Volume 44, Issue 2, Pages 368-374 (February 2006) Rat HMGCoA reductase activation in thioacetamide-induced liver injury is related to an increased reactive oxygen species content  Valentina Pallottini, Chiara Martini, Anna M. Bassi, Paola Romano, Giorgio Nanni, Anna Trentalance  Journal of Hepatology  Volume 44, Issue 2, Pages 368-374 (February 2006) DOI: 10.1016/j.jhep.2005.06.011 Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

Fig. 1 ROS content in chronic and acute TAA-treated rat liver. ROS were detected by fluorescence measurements incubating the liver homogenates from chronic (Ch.) and acute (Ac.) TAA-treated samples with 10μM dichlorodihydrofluorescein diacetate for 30min at 37°. Peroxidized or oxidized molecules were assessed by the initiation of a Fenton-type reaction by the addition of FeSO4 (10μM). For details see the text. Statistical analysis is relative to four different experiments in duplicate. *P<0.001 as from a Student's t-test with respect to its own control (basal fluorescence); #P<0,001 as from a Student's t-test with respect to the control (C). Journal of Hepatology 2006 44, 368-374DOI: (10.1016/j.jhep.2005.06.011) Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

Fig. 2 HMGCoAR activity in chronic and acute TAA-treated rat liver. Results are shown for chronic (Ch.) and acute (Ac.) TAA-treated rat liver. HMGCoAR activity was measured following the conversion of [14C]-HMG coenzyme A to [14C]-mevalonate on microsomes prepared in the presence or absence of 50mM NaF, respectively representing the expressed and the total enzyme activity. Statistical analysis is relative to four different experiments in duplicate. *P<0.001 as from a Student's t-test with respect to +NaF samples of the control (C); # P<0.005 as from a Student's t-test with respect to −NaF samples of the control (C). Journal of Hepatology 2006 44, 368-374DOI: (10.1016/j.jhep.2005.06.011) Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

Fig. 3 HMGCoA reductase levels. Western blots showing HMGCoAR protein levels in liver microsomes from chronic (Ch.) and acute (Ac.) TAA-treated rats. Proteins from 20μg solubilised tissue were resolved by SDS-PAGE, and subsequently transferred electrophoretically onto nitrocellulose and probed with anti-HMGCoAR antiserum. Panel (a) shows typical western blotting. Panel (b) shows the densitometric analysis obtained from four different experiments. *P<0.001 as from a Student's t test with respect to control (C). Journal of Hepatology 2006 44, 368-374DOI: (10.1016/j.jhep.2005.06.011) Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

Fig. 4 HMGCoAR degradation rate in chronic and acute TAA treated rat liver. Western blots showing HMGCoAR degradation rate from chronic (Ch.) (a) and acute (Ac.) (b) TAA-treated rat liver. The reductase degradation was performed on tissue lysates at 37°C for the times indicated in a buffer (pH =7.5) containing Tris-HCl 10 mM and sucrose 75 mM; the HMGCoAR levels were detected by Western blotting. The experiment were performed four times. The panels represent a typical western blotting. Journal of Hepatology 2006 44, 368-374DOI: (10.1016/j.jhep.2005.06.011) Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

Fig. 5 AMPK activation state and PP2A catalytic subunit level in chronic and acute TAA treated rat liver. Panel (a) represents the AMPK activation state, panel (b) the PP2A cathalytic subunit levels of chronic and acute TAA-treated rat liver. Proteins from 20μg solubilised tissue were resolved by SDS-PAGE. The proteins were subsequently transferred electrophoretically onto nitrocellulose and probed with anti-AMPK-P antibody, then stripped and probed with anti-AMPK antibody. The same samples were resolved by SDS-PAGE, transferred electrophoretically onto nitrocellulose and probed with anti PP2A catalytic subunit. For details see the text. The panels represent typical western blots and the densitometric analysis was obtained from four different experiments. Journal of Hepatology 2006 44, 368-374DOI: (10.1016/j.jhep.2005.06.011) Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

Fig. 6 Lipid profile of chronic and acute TAA-treated rat liver. The panel a represents the scanner image of a typical TLC plate on which were chromatographed lipid extracts of chronic and acute TAA-treated rat livers. Line 1 Control (C); line 2 Chronic (Ch.); line 3 Acute (Ac.); line 4 cholesterol standard (Chol); line 5 Phospholipids standard (PL); line 6 Fatty Acids standard (FA); line 7 Squalene standard (Sq); line 8 Triglycerides standard (Tr). The panel b shows a table containing the densitometric analysis, expressed in arbitrary units, of the spots from four different experiments. *P<0.001 as from a Student's t-test with respect to control (C). Journal of Hepatology 2006 44, 368-374DOI: (10.1016/j.jhep.2005.06.011) Copyright © 2005 European Association for the Study of the Liver Terms and Conditions