NADPH oxidase is implicated in the pathogenesis of oxidative phosphorylation dysfunction in mice fed a high-fat diet.

The aim of this study was to evaluate the role of NADPH oxidase (NADPHox) in the pathogenesis of oxidative phosphorylation (OXPHOS) dysfunction as found in mice fed a high-fat diet (HFD). C57BL/6J mice were distributed in four groups: WT/SCD: six wild-type (WT) mice fed a standard chow diet (SCD); WT/HFD, six WT mice fed a HFD; NOX2(-/-)/SCD, six NADPHox-deficient mice on a SCD; (4) NOX2(-/-)/HFD, six NADPHox-deficient mice on a HFD. After 32 weeks, we studied the liver for: histology; OXPHOS complex activity; fully assembled OXPHOS complexes and their subunits; gene expression of OXPHOS subunits; oxidative and nitrosative stress; and oxidative DNA damage. In the liver of WT/HFD mice, we found a significant decreased in the activity of all OXPHOS complexes, in fully assembled complexes, in the amount of OXPHOS subunits, and in gene expression of mitochondrial DNA-encoded subunits. 8-hydroxy-2'-deoxyguanosine was only increased in mitochondrial DNA. The liver of NOX(-/-)/HFD mice showed mild steatosis but no non-alcoholic steatohepatitis (NASH) lesions were found. OXPHOS activity, OXPHOS subunits, and assembly of subunits into OXPHOS complexes were normal in these mice. We conclude that this study shows that NADPH deficiency protects mice from developing OXPHOS dysfunction and NASH caused by a HFD.

In vitro treatment of HepG2 cells with saturated fatty acids reproduces mitochondrial dysfunction found in nonalcoholic steatohepatitis.

Activity of the oxidative phosphorylation system (OXPHOS) is decreased in humans and mice with nonalcoholic steatohepatitis. Nitro-oxidative stress seems to be involved in its pathogenesis. The aim of this study was to determine whether fatty acids are implicated in the pathogenesis of this mitochondrial defect. In HepG2 cells, we analyzed the effect of saturated (palmitic and stearic acids) and monounsaturated (oleic acid) fatty acids on: OXPHOS activity; levels of protein expression of OXPHOS complexes and their subunits; gene expression and half-life of OXPHOS complexes; nitro-oxidative stress; and NADPH oxidase gene expression and activity. We also studied the effects of inhibiting or silencing NADPH oxidase on the palmitic-acid-induced nitro-oxidative stress and subsequent OXPHOS inhibition. Exposure of cultured HepG2 cells to saturated fatty acids resulted in a significant decrease in the OXPHOS activity. This effect was prevented in the presence of a mimic of manganese superoxide dismutase. Palmitic acid reduced the amount of both fully-assembled OXPHOS complexes and of complex subunits. This reduction was due mainly to an accelerated degradation of these subunits, which was associated with a 3-tyrosine nitration of mitochondrial proteins. Pretreatment of cells with uric acid, an antiperoxynitrite agent, prevented protein degradation induced by palmitic acid. A reduced gene expression also contributed to decrease mitochondrial DNA (mtDNA)-encoded subunits. Saturated fatty acids induced oxidative stress and caused mtDNA oxidative damage. This effect was prevented by inhibiting NADPH oxidase. These acids activated NADPH oxidase gene expression and increased NADPH oxidase activity. Silencing this oxidase abrogated totally the inhibitory effect of palmitic acid on OXPHOS complex activity. We conclude that saturated fatty acids caused nitro-oxidative stress, reduced OXPHOS complex half-life and activity, and decreased gene expression of mtDNA-encoded subunits. These effects were mediated by activation of NADPH oxidase. That is, these acids reproduced mitochondrial dysfunction found in humans and animals with nonalcoholic steatohepatitis.

High-fat diet decreases activity of the oxidative phosphorylation complexes and causes nonalcoholic steatohepatitis in mice.

Nonalcoholic fatty liver disease (NAFLD) is the most frequent histological finding in individuals with abnormal liver-function tests in the Western countries. In previous studies, we have shown that oxidative phosphorylation (OXPHOS) is decreased in individuals with NAFLD, but the cause of this mitochondrial dysfunction remains uncertain. The aims of this study were to determine whether feeding mice a high-fat diet (HFD) induces any change in the activity of OXPHOS, and to investigate the mechanisms involved in the pathogenesis of this defect. To that end, 30 mice were distributed between five groups: control mice fed a standard diet, and mice on a HFD and treated with saline solution, melatonin (an antioxidant), MnTBAP (a superoxide dismutase analog) or uric acid (a scavenger of peroxynitrite) for 28 weeks intraperitoneously. In the liver of these mice, we studied histology, activity and assembly of OXPHOS complexes, levels of subunits of these complexes, gene expression of these subunits, oxidative and nitrosative stress, and oxidative DNA damage. In HFD-fed mice, we found nonalcoholic steatohepatitis, increased gene expression of TNFα, IFNγ, MCP-1, caspase-3, TGFß1 and collagen α1(I), and increased levels of 3-tyrosine nitrated proteins. The activity and assembly of all OXPHOS complexes was decreased to about 50-60%. The amount of all studied OXPHOS subunits was markedly decreased, particularly the mitochondrial-DNA-encoded subunits. Gene expression of mitochondrial-DNA-encoded subunits was decreased to about 60% of control. There was oxidative damage to mitochondrial DNA but not to genomic DNA. Treatment of HFD-fed mice with melatonin, MnTBAP or uric acid prevented all changes observed in untreated HFD-fed mice. We conclude that a HFD decreased OXPHOS enzymatic activity owing to a decreased amount of fully assembled complexes caused by a reduced synthesis of their subunits. Antioxidants and antiperoxynitrites prevented all of these changes, suggesting that nitro-oxidative stress played a key role in the pathogenesis of these alterations. Treatment with these agents might prevent the development of NAFLD in humans.

Fecal incontinence in men: causes and clinical and manometric features.

AIM: To determine the causes and characteristics of fecal incontinence in men and to compare these features with those presented by a group of women with the same problem. METHODS: We analyzed the medical history, clinical and manometric data from 119 men with fecal incontinence studied in our unit and compared these data with those obtained from 645 women studied for the same problem. Response to treatment was evaluated after 6 mo of follow-up. RESULTS: Fifteen percent of patients studied in our unit for fecal incontinence were male. Men took longer than women before asking for medical help. Ano-rectal surgery was the most common risk factor for men related to fecal incontinence. Chronic diarrhea was present in more than 40% of patients in both groups. Decreased resting and external anal sphincter pressures were more frequent in women. No significant differences existed between the sexes regarding rectal sensitivity and recto-anal inhibitory reflex. In 17.8% of men, all presenting soiling, manometric findings did not justify fecal incontinence. Response to treatment was good in both groups, as 80.4% of patients improved and fecal incontinence disappeared in 13.2% of them. CONCLUSION: In our series, it was common that men waited longer in seeking medical help for fecal incontinence. Ano-rectal surgery was the major cause of this problem. Chronic diarrhea was a predisposing factor in both sexes. Manometric differences between groups were limited to an increased frequency of hypotony of the external anal sphincter in women. Fecal incontinence was controllable in most patients.

Pioglitazone leads to an inactivation and disassembly of complex I of the mitochondrial respiratory chain.

BACKGROUND: Thiazolidinediones are antidiabetic agents that increase insulin sensitivity but reduce glucose oxidation, state 3 respiration, and activity of complex I of the mitochondrial respiratory chain (MRC). The mechanisms of the latter effects are unclear. The aim of this study was to determine the mechanisms by which pioglitazone (PGZ), a member of the thiazolidinedione class of antidiabetic agents, decreases the activity of the MRC. In isolated mitochondria from mouse liver, we measured the effects of PGZ treatment on MRC complex activities, fully-assembled complex I and its subunits, gene expression of complex I and III subunits, and [3H]PGZ binding to mitochondrial complexes. RESULTS: In vitro, PGZ decreased activity of complexes I and III of the MRC, but in vivo only complex I activity was decreased in mice treated for 12 weeks with 10 mg/kg/day of PGZ. In vitro treatment of isolated liver mitochondria with PGZ disassembled complex I, resulting in the formation of several subcomplexes. In mice treated with PGZ, fully assembled complex I was increased and two additional subcomplexes were found. Formation of supercomplexes CI+CIII2+CIVn and CI+CIII2 decreased in mouse liver mitochondria exposed to PGZ, while formation of these supercomplexes was increased in mice treated with PGZ. Two-dimensional analysis of complex I using blue native/sodium dodecyl sulfate polyacrylamide gel electrophoresis (BN/SDS-PAGE) showed that in vitro PGZ induced the formation of four subcomplexes of 600 (B), 400 (C), 350 (D), and 250 (E) kDa, respectively. Subcomplexes B and C had NADH:dehydrogenase activity, while subcomplexes C and D contained subunits of complex I membrane arm. Autoradiography and coimmunoprecipitation assays showed [3H]PGZ binding to subunits NDUFA9, NDUFB6, and NDUFA6. Treatment with PGZ increased mitochondrial gene transcription in mice liver and HepG2 cells. In these cells, PGZ decreased intracellular ATP content and enhanced gene expression of specific protein 1 and peroxisome-proliferator activated receptor (PPAR)γ coactivator 1α (PGC-1α). CONCLUSIONS: PGZ binds complex I subunits, which induces disassembly of this complex, reduces its activity, depletes cellular ATP, and, in mice and HepG2 cells, upregulates nuclear DNA-encoded gene expression of complex I and III subunits.

Sp1 and Sp3 transcription factors mediate leptin-induced collagen α1(I) gene expression in primary culture of male rat hepatic stellate cells.

Mechanisms by which leptin stimulates collagen α(1)(I) [Col1a(I)] gene expression are unclear. The purposes of this study were to identify the trans-acting factors and cis-acting elements in Col1a(I) promoter involved in this effect as well as the pathways that are implicated. In primary cultures of rat hepatic stellate cells (HSCs), we measured the effects of leptin on Col1a(I) gene and protein expression and on the binding of nuclear proteins to the Col1a(I) promoter. We found that leptin increased Col1a(I) gene and protein expression in activated HSCs. Transient transfections showed that leptin exerted its effects through elements located between -220 and -112 bp of the Col1a(I) promoter. Gel retardation assays demonstrated that leptin induced the binding of transcription factors specific protein (Sp)-1 and Sp3 to two elements located between -161 and -110 bp of the Col1a(I) promoter. Leptin-induced Sp1/Sp3 phosphorylation, but this effect was suppressed by inhibiting or silencing Janus kinase-2, phosphatidylinositol-3-kinase, nonphagocytic adenine dinucleotide phosphate (NADPH) oxidase, or ERK1/2, by the use of antioxidants or catalase, or by preventing protein-aldehyde adduct formation. Leptin provoked oxidative stress, aldehyde-protein adduct formation, and increased gene expression of some components of the NADPH oxidase complex. In conclusion, in HSCs, leptin up-regulates Col1a(I) gene expression after activating NADPH oxidase, inducing oxidative stress, aldehyde-protein adduct formation, and ERK1/2 phosphorylation, which in turn activates Sp1/Sp3 and provokes the binding of these two factors to regulatory elements located between -161 and -110 bp of the Col1a(I) promoter. These findings may contribute to a better understanding of mechanisms involved in the leptin-induced liver fibrosis.

Melatonin improves mitochondrial respiratory chain activity and liver morphology in ob/ob mice.

In previous studies, we have shown that mitochondrial respiratory chain (MRC) activity is decreased in patients with nonalcoholic steatohepatitis and in ob/ob mice and that peroxynitrite plays a pathogenic role. The present study examined whether melatonin, a peroxynitrite scavenger, prevents: (i) the in vitro effects of peroxynitrite on normal mitochondrial proteins and (ii) the development of nonalcoholic liver disease, MRC dysfunction and proteomic changes found in the mitochondrial complexes from ob/ob mice. We studied MRC activity, assembly of mitochondrial complexes and its subunits in normal mitochondrial proteins exposed to peroxynitrite in the absence and presence of melatonin. The same studies were done in mitochondrial proteins from ob/ob mice untreated and treated with melatonin. Preincubation of mitochondrial proteins from wild-type mice with melatonin prevented 3-tyrosine nitration of these proteins, eliminated the reduction in the MRC activity, the defect in the assembly of mitochondrial complexes and degradation of their subunits induced by peroxynitrite in vitro. Moreover, treatment of ob/ob mice with 10 mg/kg/day melatonin for 12 wk reduced oxidative and nitrosative stress, prevented the loss of MRC activity, protected their complexes and subunits from degradation, and favored assembling of mitochondrial complexes. In addition, this treatment improved fatty liver, decreased hepatic triglyceride concentration and increased apolipoprotein B100 in liver tissue. In conclusion, melatonin prevents the effects of peroxynitrite on mitochondrial proteins in vitro and administration of melatonin to ob/ob mice normalizes liver morphology, mitochondrial dysfunction and assembly of MRC complexes.

Calidad de vida en pacientes con incontinencia fecal y su relación con la gravedad de la misma / Quality of life and its association with the severity of fecal incontinence

Introducción La escala de Wexner es una de las más utilizadas para evaluar la gravedad de la incontinencia fecal (IF). El SF-36 y la escala específica, Fecal incontinence quality of life scale (FIQLS) son instrumentos validados en castellano para el estudio de la calidad de vida.ObjetivosEvaluar la correlación entre la gravedad de la IF y la afectación de la calidad de vida mediante las escalas SF-36 y FIQL.Material y métodosSetenta y tres pacientes con IF completaron las escalas de Wexner, SF-36 y FIQL. El punto de corte de gravedad, según lo publicado en estudios previos, fue 9 (Wexner ≥9: IF grave; Wexner <9: IF leve).ResultadosLa puntuación en las subescalas del SF-36 fueron más bajas que en la población general (p<0,001). El componente mental del SF-36 se correlacionó con los dominios «conducta» y «vergüenza» de FIQLS (p<0,05). La puntuación de Wexner se correlacionó con todos los dominios de FIQLS (p<0,01). Los resultados para todos los dominios de FIQLS fueron menores para el grupo Wexner ≥9 comparado con el grupo Wexner <9 (p<0,001). No se encontró correlación entre la puntuación de Wexner y el SF-36. La edad, IF de urgencia, frecuencia diaria, co-morbilidad y cirugía ano-rectal empeoran distintos aspectos de la calidad de vida en la escala FIQL.ConclusionesLos pacientes con IF tienen gran afectación de la calidad de vida medida con las escalas SF-36 y FIQL. La gravedad de la IF se correlaciona con la escala FIQL. Otros factores como la edad, el tipo y la frecuencia de la IF deben considerarse al evaluar la calidad de vida (AU)

Background Wexner's score is widely used to assess the severity of fecal incontinence (FI). The 36-item short form health survey (SF-36) and the Fecal Incontinence Quality of Life Scale (FIQLS), a disease-specific scale, are instruments measuring quality of life that have been validated into Spanish.AimTo evaluate quality of life in patients with FI by using the FIQL and SF-36 scales to correlate the results with the Wexner's score.MethodsWexner's score, SF-36 and FIQLS were completed by 73 patients. The cut-off point for severity, as previously published, was 9 (Wexner's score <9=mild FI; Wexner's score ≥9=severe FI).ResultsThe scores for all SF-36 sub-scales were lower in patients with FI than in the healthy Spanish population (p<0.001). The mental component of the SF-36 correlated with the subscales of «behavior» and «embarrassment» of the FIQLS (p<0.05). Wexner's score correlated with all the domains of the FIQLS (p<0.01). The results for all the domains of the FIQLS were lower for the Wexner ≥ 9 group than for the Wexner <9 group (p<0.001). There was no significant correlation between Wexner's score and the SF-36. Age, urge and daily FI, comorbidity and anorectal surgery worsened different aspects of quality of life measured by the FIQLS.ConclusionsQuality of life measured by the SF-36 and FIQLS was substantially impaired in patients with FI. The severity of FI correlated with FIQLS. Other factors such as age, type and frequency of FI should be considered when evaluating quality of life (AU)

[Quality of life and its association with the severity of fecal incontinence]. / Calidad de vida en pacientes con incontinencia fecal y su relación con la gravedad de la misma.

BACKGROUND: Wexner's score is widely used to assess the severity of fecal incontinence (FI). The 36-item short form health survey (SF-36) and the Fecal Incontinence Quality of Life Scale (FIQLS), a disease-specific scale, are instruments measuring quality of life that have been validated into Spanish. AIM: To evaluate quality of life in patients with FI by using the FIQL and SF-36 scales to correlate the results with the Wexner's score. METHODS: Wexner's score, SF-36 and FIQLS were completed by 73 patients. The cut-off point for severity, as previously published, was 9 (Wexner's score <9=mild FI; Wexner's score ≥9=severe FI). RESULTS: the scores for all SF-36 sub-scales were lower in patients with FI than in the healthy Spanish population (p<0.001). The mental component of the SF-36 correlated with the subscales of «behavior¼ and «embarrassment¼ of the FIQLS (p<0.05). Wexner's score correlated with all the domains of the FIQLS (p<0.01). The results for all the domains of the FIQLS were lower for the Wexner ≥ 9 group than for the Wexner <9 group (p<0.001). There was no significant correlation between Wexner's score and the SF-36. Age, urge and daily FI, comorbidity and anorectal surgery worsened different aspects of quality of life measured by the FIQLS. CONCLUSIONS: quality of life measured by the SF-36 and FIQLS was substantially impaired in patients with FI. The severity of FI correlated with FIQLS. Other factors such as age, type and frequency of FI should be considered when evaluating quality of life.

Mitochondrial complex I subunits are decreased in murine nonalcoholic fatty liver disease: implication of peroxynitrite.

We investigate the cause of the low activity of mitochondrial complex I found in ob/ob mice with nonalcoholic fatty liver disease. In mitochondrial proteins from ob/ob mice, we assessed complex I activity, fully assembled complex I, and its subunits, oxygen consumption, gene expression of complex I subunits, and oxidative damage to DNA. In mitochondrial proteins from the liver of ob/ob mice, complex I activity, fully assembly of this complex and complex I subunits were markedly reduced. Likewise, gene expression of mitochondrial DNA-encoded subunits was significantly decreased in obese mice, but not nuclear DNA-encoded subunits. Treatment of obese mice with uric acid, anti-TNFalpha antibody or a mimic of manganese superoxide dismutase normalized all these abnormalities. "In vitro" addition of peroxynitrite to mitochondrial proteins from wild-type mice reproduced the abnormalities found in ob/ob mice (decreased complex I activity, the amount of fully assembled complex I, and its subunits, and mitochondrial oxygen consumption). Low activity of complex I found in ob/ob mice can be ascribed to a reduced amount of fully assembled complex, which may be attributed to degradation and reduced synthesis of its subunits by peroxynitrite. Exposure of mitochondrial proteins from normal mice to peroxynitrite reproduced the proteomic abnormalities present in ob/ob mice.

Fibronectin increases survival of rat hepatic stellate cells--a novel profibrogenic mechanism of fibronectin.

UNLABELLED: The aims of this study were to determine whether fibronectin increases survival of hepatic stellate cells (HSCs) in starving conditions, and to identify the signal transduction pathways involved in this effect. METHODS: Primary culture of rat HSCs were plated on fibronectin-uncoated or coated culture wells, and grown in the presence of 0.2% or 20% fetal calf serum. Cell apoptosis was measured by an ELISA procedure. Signal transduction pathways were analyzed by inhibiting major intracellular transduction pathways with appropriated inhibitors and by detecting phosphorylated proteins. RESULTS: Fibronectin increased survival of serum deprived HSCs. This effect was abrogated by the presence of the RGD peptide, by silencing FAK expression, and by inhibiting PI3K with LY294002 or wortmannin. Growth of HSCs on fibronectin induced integrin alpha5beta1 expression, tyr397, ser473, and ser136 phosphorylation of FAK, Akt, and Bad, respectively, and the binding of phosphorylated Bad to 14-3-3 proteins. Likewise, fibronectin increased Bcl2/Bax ratio and reduced release of mitochondrial cytochrome c into the cytoplasm, formation of apoptosome, and caspase 9 and 3 activity. These effects were avoided by treatment of cells with PI3K inhibitors. CONCLUSION: Fibronectin increases survival of HSCs via a pathway involving integrin alpha5beta1 receptors, FAK, PI3K, Akt and proteins of Bcl2 family.

Interferon alpha increases metalloproteinase-13 gene expression through a polyomavirus enhancer activator 3-dependent pathway in hepatic stellate cells.

BACKGROUND/AIMS: To determine the effects of IFNalpha on MMP-13 gene expression in primary culture of hepatic stellate cells. METHODS: We measured MMP-13 mRNA, MMP-13 protein, MMP-13 luciferase activity, binding of AP1 and PEA3 to DNA, and binding of PEA3 to Jak1 and Stat1. RESULTS: IFNalpha increased MMP-13 mRNA, MMP-13 protein, and luciferase activity in cells transfected either with a luciferase plasmid driven by the MMP-13 promoter or with the same plasmid in which the AP1 binding site has been mutated. IFNalpha induced the binding of nuclear proteins to a radiolabeled PEA3 probe, but not to a AP1 probe. Supershift assays demonstrated that PEA3 and Stat1 are implicated in the formation of this complex. Immunoprecipitation assays showed that PEA3 interacts physically with Stat1 and that IFNalpha treatment increases this interaction. Downregulation of PEA3 or JAK1 with appropriated siRNAs or mutation of the PEA3 binding site in the MMP-13 promoter abrogated the effects of IFNalpha on MMP-13 gene expression. Finally, IFNalpha induced the binding of PEA3 to JAK1, as well as PEA3 tyrosine and serine phosphorylation. CONCLUSIONS: IFNalpha determines the binding of PEA3 to JAK1 and its tyrosine phosphorylation. Activated PEA3 binds to MMP-13 promoter and activates its expression.

Effects of rosiglitazone on the liver histology and mitochondrial function in ob/ob mice.

UNLABELLED: Insulin resistance is present in almost all patients with nonalcoholic steatohepatitis (NAFLD), and mitochondrial dysfunction likely plays a critical role in the progression of fatty liver into nonalcoholic steatohepatitis. Rosiglitazone, a selective ligand of peroxisome proliferator-activated receptor gamma (PPARgamma), is an insulin sensitizer drug that has been used in a number of insulin-resistant conditions, including NAFLD. The aim of this study was to analyze the effects of rosiglitazone on the liver histology and mitochondrial function in a model of NAFLD. All studies were carried out in wild-type and leptin-deficient (ob/ob) C57BL/6J mice. Ob/ob mice were treated with 1 mg/kg/day, and activity of mitochondrial respiratory chain (MRC), beta-oxidation, lipid peroxidation, glutathione content in mitochondria, and 3-tyrosine-nitrated proteins in mitochondria were measured. In addition, histological and ultrastructural changes induced by rosiglitazone were also noted. Rosiglitazone treatment increased liver steatosis, particularly microvesicular steatosis. In these animals, mitochondria were markedly swollen with cristae peripherally placed. In ob/ob mice, this drug increased PPARgamma protein expression and lipid peroxide content in liver tissue and decreased glutathione concentration in mitochondria. Rosiglitazone suppressed the activity of complex I of the MRC in ob/ob mice, but did not affect beta-oxidation. 3-Tyrosine nitrated mitochondrial proteins, significantly increased in ob/ob mice, were not modified by rosiglitazone treatment. CONCLUSION: Treatment of ob/ob mice with rosiglitazone did not reverse histological lesions of NAFLD or improve MRC activity. On the contrary, rosiglitazone reduced activity of complex I and increased oxidative stress and liver steatosis.

Outcome of acute idiosyncratic drug-induced liver injury: Long-term follow-up in a hepatotoxicity registry.

A chronic adverse reaction may occur in some instances of drug-induced liver injury (DILI), even despite drug cessation. In our study, we obtained records from a Spanish registry and evaluated cases of DILI with biochemical evidence of long-term damage. Chronic outcome was defined as a persistent biochemical abnormality of hepatocellular pattern of damage more than 3 months after drug withdrawal or more than 6 months after cholestatic/mixed damage. Data on 28 patients with a chronic clinical evolution (mean follow-up 20 months) between November 1995 and October 2005 were retrieved (18 female; overall mean age 55 yr) and accounted for 5.7% of total idiosyncratic DILI cases (n = 493) submitted to the registry. The main drug classes were cardiovascular and central nervous system (28.5% and 25%, respectively), which, in contrast, represented only 9.8% and 13%, respectively, of all DILI cases. The most frequent causative drugs were amoxicillin-clavulanate (4 of 69 cases), bentazepam (3 of 7 cases), atorvastatin (2 of 7 cases), and captopril (2 of 5 cases). Patients with cholestatic/mixed injury (18 of 194 cases [9%]) were more prone to chronicity than patients with hepatocellular injury (10 of 240 cases; P < .031). In the case of chronic hepatocellular injury, 3 patients progressed to cirrhosis and 2 to chronic hepatitis. In the cholestatic/mixed group, liver biopsy indicated cirrhosis in 1 patient and ductal lesions in 3 patients. In conclusion, cholestatic/mixed type of damage is more prone to become chronic while, in the hepatocellular pattern, the severity is greater. Cardiovascular and central nervous system drugs are the main groups leading to chronic liver damage.

Uric acid and anti-TNF antibody improve mitochondrial dysfunction in ob/ob mice.

The mechanisms responsible for low mitochondrial respiratory chain (MRC) activity in the liver of patients with nonalcoholic steatohepatitis are unknown. In this study, we examined the cause of this dysfunction in ob/ob mice. Forty-six mice were distributed in six groups: group I: C57BL/6J mice; group II: C57BL/6J Lep(-/-) mice (ob/ob); group III, ob/ob mice treated with manganese [III] tetrakis (5,10,15,20 benzoic acid) porphyrin (MnTBAP); group IV, ob/ob mice treated with IgG1 immunoglobulin; group V, ob/ob mice treated with anti-TNF antibody; group VI: ob/ob mice treated with uric acid. In liver tissue, we measured MRC activity, fatty acid beta-oxidation, tumor necrosis factor (TNF), inducible nitric oxide synthase (iNOS), 3-tyrosine-nitrated proteins, 3-tyrosine-nitrated mitochondrial proteins, including cytochrome c and ND4 subunit of complex I. MRC activity was decreased in ob/ob mice. TNF levels, iNOS protein expression, and tyrosine nitrated proteins were markedly increased in the liver of ob/ob mice. In these animals, mitochondrial proteins were markedly tyrosine nitrated, particularly the ND4 subunit of complex I and cytochrome c. Treatment of these animals with uric acid, a peroxynitrite scavenger, anti-TNF antibody, or MnTBAP decreased tyrosine nitrated proteins, improved the activity of MRC complexes, and led to a marked regression of hepatic steatosis and inflammation. In conclusion, MRC dysfunction and liver lesions found in ob/ob mice are likely to reflect the tyrosine nitration of mitochondrial proteins by peroxynitrite or a peroxynitrite-derivate radical. Increased hepatic TNF and iNOS expression might enhance peroxynitrite formation and inhibition of MRC complexes.

Interleukin-6 increases rat metalloproteinase-13 gene expression through Janus kinase-2-mediated inhibition of serine/threonine phosphatase-2A.

Interleukin-6 (IL-6) increases metalloproteinase-13 (MMP-13) gene expression by increasing phosphorylated c-Jun and by inhibiting serine/threonine phosphatase-2A (PP2A) activity. We investigated the mechanisms by which IL-6 induces c-Jun phosphorylation and PP2A inactivation in Rat-1 fibroblasts. We show that IL-6 increased MMP-13 mRNA, phosphorylated c-Jun, and activator protein 1 (AP1) binding activity without increasing c-Jun-N-terminal kinase (JNK) activity. These effects did not seem to be mediated by ERK, p38 MAP kinase, phosphatidylinositol-3-kinase, calmoduline-dependent protein kinase, protein kinase C (PKC) or protein kinase A since inhibition with specific inhibitors did not abrogate these effects. IL-6 increases PP2A catalytic subunit tyrosine phosphorylation. Inhibition of the tyrosine kinase Jak2, with the specific inhibitor AG490, abrogated this effect. Likewise, this Jak2 inhibitor blocked the effects of IL-6 on c-Jun phosphorylation, AP1 binding activity and metalloproteinase-13 gene expression. We conclude that IL-6 increases MMP-13 gene expression by activation of Jak2, resulting in tyrosine phosphorylation of the catalytic subunit of PP2A, which in turn decreases PP2A activity and prolongs c-Jun phosphorylation.