- Email: [email protected]
[721] OVEREXPRESSION OF MANGANESE SUPEROXIDE DISMUTASE TRIGGERS MITOCHONDRIA!. DAMAGE AFTER CHRONIC ALCOHOL INTOXICATION IN MOUSE LIVER
POSTERS
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p=O.O077)and GAHS 39 (80% vs 41% p=O.O143). The difference at 84 days was not significant. Conclusions: The short term (28 days) outcome of patients with apparent AH after stabilisation from a GI bleed appears to be significantly better compared with those who do not present with GI bleeding. Standard clinical criteria for diagnosis of AH may not be applicable in this group and the threshold for treatment of these patients should be different. Patients who present with GI bleeding should be excluded from future AH trials as they are not a comparable group.
17211 OVEREXPRESSION OF MANGANESE SUPEROXIDE DISMUTASE TRIGGERS MITOCHONDRIAL DAMAGE AFTER CHRONIC ALCOHOL INTOXICATION IN MOUSE LIVER I. Larosche’, P. Letteron’, B. Fromenty’, A. Abbey-Toby1, N. Vadrot’, G. Feldmann’, H. Van Remmen2, A. Richardson2, D. Pessayrel , A. Mansouri’ . ‘INSERM U773, Faculti de Midecine Xavier Bichat, Paris, fiance; 2Departnzent of Cellular and Strnctural Biology, University of RJX~XLES Health Science Center at Sun Antonio, Sun Antonio, TX, USA E-mail: [email protected] Background and Aims: Long-term alcohol consumption causes morphologic and functional changes of liver mitochondria due to increased generation of reactive oxygen radicals. Overexpression of the mitochondrial isoform of superoxide dismutase (MnSOD) might therefore alleviate hepatic lesions mediated by oxidative stress, but controversies remain. Indeed, MnSOD overexpression in rat protects against alcohol-induced hepatic lesions whereas in humans a genetic dimorphism increasing mitochondrial MnSOD activity is a risk factor to develop alcoholic liver diseases. The aim of the present study was therefore to determine whether overexpression of MnSOD affects mitochondrial damage in a mouse model of chronic ethanol exposure. Methods: Wild type MnSOD+’+ mice (WT) and overexpressing transgenic MnSOD+++ mice (TG) were fed ethanol for 7 weeks (20% in drinking water) and hepatic mitochondrial damage was evaluated. Results: Chronic ethanol feeding resulted in decreased hepatic mitochondrial DNA (mtDNA) content and increased mtDNA damage in TG mice, whereas no alterations were observed in WT mice. Using Blue Native-PAGE method, we demonstrated that chronic alcohol consumption decreased the levels and activities of complexes 1 and V in TG mice compared to their respective littermates. Alcohol-induced inhibition of mitochondrial respiration may increase superoxide anion formation leading to an overproduction of hydrogen peroxide (H2O2) in TG mice. In combination with mobilization of iron caused by alcohol, H202 may form the reactive hydroxyl radical (OH.). Hepatic iron was slightly but not significantly elevated in intoxicated TG mice but not in WT mice. Induction and/or inactivation of the antioxidant defense systems may further modulate mitochondrial damage in intoxicated mice. Cu, ZnSOD activity remained unchanged in intoxicated WT and TG mice whereas catalase activity decreased in TG mice fed alcohol. Mitochondria1 GPx-1 activity and GSH were decreased in WT mice fed alcohol but not in TG mice, and MnSOD/GPx-1 ratio was increased in intoxicated TG mice, thus further increasing H202 production. Furthermore, intoxicated TG mice showed an elevated content of carbonylated hepatic proteins, which may further impair respiratory chain function. Conclusions: Mice overexpressing MnSOD are more prone to mtDNA damage after a chronic alcohol exposure, resulting in impairment of mtDNA-encoded respiratory chain complexes thus leading to increased mitochondrial oxidative stress.
17221 ASSESSMENT UTILITY OF THE ROUSSEL UCLAF CAUSALITY METHOD (RUCAM) IN THE EVALUATION OF HEPATOTOXICITY DISCOVERED DURING CLINICAL TRIALS - THE CASE OF XIMELAGATRAN D. Larrev’, J.H. Lewis2, R. Olsson3, WM. Lee4, L. Frison5, M. Keisu‘. ’Lioer linit, INSERM U632, Montpellier School q f Medicine,
St Eloi Hospital, Montpellier, France; ’Gastroenterology, Hepatology Depwtnzent, Georgetown University Medical Centel; Washington, USA; 3 Hepatology, Sahlgrenska Unioersity Hospital, Gothenhnrg, Sweden; 4Hepatology, liniversity of Texas Southwestern Medical Centev, Dallas, TX, USA; -5Researchand Development, AstraZeneca, Molndal, Sweden E-mail: dom-larreyejchu-montpellier.fr Introduction: RUCAM was developed >20 years ago as a means to more objectively assess possible hepatotoxicity in marketed drugs. It has been used increasingly for this purpose worldwide, although up to nownever to evaluate a single drug still in development. Aim: RUCAM was applied to review potentially serious adverse hepatobiliary events that occurred during the phase 2 and 3 long-term clinical trials of ximelagatran (Exanta, AstraZeneca, Molndal, Sweden). Four hepatologists independently reviewed and scored instances of ALT > IOxULN; ALT > 3 x ULN plus bilirubin > 1.5x ULN; any bilirubin > 3 x ULN; first ALT > 3 xULN after 6 months of therapy and death any time after ALT > 3 xULN. 19816948 ximelagatran recipients and 35/6230 control patients were scored. RUCAM score concordance between assessors was determined for the first 50 cases and compared to the last 50. Results: causality classification: highly probable O%, possible or probable 64%, unlikely and excluded 36% (vs 91% in comparator cases). Overall, 86% of cases judged to be possibleiprobable were hepatocellular in nature. Problems and ambiguities in the RUCAM were encountered. For the first 50 patients, learning period was observed with score differences between the assessors leading to review discussion for a final score. The concordance improved throughout the course of investigation and was better for the last 50 assessments vs the first 50 (kappa statistic 0.56-0.79 vs 0.37-0.74). Variability factors were mainly the points awarded for age >55 yr and the evaluation of alcohol consumption impact. Other difficulties were: score reduction for latency period >90 days (whereas liver injury mostly occurred between 90-1 80 days); no point for transient ALT elevation which could nevertheless be caused by the drug; no point for delayed ALT elevation beyond treatment discontinuation; no point for “known liver reaction” whereas trial progression suggested a higher frequency of ALT elevation. Conclusion: The RUCAM is adapted to assess hepatotoxicity of drugs with already lmown liver profile. It remains useful for assessing new agents in clinical trial setting. However, modifications should be considered to enhance its objectivity and accuracy in future studies to fit in with specific compounds.
17231 LIVER MITOCHONDRIAL FUNCTION AND BlOGENESlS IN PATIENTS WITH NAFLD: HEPATIC EXPRESSION OF PGCI-ALPHA, COX2 AND COX4 M. Lemoine’,2, M. Kim2, V. Ratziu3, V. Barbu2, J.P. Bastard2,
D. Wendum4, F. Paye5, R. Poupon1,2, C. Housset2, J. Capeau2, L. Serfaty’,2. ‘Liver linit, Saint-Antoine Hospital, APHF: Paris; ’INSERM U6K0, Facult?, of Medicine, UPMC, Paris; ‘Liuer Unit, Piti& Sulpitridre Hospital, APHR Paris; 4Pathology Department, Saint-Antoine Hospital, APHR Paris; -5SurgeryDepartment, Saint-Antoine Hospital, APHF: Paris, Frunce E-mail: [email protected] NASH is associated with mitochondrial dysfunction. PPAR gamma coactivator l a (PGCla) is involved in mitochondrial biogenesis. The subunit Cox2 of cytochrome c oxidase (Cox) is encoded by the mitochondrial DNA
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p=O.O077)and GAHS 39 (80% vs 41% p=O.O143). The difference at 84 days was not significant. Conclusions: The short term (28 days) outcome of patients with apparent AH after stabilisation from a GI bleed appears to be significantly better compared with those who do not present with GI bleeding. Standard clinical criteria for diagnosis of AH may not be applicable in this group and the threshold for treatment of these patients should be different. Patients who present with GI bleeding should be excluded from future AH trials as they are not a comparable group.
17211 OVEREXPRESSION OF MANGANESE SUPEROXIDE DISMUTASE TRIGGERS MITOCHONDRIAL DAMAGE AFTER CHRONIC ALCOHOL INTOXICATION IN MOUSE LIVER I. Larosche’, P. Letteron’, B. Fromenty’, A. Abbey-Toby1, N. Vadrot’, G. Feldmann’, H. Van Remmen2, A. Richardson2, D. Pessayrel , A. Mansouri’ . ‘INSERM U773, Faculti de Midecine Xavier Bichat, Paris, fiance; 2Departnzent of Cellular and Strnctural Biology, University of RJX~XLES Health Science Center at Sun Antonio, Sun Antonio, TX, USA E-mail: [email protected] Background and Aims: Long-term alcohol consumption causes morphologic and functional changes of liver mitochondria due to increased generation of reactive oxygen radicals. Overexpression of the mitochondrial isoform of superoxide dismutase (MnSOD) might therefore alleviate hepatic lesions mediated by oxidative stress, but controversies remain. Indeed, MnSOD overexpression in rat protects against alcohol-induced hepatic lesions whereas in humans a genetic dimorphism increasing mitochondrial MnSOD activity is a risk factor to develop alcoholic liver diseases. The aim of the present study was therefore to determine whether overexpression of MnSOD affects mitochondrial damage in a mouse model of chronic ethanol exposure. Methods: Wild type MnSOD+’+ mice (WT) and overexpressing transgenic MnSOD+++ mice (TG) were fed ethanol for 7 weeks (20% in drinking water) and hepatic mitochondrial damage was evaluated. Results: Chronic ethanol feeding resulted in decreased hepatic mitochondrial DNA (mtDNA) content and increased mtDNA damage in TG mice, whereas no alterations were observed in WT mice. Using Blue Native-PAGE method, we demonstrated that chronic alcohol consumption decreased the levels and activities of complexes 1 and V in TG mice compared to their respective littermates. Alcohol-induced inhibition of mitochondrial respiration may increase superoxide anion formation leading to an overproduction of hydrogen peroxide (H2O2) in TG mice. In combination with mobilization of iron caused by alcohol, H202 may form the reactive hydroxyl radical (OH.). Hepatic iron was slightly but not significantly elevated in intoxicated TG mice but not in WT mice. Induction and/or inactivation of the antioxidant defense systems may further modulate mitochondrial damage in intoxicated mice. Cu, ZnSOD activity remained unchanged in intoxicated WT and TG mice whereas catalase activity decreased in TG mice fed alcohol. Mitochondria1 GPx-1 activity and GSH were decreased in WT mice fed alcohol but not in TG mice, and MnSOD/GPx-1 ratio was increased in intoxicated TG mice, thus further increasing H202 production. Furthermore, intoxicated TG mice showed an elevated content of carbonylated hepatic proteins, which may further impair respiratory chain function. Conclusions: Mice overexpressing MnSOD are more prone to mtDNA damage after a chronic alcohol exposure, resulting in impairment of mtDNA-encoded respiratory chain complexes thus leading to increased mitochondrial oxidative stress.
17221 ASSESSMENT UTILITY OF THE ROUSSEL UCLAF CAUSALITY METHOD (RUCAM) IN THE EVALUATION OF HEPATOTOXICITY DISCOVERED DURING CLINICAL TRIALS - THE CASE OF XIMELAGATRAN D. Larrev’, J.H. Lewis2, R. Olsson3, WM. Lee4, L. Frison5, M. Keisu‘. ’Lioer linit, INSERM U632, Montpellier School q f Medicine,
St Eloi Hospital, Montpellier, France; ’Gastroenterology, Hepatology Depwtnzent, Georgetown University Medical Centel; Washington, USA; 3 Hepatology, Sahlgrenska Unioersity Hospital, Gothenhnrg, Sweden; 4Hepatology, liniversity of Texas Southwestern Medical Centev, Dallas, TX, USA; -5Researchand Development, AstraZeneca, Molndal, Sweden E-mail: dom-larreyejchu-montpellier.fr Introduction: RUCAM was developed >20 years ago as a means to more objectively assess possible hepatotoxicity in marketed drugs. It has been used increasingly for this purpose worldwide, although up to nownever to evaluate a single drug still in development. Aim: RUCAM was applied to review potentially serious adverse hepatobiliary events that occurred during the phase 2 and 3 long-term clinical trials of ximelagatran (Exanta, AstraZeneca, Molndal, Sweden). Four hepatologists independently reviewed and scored instances of ALT > IOxULN; ALT > 3 x ULN plus bilirubin > 1.5x ULN; any bilirubin > 3 x ULN; first ALT > 3 xULN after 6 months of therapy and death any time after ALT > 3 xULN. 19816948 ximelagatran recipients and 35/6230 control patients were scored. RUCAM score concordance between assessors was determined for the first 50 cases and compared to the last 50. Results: causality classification: highly probable O%, possible or probable 64%, unlikely and excluded 36% (vs 91% in comparator cases). Overall, 86% of cases judged to be possibleiprobable were hepatocellular in nature. Problems and ambiguities in the RUCAM were encountered. For the first 50 patients, learning period was observed with score differences between the assessors leading to review discussion for a final score. The concordance improved throughout the course of investigation and was better for the last 50 assessments vs the first 50 (kappa statistic 0.56-0.79 vs 0.37-0.74). Variability factors were mainly the points awarded for age >55 yr and the evaluation of alcohol consumption impact. Other difficulties were: score reduction for latency period >90 days (whereas liver injury mostly occurred between 90-1 80 days); no point for transient ALT elevation which could nevertheless be caused by the drug; no point for delayed ALT elevation beyond treatment discontinuation; no point for “known liver reaction” whereas trial progression suggested a higher frequency of ALT elevation. Conclusion: The RUCAM is adapted to assess hepatotoxicity of drugs with already lmown liver profile. It remains useful for assessing new agents in clinical trial setting. However, modifications should be considered to enhance its objectivity and accuracy in future studies to fit in with specific compounds.
17231 LIVER MITOCHONDRIAL FUNCTION AND BlOGENESlS IN PATIENTS WITH NAFLD: HEPATIC EXPRESSION OF PGCI-ALPHA, COX2 AND COX4 M. Lemoine’,2, M. Kim2, V. Ratziu3, V. Barbu2, J.P. Bastard2,
D. Wendum4, F. Paye5, R. Poupon1,2, C. Housset2, J. Capeau2, L. Serfaty’,2. ‘Liver linit, Saint-Antoine Hospital, APHF: Paris; ’INSERM U6K0, Facult?, of Medicine, UPMC, Paris; ‘Liuer Unit, Piti& Sulpitridre Hospital, APHR Paris; 4Pathology Department, Saint-Antoine Hospital, APHR Paris; -5SurgeryDepartment, Saint-Antoine Hospital, APHF: Paris, Frunce E-mail: [email protected] NASH is associated with mitochondrial dysfunction. PPAR gamma coactivator l a (PGCla) is involved in mitochondrial biogenesis. The subunit Cox2 of cytochrome c oxidase (Cox) is encoded by the mitochondrial DNA