- Email: [email protected]
680 NVP BEZ235-AN ALONE OR IN COMBINATION WITH CHEMOTHERAPY EFFECTIVELY INHIBITS HEPATOCELLULAR CARCINOMA GROWTH
POSTERS tested, long treatment of TGF-b (2–5 ng/mL for 5 days) significantly blocked the anti-tumor effect of sorafenib. Western blotting showed that sorafenib decreased the levels of phosphorylated ERK, STAT3, PDK1 and mTOR in hepatoma cells, irrespective of with or without additional cytokines. Sorafenib also decreased the level of p38 phosphorylation, but in the cells pre-treated with TGF-b for long time (2–5 ng/mL for 5 days), phospho-p38 remained unchanged. SB203580 reverted the anti-tumor effect of sorafenib in the cells pre-treated with TGF-b, showing the ratio of apoptotic cells almost equal to the cells treated with sorafenib alone. Conclusions: TGF-b is a strong modifier for sorafenib resistance in hepatoma. TGF-b may be a useful biomarker of efficacy prediction of a sorafenib treatment in HCC, and TGF-b/p38 signaling could be a potential therapeutic target for overcoming sorafenib resistance. 678 AMP-ACTIVATED PROTEIN KINASE (AMPK) IS REQUIRED FOR G1/S PROGRESSION DURING LIVER REGENERATION G. Merlen, J.-E. Guidotti, M. Foretz, B. Viollet, C. Desdouets. Institut Cochin, INSERM U1016, CNRS UMR8104, Universit´e Paris Descartes (UMR S1016), Paris, France E-mail: [email protected] Background and Aim: AMPK is a cellular energy sensor that is conserved in eukaryotes. Elevated AMP/ATP ratio activates AMPK, which inhibits energy consuming processes and activates energyproducing processes to restore the energy homeostasis inside the cell. AMPK activators are used for the treatment of type II diabetes. Recently, several research groups have reported that AMPK links energy status to cell structure, cell cycle and mitosis. In this context, our aim is to assess a potential role of AMPK on liver regeneration. Methods: 2/3 partial hepatectomy (PH) was performed in male AMPK catalytic subunit a1−/− and wild type (C57Bl6J) mice (12–14 weeks-old). Animal were sacrified at 7 time points post PH and the remaining livers weighed to calculate the restoration of liver mass. Cell proliferation was determined by immunohistochemistry (BrdU and pHH3 labelling) and by western blot (Cyclins D, E, A, B expression). Cell cycle progression have been also analysed on isolated hepatocytes on primary culture. Results: In AMPK a1−/− mice, hepatocyte proliferation was delayed without altering liver mass regeneration after PH. We observed no difference in the priming phase between mutant and control mice. By contrast, we showed that AMPK a1−/− mice display a drastic impaired of G1/S progression. The percentage of hepatocytes in S phase, 38 hours after hepatectomy, was reduced by 85% in AMPK a1−/− mice compared to wild type controls. We next evaluated component of the cell cycle machinery involved in the progression through G1/S. We discovered that level of cyclin D and cyclin E was equal in both models. However, we observed a significant delay in cyclin A expression. Moreover, we observed the same delay on cell cycle progression on primary cultured hepatocytes. BrdU incorporation showed decreased numbers of positive nuclei, at 36 h, in AMPK a1−/− hepatocytes (43%) compared to controls, correlated with a specific delay in cyclin A expression. Conclusions: Our results suggest that AMPK plays a role in the initiation of S phase during hepatocyte proliferation. Our hypothesis is that a decrease of AMPK activity leads to a reduction of the “energetic information” essential for S phase entry.
679 ACTIVATION OF WNT/b-CATENIN SIGNALING PATHWAY IN THE LIVER OF TRANSGENIC MICE OVEREXPRESSING GROWTH HORMONE J.G. Miquet1 , C.S. Martinez1 , L. Gonzalez ´ 1 , M.E. D´ıaz1 , E. Zotta2 , A. Bartke3 , D. Turyn1 , A.I. Sotelo1 . 1 Department of Biological Chemistry – IQUIFIB, School of Pharmacy and Biochemistry, University of Buenos Aires – CONICET, 2 Department of Physiology, School of Medicine, University of Buenos Aires, Ciudad Autonoma de Buenos Aires, Argentina; 3 Departments of Internal Medicine and Physiology, School of Medicine, Southern Illinois University, Springfield, IL, USA E-mail: [email protected] Background and Aims: Transgenic mice overexpressing growth hormone (GH) exhibit hepatomegaly due to hypertrophy and hyperplasia. Throughout lifespan, transgenic mice present high levels of hepatocellular replication and at advanced ages develop liver tumors, mainly hepatocellular carcinoma (HCC). Wnt/b-catenin signaling regulates cell proliferation, apoptosis and differentiation, and is believed to play a role in carcinogenesis. A high percentage of human HCC show high levels of b-catenin, usually located at the cytoplasm or nucleus rather than at the plasma membrane. Control of b-catenin stability is regulated by GSK-3b, which phosphorylates b-catenin, promoting its degradation. The objective of this work was to evaluate if Wnt/b-catenin signaling is dysregulated in the liver of GH-transgenic mice to assess its possible association with the liver pathology observed in these animals. Methods: Young adult transgenic mice overexpressing GH, which present preneoplastic liver pathology, were studied; non-transgenic siblings were used as controls. Liver samples were subjected to qRT-PCR, Western-blotting and immunohistochemistry to evaluate gene expression, protein content and cellular localization of the mediators under study. Results: b-catenin protein content was 2-fold increased in the liver of GH-overexpressing mice, although its mRNA levels were lower compared to normal siblings (P < 0.002). Immunohistochemical analyses revealed that b-catenin was mainly located in the nucleus or in the cytoplasm in transgenic mice. Transgenic mice presented GSK-3b mRNA levels similar to those observed for control mice, but its protein content was elevated, with a parallel increase in its phosphorylation levels at serine-9, which renders GSK-3b inactive (P < 0.001). The expression of the b-catenin target genes cyclin D1, c-myc and c-jun was increased in transgenic mice liver (P < 0.005). Conclusions: Wnt/b-catenin signaling is dysregulated in the liver of GH-transgenic mice. The increased protein levels and nuclear/ cytoplasmic localization of b-catenin, without an increase in its mRNA levels, suggest that the upregulation of b-catenin may be associated with increased protein stability, in accordance with the overexpression of inactive GSK-3b found in these mice. Elevated levels of inactive GSK-3b were previously reported in HCC harboring b-catenin accumulation. We propose aberrant Wnt/b-catenin signaling may be involved in the preneoplastic liver pathology observed in GH-overexpressing transgenic mice. 680 NVP BEZ235-AN ALONE OR IN COMBINATION WITH CHEMOTHERAPY EFFECTIVELY INHIBITS HEPATOCELLULAR CARCINOMA GROWTH D. Pothiraju, M.P. Manns, A. Vogel. Hannover Medical School, Hannover, Germany E-mail: [email protected] Background and Aims: Hepatocellular carcinoma (HCC) is the fifth most frequent cancer and third most lethal cancer worldwide. Activated PI3K/ AKT/ mTOR pathway has recently emerged as a pivotal contributor to HCC development. Aberrant mTOR signaling has been identified in up to 50–60% of HCC cases, associated
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POSTERS with insulin-like growth factor pathway activation and PTEN dysregulation. The aim of this study was to delineate the role of PI3k and mTOR inhibitor (NVP BEZ235-AN) in HCC. Methods: HuH7 cells were treated with dual inhibitor i.e. AKT & mTOR (NVP BEZ235-AN) inhibitor, Doxorubicin, Cisplatin, 5Fluoro Uracil and Irinotecan alone or in combination for 48hrs. Cell proliferation was measured using MTT assay. Cell cycle (PI staining) and apoptosis (Annexin vs PI) analysis were carried out by FACS. Signaling mechanism of these drugs was confirmed by western blots. In in vivo studies, nude mice were transplanted with Huh7 cells. Tumor size was measured. Tissue pathological analysis was done by HE staining, proliferation was done by IHC using Ki67 antigen and pHis3 anitibody. Apoptosis was measured by Tunel staining. Results: All drugs had an inhibitory effect on cell viability. Combination of BEZ with cisplatin significantly decreased the viability of hepatoma cells compared to Doxorubicin, Irinotecan and 5FU. BEZ treatment inhibits the mTOR phosphorylation and higher concentration of BEZ decreased AKT phosphorylation. Tumor size was significantly decreased in combination treated mice compared to single treated mice. Proliferating cell count was less in treated samples compared to control. Apoptotic cell number was more in treated mice compared to control mice. Conclusion: BEZ effectively induces loss of cell viability. Additionally, BEZ and cisplatin showed synergistic effect on cell proliferation inhibition. Xenograft models confirmed reduced tumor growth in single treatment and moreover, tumor growth was highly regressed in combination of BEZ with cisplatin treatment compared to single treatment. 681 THE COMBINATION OF NEXAVAR AND SIRNA-VEGF INHIBITS THE NEXAVAR-INDUCED INCREASE OF VEGF RESULTING IN ENHANCED ANGIOSTATIC EFFECTS E. Raskopf, T. Sauerbruch, V. Schmitz. Department of Internal Medicine I, University of Bonn, Bonn, Germany E-mail: [email protected] Background and Aims: Nexavar is an established medication in the treatment of advanced HCC and effects on the tumour cell itself are in focus of current research. However, effects on endothelial cells are barely investigated. Therefore, we investigated the effects of Nexavar alone and in combination with siRNA targeting VEGF in human and murine endothelial cells in vitro. Methods: SVEC4–10 and HUVE cells were incubated with different Nexavar-concentrations (0, 1, 2.5, 5 and 10 mM) and analysed for cell viability, proliferation, apoptosis, intracellular signal transduction and VEGF-secretion. In analogy to the mono-therapy, Nexavar was combined with siRNA-VEGF to investigate possible synergistic effects on endothelial cells. Results: Treatment with Nexavar inhibited cell viability in a dose dependent-manner in both cell lines (−85% for HUVE-cells and −90% for SVEC4–10 at 10 mM). Cell proliferation was affected in a similar way. Apoptosis was not influenced under Nexavartreatment. Analysis of intracellular signal transduction showed that phosphorylation of AKT was reduced in a dose-dependent manner. Interestingly, VEGF was increased following Nexavar treatment, peaking at 5 mM Nexavar in both cell lines. This increase was reversed using siRNA targeting VEGF. VEGF-secretion was decreased by 50% compared to the control-siRNA. Furthermore, cell viability was also further decreased (by 30%) after combination of Nexavar with siRNA-VEGF compared to Nexavar alone. Conclusions: Treatment with Nexavar reduced cell viability in both endothelial cell lines, whereas VEGF-levels were increased. This increase was antagonised by siRNA-VEGF leading to an enhanced efficacy of Nexavar. Therefore the combination of both treatment strategies could reduce Nexavar medication and corresponding side effects. S274
682 TAURO-b-MURICHOLIC ACID REDUCES GLYCOCHENODEOXYCHOLIC ACID-INDUCED APOPTOSIS IN A HUMAN HEPATOMA CELL LINE BY RESTORING THE MITOCHONDRIAL MEMBRANE POTENTIAL P. Kleiss1 , G. Denk1 , R. Wimmer1 , H. Zischka2 , C. Rust1 . 1 Department of Medicine 2 – Grosshadern, Ludwig-Maximilians University of Munich, 2 Institute of Toxicology, Helmholtz Zentrum Munich, German Research Center for Environmental Health, Munich, Germany E-mail: [email protected] Introduction and Aim: b-muricholic acid (bMCA) is a natural trihydroxylated bile acid and constitutes the major bile acid synthesized in rat and mouse. Due to the presence of a hydroxyl group in the 6b position of the steroid ring, bMCA is more hydrophilic than ursodeoxycholic acid. In the past, bMCA has been evaluated for the dissolution of cholesterol gallstones (Montet et al., Biochemica et Biopysica Acta 1987: 918: 1–19). However, it is unknown if bMCA has beneficial effects on cell death induced by toxic bile acids. Therefore, our aim was to determine the effect of bMCA on apoptosis induced by glycochenodeoxycholic acid (GCDCA) in a human hepatoma cell line stably transfected with the sodium taurocholate cotransporting poylpetide (HepG2-Ntcp). Methods: Ntcp-transfected HepG2 were stimulated for 4 hours with GCDCA, tauro-b-muricholic acid (TbMCA), tauroursodeoxycholic acid (TUDCA) at a concentration of 25 mM each and DMSO (‰, respectively. Apoptosis was then quantified biochemically in a caspase 3/7-assay as well as morphologically after Hoechst33342 staining. The mitochondrial membrane potential (MMP) was measured fluorometrically using JC-1 (5,5 ,6,6 -tetrachloro-1,1 ,3,3 tetraethyl-benzimidazol-carbocyaniniodide) at a concentration of 2 mM at 2, 4 and 6 hours after stimulation with bile acids (25 mM, 50 mM and 100 mM) alone or in combination. Results: In HepG2-Ntcp cells, GCDCA induced a 18±8-fold increase in apoptosis after 4 hours, respectively, compared to untreated cells as determined in caspase 3/7-assays (n = 6, each). Simultaneous incubation with TbMCA reduced apoptosis to 49% as compared to 44% with TUDCA, both significant differences (p < 0.01 vs. GCDCA, each; n = 6). Similar results were obtained when apoptosis was assessed morphologically by Hoechst-33342 staining. While GCDCA (100mM) reduced the MMP to 58% (2 h), 45% (4 h) and 34% (6 h), combination treatment with TbMCA restored the MMP to control levels at all time points (n = 4). Conclusions: TbMCA significantly reduced hepatocyte apoptosis induced by low micromolar concentrations of GCDCA by restoring the MMP. Thus, TbMCA might prove useful in ameliorating liver injury in cholestasis. 683 RECOMBINANT ADENOVIRUS IL-24-BAX PROMOTES APOPTOSIS OF HEPATOCELLULAR CARCINOMA CELLS IN VITRO AND IN VIVO L. Shi. Department of Comprehensive Treatment, Eastern Hepatobiliary Surgery Hospital, The Second Military Medical University, Shanghai, China E-mail: [email protected] Background and Aims: Gene therapy promises to become an alternative choice for treatment in hepatic cancer. In many cancers, the delivery of chimeric proteins by adenovirus vector has been reported to induce apoptosis. The present study was performed to evaluate if the recombinant adenovirus IL-24-Bax (Ad.IL-24-Bax) can induce apoptosis in hepatocellular carcinoma cells in vitro and in vivo. Methods: Several recombinant adenoviruses (Ad.Luc, Ad.IL-24, Ad.Bax, and Ad.IL-24-Bax) were constructed and the expression of their encoded proteins was measured. The effects of the recombinant adenovirus on hepatocellular carcinoma cells (Hep3B, HepG2, and PLC/PRF/5) and the normal hepatocyte cell line L02 were investigated through cell viability and apoptosis assays in
Journal of Hepatology 2011 vol. 54 | S209–S361
679 ACTIVATION OF WNT/b-CATENIN SIGNALING PATHWAY IN THE LIVER OF TRANSGENIC MICE OVEREXPRESSING GROWTH HORMONE J.G. Miquet1 , C.S. Martinez1 , L. Gonzalez ´ 1 , M.E. D´ıaz1 , E. Zotta2 , A. Bartke3 , D. Turyn1 , A.I. Sotelo1 . 1 Department of Biological Chemistry – IQUIFIB, School of Pharmacy and Biochemistry, University of Buenos Aires – CONICET, 2 Department of Physiology, School of Medicine, University of Buenos Aires, Ciudad Autonoma de Buenos Aires, Argentina; 3 Departments of Internal Medicine and Physiology, School of Medicine, Southern Illinois University, Springfield, IL, USA E-mail: [email protected] Background and Aims: Transgenic mice overexpressing growth hormone (GH) exhibit hepatomegaly due to hypertrophy and hyperplasia. Throughout lifespan, transgenic mice present high levels of hepatocellular replication and at advanced ages develop liver tumors, mainly hepatocellular carcinoma (HCC). Wnt/b-catenin signaling regulates cell proliferation, apoptosis and differentiation, and is believed to play a role in carcinogenesis. A high percentage of human HCC show high levels of b-catenin, usually located at the cytoplasm or nucleus rather than at the plasma membrane. Control of b-catenin stability is regulated by GSK-3b, which phosphorylates b-catenin, promoting its degradation. The objective of this work was to evaluate if Wnt/b-catenin signaling is dysregulated in the liver of GH-transgenic mice to assess its possible association with the liver pathology observed in these animals. Methods: Young adult transgenic mice overexpressing GH, which present preneoplastic liver pathology, were studied; non-transgenic siblings were used as controls. Liver samples were subjected to qRT-PCR, Western-blotting and immunohistochemistry to evaluate gene expression, protein content and cellular localization of the mediators under study. Results: b-catenin protein content was 2-fold increased in the liver of GH-overexpressing mice, although its mRNA levels were lower compared to normal siblings (P < 0.002). Immunohistochemical analyses revealed that b-catenin was mainly located in the nucleus or in the cytoplasm in transgenic mice. Transgenic mice presented GSK-3b mRNA levels similar to those observed for control mice, but its protein content was elevated, with a parallel increase in its phosphorylation levels at serine-9, which renders GSK-3b inactive (P < 0.001). The expression of the b-catenin target genes cyclin D1, c-myc and c-jun was increased in transgenic mice liver (P < 0.005). Conclusions: Wnt/b-catenin signaling is dysregulated in the liver of GH-transgenic mice. The increased protein levels and nuclear/ cytoplasmic localization of b-catenin, without an increase in its mRNA levels, suggest that the upregulation of b-catenin may be associated with increased protein stability, in accordance with the overexpression of inactive GSK-3b found in these mice. Elevated levels of inactive GSK-3b were previously reported in HCC harboring b-catenin accumulation. We propose aberrant Wnt/b-catenin signaling may be involved in the preneoplastic liver pathology observed in GH-overexpressing transgenic mice. 680 NVP BEZ235-AN ALONE OR IN COMBINATION WITH CHEMOTHERAPY EFFECTIVELY INHIBITS HEPATOCELLULAR CARCINOMA GROWTH D. Pothiraju, M.P. Manns, A. Vogel. Hannover Medical School, Hannover, Germany E-mail: [email protected] Background and Aims: Hepatocellular carcinoma (HCC) is the fifth most frequent cancer and third most lethal cancer worldwide. Activated PI3K/ AKT/ mTOR pathway has recently emerged as a pivotal contributor to HCC development. Aberrant mTOR signaling has been identified in up to 50–60% of HCC cases, associated
Journal of Hepatology 2011 vol. 54 | S209–S361
S273
POSTERS with insulin-like growth factor pathway activation and PTEN dysregulation. The aim of this study was to delineate the role of PI3k and mTOR inhibitor (NVP BEZ235-AN) in HCC. Methods: HuH7 cells were treated with dual inhibitor i.e. AKT & mTOR (NVP BEZ235-AN) inhibitor, Doxorubicin, Cisplatin, 5Fluoro Uracil and Irinotecan alone or in combination for 48hrs. Cell proliferation was measured using MTT assay. Cell cycle (PI staining) and apoptosis (Annexin vs PI) analysis were carried out by FACS. Signaling mechanism of these drugs was confirmed by western blots. In in vivo studies, nude mice were transplanted with Huh7 cells. Tumor size was measured. Tissue pathological analysis was done by HE staining, proliferation was done by IHC using Ki67 antigen and pHis3 anitibody. Apoptosis was measured by Tunel staining. Results: All drugs had an inhibitory effect on cell viability. Combination of BEZ with cisplatin significantly decreased the viability of hepatoma cells compared to Doxorubicin, Irinotecan and 5FU. BEZ treatment inhibits the mTOR phosphorylation and higher concentration of BEZ decreased AKT phosphorylation. Tumor size was significantly decreased in combination treated mice compared to single treated mice. Proliferating cell count was less in treated samples compared to control. Apoptotic cell number was more in treated mice compared to control mice. Conclusion: BEZ effectively induces loss of cell viability. Additionally, BEZ and cisplatin showed synergistic effect on cell proliferation inhibition. Xenograft models confirmed reduced tumor growth in single treatment and moreover, tumor growth was highly regressed in combination of BEZ with cisplatin treatment compared to single treatment. 681 THE COMBINATION OF NEXAVAR AND SIRNA-VEGF INHIBITS THE NEXAVAR-INDUCED INCREASE OF VEGF RESULTING IN ENHANCED ANGIOSTATIC EFFECTS E. Raskopf, T. Sauerbruch, V. Schmitz. Department of Internal Medicine I, University of Bonn, Bonn, Germany E-mail: [email protected] Background and Aims: Nexavar is an established medication in the treatment of advanced HCC and effects on the tumour cell itself are in focus of current research. However, effects on endothelial cells are barely investigated. Therefore, we investigated the effects of Nexavar alone and in combination with siRNA targeting VEGF in human and murine endothelial cells in vitro. Methods: SVEC4–10 and HUVE cells were incubated with different Nexavar-concentrations (0, 1, 2.5, 5 and 10 mM) and analysed for cell viability, proliferation, apoptosis, intracellular signal transduction and VEGF-secretion. In analogy to the mono-therapy, Nexavar was combined with siRNA-VEGF to investigate possible synergistic effects on endothelial cells. Results: Treatment with Nexavar inhibited cell viability in a dose dependent-manner in both cell lines (−85% for HUVE-cells and −90% for SVEC4–10 at 10 mM). Cell proliferation was affected in a similar way. Apoptosis was not influenced under Nexavartreatment. Analysis of intracellular signal transduction showed that phosphorylation of AKT was reduced in a dose-dependent manner. Interestingly, VEGF was increased following Nexavar treatment, peaking at 5 mM Nexavar in both cell lines. This increase was reversed using siRNA targeting VEGF. VEGF-secretion was decreased by 50% compared to the control-siRNA. Furthermore, cell viability was also further decreased (by 30%) after combination of Nexavar with siRNA-VEGF compared to Nexavar alone. Conclusions: Treatment with Nexavar reduced cell viability in both endothelial cell lines, whereas VEGF-levels were increased. This increase was antagonised by siRNA-VEGF leading to an enhanced efficacy of Nexavar. Therefore the combination of both treatment strategies could reduce Nexavar medication and corresponding side effects. S274
682 TAURO-b-MURICHOLIC ACID REDUCES GLYCOCHENODEOXYCHOLIC ACID-INDUCED APOPTOSIS IN A HUMAN HEPATOMA CELL LINE BY RESTORING THE MITOCHONDRIAL MEMBRANE POTENTIAL P. Kleiss1 , G. Denk1 , R. Wimmer1 , H. Zischka2 , C. Rust1 . 1 Department of Medicine 2 – Grosshadern, Ludwig-Maximilians University of Munich, 2 Institute of Toxicology, Helmholtz Zentrum Munich, German Research Center for Environmental Health, Munich, Germany E-mail: [email protected] Introduction and Aim: b-muricholic acid (bMCA) is a natural trihydroxylated bile acid and constitutes the major bile acid synthesized in rat and mouse. Due to the presence of a hydroxyl group in the 6b position of the steroid ring, bMCA is more hydrophilic than ursodeoxycholic acid. In the past, bMCA has been evaluated for the dissolution of cholesterol gallstones (Montet et al., Biochemica et Biopysica Acta 1987: 918: 1–19). However, it is unknown if bMCA has beneficial effects on cell death induced by toxic bile acids. Therefore, our aim was to determine the effect of bMCA on apoptosis induced by glycochenodeoxycholic acid (GCDCA) in a human hepatoma cell line stably transfected with the sodium taurocholate cotransporting poylpetide (HepG2-Ntcp). Methods: Ntcp-transfected HepG2 were stimulated for 4 hours with GCDCA, tauro-b-muricholic acid (TbMCA), tauroursodeoxycholic acid (TUDCA) at a concentration of 25 mM each and DMSO (‰, respectively. Apoptosis was then quantified biochemically in a caspase 3/7-assay as well as morphologically after Hoechst33342 staining. The mitochondrial membrane potential (MMP) was measured fluorometrically using JC-1 (5,5 ,6,6 -tetrachloro-1,1 ,3,3 tetraethyl-benzimidazol-carbocyaniniodide) at a concentration of 2 mM at 2, 4 and 6 hours after stimulation with bile acids (25 mM, 50 mM and 100 mM) alone or in combination. Results: In HepG2-Ntcp cells, GCDCA induced a 18±8-fold increase in apoptosis after 4 hours, respectively, compared to untreated cells as determined in caspase 3/7-assays (n = 6, each). Simultaneous incubation with TbMCA reduced apoptosis to 49% as compared to 44% with TUDCA, both significant differences (p < 0.01 vs. GCDCA, each; n = 6). Similar results were obtained when apoptosis was assessed morphologically by Hoechst-33342 staining. While GCDCA (100mM) reduced the MMP to 58% (2 h), 45% (4 h) and 34% (6 h), combination treatment with TbMCA restored the MMP to control levels at all time points (n = 4). Conclusions: TbMCA significantly reduced hepatocyte apoptosis induced by low micromolar concentrations of GCDCA by restoring the MMP. Thus, TbMCA might prove useful in ameliorating liver injury in cholestasis. 683 RECOMBINANT ADENOVIRUS IL-24-BAX PROMOTES APOPTOSIS OF HEPATOCELLULAR CARCINOMA CELLS IN VITRO AND IN VIVO L. Shi. Department of Comprehensive Treatment, Eastern Hepatobiliary Surgery Hospital, The Second Military Medical University, Shanghai, China E-mail: [email protected] Background and Aims: Gene therapy promises to become an alternative choice for treatment in hepatic cancer. In many cancers, the delivery of chimeric proteins by adenovirus vector has been reported to induce apoptosis. The present study was performed to evaluate if the recombinant adenovirus IL-24-Bax (Ad.IL-24-Bax) can induce apoptosis in hepatocellular carcinoma cells in vitro and in vivo. Methods: Several recombinant adenoviruses (Ad.Luc, Ad.IL-24, Ad.Bax, and Ad.IL-24-Bax) were constructed and the expression of their encoded proteins was measured. The effects of the recombinant adenovirus on hepatocellular carcinoma cells (Hep3B, HepG2, and PLC/PRF/5) and the normal hepatocyte cell line L02 were investigated through cell viability and apoptosis assays in
Journal of Hepatology 2011 vol. 54 | S209–S361