to activate NF-κB, this study examined for additive/synergistic effects of ammonia and cytokines in the activation of NF-κB and their role in astrocyte swelling.MethodsPrimary cultures of astrocytes were treated with ammonia and cytokines (TNF-α, IL-1, IL-6, IFN-γ, each at 10 ng/ml), individually or in combination, and cell volume was determined by the [3H]-O-methylglucose equilibration method. The effect of ammonia and cytokines on the activation of NF-κB was determined by immunoblots.ResultsCell swelling was increased by ammonia (43%) and by cytokines (37%) at 24 h. Simultaneous co-treatment with cytokines and ammonia showed no additional swelling. By contrast, cultures pretreated with ammonia for 24 h and then exposed to cytokines for an additional 24 h, showed a marked increase in astrocyte swelling (129%). Treatment of cultures with ammonia or cytokines alone also activated NF-κB (80-130%), while co-treatment had no additive effect. However, in cultures pre-treated with ammonia for 24 h, cytokines induced a marked activation of NF-κB (428%). BAY 11-7082, an inhibitor of NF-κB, completely blocked the astrocyte swelling in cultures pre-treated with ammonia and followed by the addition of a mixture of cytokines.ConclusionOur results indicate that ammonia and a mixture of cytokines each cause astrocyte swelling but when these agents are added simultaneously, no additive effects were found. On the other hand, when cells were initially treated with ammonia and 24 h later given a mixture of cytokines, a marked potentiation in cell swelling and NF-κB activation occurred. These data suggest that the potentiation in cell swelling is a consequence of the initial activation of NF-κB by ammonia. These findings provide a likely mechanism for the exacerbation of brain edema in patients with ALF in the setting of sepsis/inflammation.

The Na–K–Cl cotransporter in the brain edema of acute liver failure

Astrocyte swelling and brain edema associated with increased intracranial pressure are major complications of acute liver failure (ALF). The mechanism for such astrocyte swelling/brain edema, however, is not well understood. We recently found that ammonia, a key etiological factor in ALF, caused the activation of the Na-K-Cl cotransporter-1 (NKCC1) in cultured astrocytes, and that inhibition of such activation led to a reduction in astrocyte swelling, suggesting that NKCC1 activation may be an important factor in the mechanism of brain edema in ALF. To determine whether NKCC activation is also involved in brain edema in vivo, we examined whether NKCC activation occurs in the thioacetamide (TAA) rat model of ALF and determined whether treatment with the NKCC inhibitor bumetanide reduces the severity of brain edema in TAA-treated rats. Brain water content was measured using the gravimetric method. NKCC1 phosphorylation and protein expression were measured by Western blots. NKCC activity was measured in brain cortical slices. NKCC activity was elevated in brain cortical slices of TAA-treated rats as compared to sham animals. Western blot analysis showed significant increases in total as well as phosphorylated (activated) NKCC1 protein expression in the cortical tissue. These findings were associated with a significant increase in brain water content which was attenuated by treatment with the NKCC inhibitor bumetanide. Our studies suggest the involvement of NKCC in the development of brain edema in experimental ALF, and that targeting NKCC may represent a useful therapeutic strategy in humans with ALF.

Brain Aquaporin-4 in Experimental Acute Liver Failure

Intracranial hypertension caused by brain edema and associated astrocyte swelling is a potentially lethal complication of acute liver failure (ALF). Mechanisms of edema formation are not well understood, but elevated levels of blood and brain ammonia and its by-product glutamine have been implicated in this process. Since aquaporin-4 (AQP4) has been implicated in brain edema in other conditions, we examined its role in a rat model of ALF induced by the hepatotoxin thioacetamide. Rats with ALF showed increased AQP4 protein in the plasma membrane (PM). Total tissue levels of AQP4 protein and mRNA levels were not altered, indicating that increased AQP4 is not transcriptionally mediated but likely reflects a more stable anchoring of AQP4 to the PM and/or interference with its degradation. An increase inAQP4 immunoreactivity in thePM was observed in perivascular astrocytes in ALF. Rats with ALF also showed increased levels of α-syntrophin, a protein involved in anchoringAQP4 to perivascular astrocytic end-feet. Increased AQP4 andα-syntrophin levels were inhibited by L-histidine, an inhibitor of glutamine transport into mitochondria, suggesting a role for glutamine in the increase of PM levels of AQP4. These results indicate that increased AQP4 PM levels in perivascular astrocytic end-feet are likely critical to the development of brain edema in ALF.

Pathogenesis and diagnosis of hepatic encephalopathy

Hepatic encephalopathy (HE) is a common and potentially devastating neuropsychiatric complication of acute liver failure and cirrhosis. Even in its mildest form, minimal HE (MHE), the syndrome significantly impacts daily living and heralds progression to overt HE. There is maturity in the scientific understanding of the cellular processes that lead to functional and structural abnormalities in astrocytes. Hyperammonemia and subsequent cell swelling is a key pathophysiological abnormality, but this aspect alone is insufficient to fully explain the complex neurotransmitter abnormalities that may be observable using sophisticated imaging techniques. Inflammatory cytokines, reactive oxygen species activation and the role of neurosteroids on neurotransmitter binding sites are emerging pathological lines of inquiry that have yielded important new information on the processes underlying HE and offer promise of future therapeutic targets. Overt HE remains a clinical diagnosis and the neurophysiological and imaging modalities used in research studies have not transferred successfully to the clinical situation. MHE is best characterized by psychometric evaluation, but these tests can be lengthy to perform and require specific expertise to interpret. Simpler computer-based tests are now available and perhaps offer an opportunity to screen, diagnose and monitor MHE in a clinical scenario, although large-scale studies comparing the different techniques have not been undertaken. There is a discrepancy between the depth of understanding of the pathophysiology of HE and the translation of this understanding to a simple, easily understood diagnostic and longitudinal marker of disease. This is a present area of focus for the management of HE.

Treatment of hepatic encephalopathy by on-line hemodiafiltration: a case series study

BackgroundIt is thought that a good survival rate of patients with acute liver failure can be achieved by establishing an artificial liver support system that reliably compensates liver function until the liver regenerates or a patient undergoes transplantation. We introduced a new artificial liver support system, on-line hemodiafiltration, in patients with acute liver failure.MethodsThis case series study was conducted from May 2001 to October 2008 at the medical intensive care unit of a tertiary care academic medical center. Seventeen consecutive patients who admitted to our hospital presenting with acute liver failure were treated with artificial liver support including daily on-line hemodiafiltration and plasma exchange.ResultsAfter 4.9 ± 0.7 (mean ± SD) on-line hemodiafiltration sessions, 16 of 17 (94.1%) patients completely recovered from hepatic encephalopathy and maintained consciousness for 16.4 ± 3.4 (7-55) days until discontinuation of artificial liver support (a total of 14.4 ± 2.6 [6-47] on-line hemodiafiltration sessions). Significant correlation was observed between the degree of encephalopathy and number of sessions of on-line HDF required for recovery of consciousness. Of the 16 patients who recovered consciousness, 7 fully recovered and returned to society with no cognitive sequelae, 3 died of complications of acute liver failure except brain edema, and the remaining 6 were candidates for liver transplantation; 2 of them received living-related liver transplantation but 4 died without transplantation after discontinuation of therapy.ConclusionsOn-line hemodiafiltration was effective in patients with acute liver failure, and consciousness was maintained for the duration of artificial liver support, even in those in whom it was considered that hepatic function was completely abolished.

Neuropathological changes in the brain of pigs with acute liver failure

Objective. Cerebral edema is a serious complication of acute liver failure (ALF), which may lead to intracranial hypertension and death. An accepted tenet has been that the blood-brain barrier is intact and that brain edema is primarily caused by a cytotoxic etiology due to hyperammonemia. However, the neuropathological changes in ALF have been poorly studied. Using a well characterized porcine model we aimed to investigate ultrastructural changes in the brain from pigs suffering from ALF. Materials and methods. Sixteen female Norwegian Landrace pigs weighing 27–35 kg were randomised into two groups: ALF (n = 8) and sham operated controls (n = 8). ALF was induced with an end-to-side portacaval shunt followed by ligation of the hepatic arteries. Biopsies were harvested from three different areas of the brain (frontal lobe, cerebellum, and brain stem) following eight hours of ALF and analyzed using electron microscopy. Results. Profound perivascular and interstitial edema were found in all three areas. Disruption of pericytic and astrocytic processes were seen, reflecting breakdown/lesion of the blood-brain barrier in animals suffering from ALF. Furthermore, neurons and axons were edematous and surrounded by vesicles. Severe damage to Purkinje neuron (necrosis) and damaged myelin were seen in the cerebellum and brain stem, respectively. Biopsies from sham operated animals were normal. Conclusions. Our data support the concept that vasogenic brain edema plays an important role in the development of intracranial hypertension in pigs with ALF.

Ammonia and proinflammatory cytokines modify expression of genes coding for astrocytic proteins implicated in brain edema in acute liver failure

There is evidence to suggest that, in acute liver failure (ALF), brain ammonia and proinflammatory cytokines may act synergistically to cause brain edema and its complications (intracranial hypertension, brain herniation). However, the molecular mechanisms involved remain to be established. In order to address this issue, semi-quantitative RT-PCR was used to measure the expression of genes coding for astrocytic proteins with an established role in cell volume regulation in cerebral cortical astrocytes exposed to toxic agents previously identified in experimental and clinical ALF. Such agents include ammonia, the proinflammatory cytokine interleukin-1beta (IL-1beta) and combinations of the two. Exposure of cultured astrocytes to recombinant IL-1beta (but not ammonia) resulted in increased expression of aquaporin-4 (AQP-4). Both ammonia and proinflammatory mediators led to decreased expression of glial fibrillary acidic protein (GFAP), a cytoskeletal protein, but these effects were not additive. On the other hand, heme oxygenase-1 (HO-1) and inducible nitric oxide synthase (iNOS) expression were significantly increased by exposure to both ammonia and proinflammatory mediators and although modest, these effects were additive suggestive of a synergistic mechanism. These findings suggest that worsening of brain edema and its complications in ALF due to proinflammatory mechanisms may result from exacerbation of oxidative stress-related mechanisms rather than upregulation of AQP-4 or decreases in expression of the astrocytic structural protein GFAP.

Brain Edema in Acute Liver Failure: Inhibition by L-Histidine

Brain edema and the associated increase in intracranial pressure are potentially lethal complications of acute liver failure (ALF). Astrocyte swelling (cytotoxic edema) represents a significant component of the brain edema in ALF, and elevated blood and brain ammonia levels have been strongly implicated in its formation. We earlier showed in cultured astrocytes that oxidative stress (OS) and the mitochondrial permeability transition (mPT) play major roles in the mechanism of ammonia-induced astrocyte swelling. Glutamine, a byproduct of ammonia metabolism, has also been shown to induce OS, the mPT, and astrocyte swelling. Such effects of glutamine were suggested to be mediated by its hydrolysis in mitochondria, potentially yielding high levels of ammonia in this organelle and leading to OS and the mPT. L-histidine, an inhibitor of mitochondrial glutamine transport, was recently shown to mitigate OS, mPT, and cell swelling in cultured astrocytes treated with ammonia. The present study examined whether L-histidine similarly abolishes OS, the mPT, and brain edema in a rat model of ALF. Treatment of rats with thioacetamide caused a significant degree of brain edema, which was associated with induction of OS and the mPT. These changes were completely abolished by L-histidine, supporting a key role of mitochondrial glutamine transport and hydrolysis in the mechanism of the brain edema associated with ALF.

This Month in Gastroenterology

This Month in Gastroenterology

IL-1 or TNF receptor gene deletion delays onset of encephalopathy and attenuates brain edema in experimental acute liver failure

Previous reports suggested that brain-derived proinflammatory cytokines are involved in the pathogenesis of hepatic encephalopathy (HE) and brain edema in acute liver failure (ALF). To further address this issue, expression of interleukin-1beta (IL-1β), tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) mRNAs were measured in the brains of mice with acute liver failure resulting from exposure to azoxymethane. In addition, time to severe encephalopathy (coma) was assessed in mice lacking genes coding for interferon-gamma, the tumor necrosis factor receptor-1 or the interleukin-1 type 1 receptor. Interleukin-1beta, tumor necrosis factor-alpha and interferon-gamma expression were quantified using RT-PCR. Significant increases in interleukin-1beta and tumor necrosis factor-alpha mRNA were observed in the frontal cortex of azoxymethane-treated wild-type mice at coma stages of encephalopathy. Interferon-gamma, however, could not be detected in the brains of these animals. Onset of severe encephalopathy (coma) and brain edema in ALF mice were significantly delayed in interleukin-1 type 1 receptor or tumor necrosis factor receptor-1 knockout mice. Deletion of the interferon-gamma gene, on the other hand, had no significative effect on the neurological status or brain water content of acute liver failure mice. These results demonstrate that toxic liver injury resulting from exposure to azoxymethane is associated with selective induction of proinflammatory cytokines in the brain and that deletion of tumor necrosis factor receptor-1 or interlukin-1 type 1 receptor delays the onset of coma and brain edema in this model of acute liver failure. These findings further support a role for selective brain-derived cytokines in the pathogenesis of the cerebral complications in acute liver failure and suggest that anti-inflammatory strategies could be beneficial in their prevention.

Diffusion Tensor Imaging Supports the Cytotoxic Origin of Brain Edema in a Rat Model of Acute Liver Failure

Brain edema is a severe complication of acute liver failure (ALF) that has been related to ammonia concentrations. Two mechanisms have been proposed in the pathogenesis: vasogenic edema that is secondary to the breakdown of the blood-brain barrier and cytotoxic edema caused by ammonia metabolites in astrocytes. We applied magnetic resonance techniques to assess the intracellular or extracellular distribution of brain water and metabolites in a rat model of devascularized ALF. The brain water content was assessed by gravimetry and blood-brain barrier permeability was determined from the transfer constant of (14)C-labeled sucrose. Rats with ALF had a progressive decrease in the apparent diffusion coefficient (ADC) in all brain regions. The average decrease in ADC was significant in precoma (-14%) and coma stages (-20%). These changes, which indicate an increase of the intracellular water compartment, were followed by a significant increase in total brain water (coma 82.4% +/- 0.3% vs sham 81.6% +/- 0.3%; P = .0001). Brain concentrations of glutamine (6 hours, 540%; precoma, 851%; coma, 1086%) and lactate (6 hours, 166%; precoma, 998%; coma, 3293%) showed a marked increase in ALF that paralleled the decrease in ADC and neurologic outcome. In contrast, the transfer constant of (14)C-sucrose was unaltered. The pathogenesis of brain edema in an experimental model of ALF involves a cytotoxic mechanism: the metabolism of ammonia in astrocytes induces an increase of glutamine and lactate that appears to mediate cellular swelling. Therapeutic measures should focus on removing ammonia and improving brain energy metabolism.

The roles of tumor necrosis factor-alpha in colon tight junction protein expression and intestinal mucosa structure in a mouse model of acute liver failure

BackgroundSpontaneous bacterial peritonitis (SBP) is a common clinical disease and one of the most severe complications of acute liver failure (ALF). Although the mechanism responsible for SBP is unclear, cytokines play an important role. The aim of this study was to investigate the effects of tumor necrosis factor-alpha (TNF-α) on the structure of the intestinal mucosa and the expression of tight junction (Zona Occludens 1; ZO-1) protein in a mouse model of ALF.MethodsWe induced ALF using D-galactosamine/lipopolysaccharide (GalN/LPS) or GalN/TNF-α and assessed the results using transmission electron microscopy, immunohistochemistry, Western blotting, ELISA and real-time quantitative PCR. The effects of administration of anti-TNF-α IgG antibody or anti-TNF-α R1 antibody before administration of GalN/LPS or GalN/TNF-α, respectively, on TNF-α were also assessed.ResultsMorphological abnormalities in the intestinal mucosa of ALF mice were positively correlated with serum TNF-α level. Electron microscopic analysis revealed tight junction (TJ) disruptions, epithelial cell swelling, and atrophy of intestinal villi. Gut bacteria invaded the body at sites where TJ disruptions occurred. Expression of ZO-1 mRNA was significantly decreased in both ALF models, as was the level of ZO-1 protein. Prophylactic treatment with either anti-TNF-α IgG antibody or anti-tumor necrosis factor-a receptor1 (anti-TNF-α R1) antibody prevented changes in intestinal tissue ultrastructure and ZO-1 expression.ConclusionTNF-α affects the structure of the intestinal mucosa, decreases expression of ZO-1, and affects the morphology of the colon in a mouse model of ALF. It also may participate in the pathophysiological mechanism of SBP complicated to ALF.

Therapeutic hypothermia for acute liver failure

Cerebral edema is a potentially life-threatening complication of acute liver failure, the syndrome of abrupt loss of liver function in a patient with a previously healthy liver. Although the prevalence of cerebral edema appears to be decreasing, patients with rapidly progressive (hyperacute) liver failure, such as after acetaminophen overdose, remain at highest risk. In severe cases of cerebral edema, intracranial hypertension develops and leads to brain death after brainstem herniation or to anoxic brain injury and permanent neurologic impairment. Intracranial hypertension in patients with acute liver failure often can be temporarily controlled by manipulating body position, increasing the degree of sedation, and increasing blood osmolarity through pharmacologic means. However, these maneuvers often postpone, but do not eliminate, the risk of brainstem herniation unless orthotopic liver transplantation or spontaneous liver regeneration follows in short order. To buy time, the induction of therapeutic hypothermia (core temperature 32 degrees C-35 degrees C) has been shown to effectively bridge patients to transplant. Similar to the experience in patients with cerebral edema after other neurologic insults, hypothermia reduces cerebral edema and intracranial hypertension in patients with acute liver failure by decreasing splanchnic ammonia production, restoring normal regulation of cerebral hemodynamics, and lowering oxidative metabolism within the brain. Hypothermia may also ameliorate the degree of liver injury. Hypothermia has not been adequately studied for its safety and theoretically may increase the risk of infection, cardiac dysrhythmias, and bleeding, all complications independently associated with acute liver failure. Therefore, although an ample body of experimental and human data provides a rationale for the use of therapeutic hypothermia in patients with acute liver failure, multicenter, randomized, controlled clinical trials are needed to confirm that hypothermia secures brain viability and improves survival without causing harm.

Minocycline attenuates oxidative/nitrosative stress and cerebral complications of acute liver failure in rats

In the present study, the effects of minocycline on progression of encephalopathy and brain edema in rats with acute liver failure (ALF) resulting from hepatic devascularization were studied in relation to the antioxidant action of the drug. ALF rats were sacrificed at precoma and coma stages of encephalopathy along with their appropriate sham-operated controls. Minocycline-treated ALF rats were sacrificed in parallel with comatose vehicle-treated ALF controls. Microglial activation was assessed using CD11b/c (OX-42) immunohistochemistry. Nitrite/nitrate levels in plasma and brain were measured using the Griess reaction. Expression of nitric oxide synthase (NOS) isoforms and heme oxygenase-1 (HO-1) were measured using real-time quantitative PCR and Western blot analysis. Increased nitrite/nitrate levels were observed in the plasma of ALF rats at coma stage of encephalopathy compared to sham-operated controls. Increased expression of HO-1 mRNA and protein was observed in the frontal cortex of ALF rats at both precoma and coma stages of encephalopathy. Significant increases in expression of endothelial (eNOS) and inducible (iNOS) isoforms of NOS mRNA and protein occurred only at coma stages of encephalopathy accompanied by increased brain nitrite/nitrate concentrations. As expected, minocycline attenuated microglial activation as confirmed by decreased OX-42 immunoreactivity, normalized nitrite/nitrate levels in brain and significantly attenuated HO-1, eNOS and iNOS expression. These results indicate that the beneficial effect of minocycline on the neurological complications of ALF is mediated, at least in part, by reduction of oxidative/nitrosative stress.

Cerebral inflammation contributes to encephalopathy and brain edema in acute liver failure: protective effect of minocycline

Encephalopathy and brain edema are serious complications of acute liver failure (ALF). The precise pathophysiologic mechanisms responsible have not been fully elucidated but it has been recently proposed that microglia-derived proinflammatory cytokines are involved. In the present study we evaluated the role of microglial activation and the protective effect of the anti-inflammatory drug minocycline in the pathogenesis of hepatic encephalopathy and brain edema in rats with ALF resulting from hepatic devascularisation. ALF rats were killed 6 h after hepatic artery ligation before the onset of neurological symptoms and at coma stages of encephalopathy along with their appropriate sham-operated controls and in parallel with minocycline-treated ALF rats. Increased OX-42 and OX-6 immunoreactivities confirming microglial activation were accompanied by increased expression of interleukins (IL-1beta, IL-6) and tumor necrosis factor-alpha (TNF-alpha) in the frontal cortex at coma stage of encephalopathy in ALF rats compared with sham-operated controls. Minocycline treatment prevented both microglial activation as well as the up-regulation of IL-1beta, IotaL-6 and TNF-alpha mRNA and protein expression with a concomitant attenuation of the progression of encephalopathy and brain edema. These results offer the first direct evidence for central proinflammatory mechanisms in the pathogenesis of brain edema and its complications in ALF and suggest that anti-inflammatory agents may be beneficial in these patients.

Hypothermia attenuates oxidative/nitrosative stress, encephalopathy and brain edema in acute (ischemic) liver failure

Encephalopathy and brain edema are serious complications of acute liver failure (ALF). The precise pathophysiologic mechanisms responsible have not been fully elucidated but it has been suggested that oxidative/nitrosative stress is involved. In the present study we evaluated the role of oxidative/nitrosative stress in the pathogenesis of hepatic encephalopathy and brain edema in rats with ALF resulting from hepatic devascularization. We also studied the effect of hypothermia, a treatment previously shown to delay the progression of encephalopathy and the onset of brain edema, on ALF-induced oxidative stress. ALF rats were sacrificed at precoma and coma stages of encephalopathy along with their appropriate sham-operated controls. Hypothermic ALF rats were sacrificed in parallel with normothermic comatose ALF rats. Nitric oxide production in plasma and brain was assessed indirectly by measuring the level of its stable end products, nitrite/nitrate (NOx), using the Griess reagent. Expression of nitric oxide synthase (NOS) isoforms and heme oxygenase-1 (HO-1) were measured using real-time quantitative PCR, Western blot analysis and immunohistochemistry. Increased nitrite/nitrate levels were observed in the plasma and frontal cortex in ALF rats at coma stage of encephalopathy compared to sham-operated controls. Increased expression of HO-1 protein and mRNA was observed in the frontal cortex of ALF rats at both precoma and coma stages of encephalopathy. Significant increases in expression of endothelial and inducible NOS mRNA isoforms also occurred at precoma and coma stages of encephalopathy. Expression of the neuronal nitric oxide synthase isoform (nNOS) was not altered by ALF. Hypothermia normalized nitrite/nitrate levels in brain and significantly attenuated HO-1, eNOS and iNOS expression. These results suggest that, oxidative/nitrosative stress participates in the pathogenesis of brain edema and its complications in ALF and that the beneficial effect of hypothermia depends in part on its ability to inhibit oxidative/nitrosative stress-related mechanisms.