T1957

Utility of Serum Fibrosis Marker to Screen Significant Fibrosis in PatientsWith Chronic Hepatitis; New Scoring System Consisted of Type IV Collagenand AP IndexMasatoshi Ishigami, Kazuhiko Hayashi, Akihiro Itoh, Yoshiki Hirooka, Yoshiaki Katano,Hidemi Goto

(Backgrounds)To detect significant fibrosis in chronic hepatitis is important to determinethe timing of treatment intervention. Liver biopsy is still a gold standard until now, thoughit is quite invasive. In this study, we evaluate the utility of some laboratory tests and indecesthat are related to liver fibrosis and found new formulas that is considered to be moreeffective than previously reported indeces and proposed as one of the noninvasive and easydiagnostic tool. (Patients and Methods) 95 patients with chronic hepatitis who have alsodone liver biopsy were included in this study. We evaluated the relationship betweensignificant fibrosis (Metavir score≧2) and age, sex, serum AST, ALT, γGTP, hyaruronan,Procollagen-III-peptide (P-III-P), type IV collagen, platelet counts (Plt) as single serummarker, and AST/ALT ratio (AAR), AP index, and AST to platelet ratio index (APRI) asfibrosis indeces. In univariate analysis, Students' t-test was applied in continuous values andChi-square test was applied in categorical values. In multivariate analysis, logistic regressionanalysis was done. P<0.05 was considered as statistically significant. And to evaluate theutility of new diagnostic index, ROC analysis was done and AUROCwas evaluated. (Results)Inunivariate analysis, higher age (p=0.0067), higher AST (p=0.0073), higher hyaruronan (p=0.0015), higher P-III-P (p<0.0001), higher type IV collagen (p<0.0001), lower Plt(p<0.0001),higher AP index (p<0.0001), and higher APRI (p<0.0001) was selected, and inmultivariate analysis, Type IV collagen (p=0.0360) and AP index (p=0.0106) were selectedas significant factors. Then we made new scoring system consisted of type IV collagen(≧150:1point,<150:0point) and AP index (≧6: 1 point, <6: 0 point). AUROC of this newscore was 0.834. Score 2 points predicted 86.7% (26/30) of patients with significant fibrosisand Score 0 ruled out 96.9% (31/32) of patients with mild fibrosis. (Conclusions)This newdiagnostic score consisted of routine laboratory test might be one of the effective andconvenient tools to determine the indication of liver biopsy that is quite invasive for thepatients, and the timing of treatment intervention in patients with chronic hepatitis.

T1958

Liver Gene Expression Signature of Mild Fibrosis in Chronic Hepatitis CA. Ducès, M. Lapalus, Ivan Bièche, Benedicte Jardin-Watelet, Eve Dupas, S. De Muynck,Isabelle Molina, N. Lambert, B. Sallenave, E. Nicolas, E. Estrabaud, Nathalie Jullian, M.Martinot-Peignoux, Olivier Lada, V. Paradis, Dominique Valla, Pierre Bedossa, DanielLaune, M. Vidaud, Patrick Marcellin, T. Asselah

BACKGROUND AND AIMS In chronic hepatitis C, the transition from mild to moderatefibrosis seems to be a major prognostic step. The aim of this study was to identify molecularmarkers that could differentiate mild from moderate fibrosis in chronic hepatitis C.METHODS Liver biopsies from 244 untreated patients (or prior to treatment) were studied.Among these patients 66% had mild fibrosis (F1, Metavir score) and 34% septal fibrosis(F2). Patients weremainly infectedwith genotype 1 (56%), 2 (10.5%), 3 (10.5%), 4 (20%) and5-6 (3%). Real-time quantitative RT-PCR assays were used to analyse the mRNA expression of51 genes selected from the literature and involved in various cellular and molecular mechan-isms associated with liver gene expression during hepatitis C infection. RESULTS Theexpression profiles of mild fibrosis differ significantly from those of moderate fibrosis, and24 genes were found to be upregulated in F2 patients. These genes are involved in extracellularmatrix production and remodelling (matrix proteases family), or in cell-cell and cell-extracel-lular matrix interactions. Disregulated genes also belongs to growth factors/cytokines families(CXC chemokines), and are involved in cell cycle. CONCLUSION Mild and moderate fibrosishave different liver gene expression. The most notable changes occurred mainly in cell-matrix turn-over. The genes included in the signature encode molecules secreted in theserum and provide a logical functional approach for the development of serum markers offibrosis progression.

T1959

Telmisartan Had a Hepatoprotective and Anti-Fibrotic Effects in Patients WithChronic Hepatitis C Treated With Pegylated Interferon and RibavirinAlaa E. El-sisi, Asem Elfert, Magda El-sayad, Sherin Zakaria

Background & Aims: Telmisartan is an angiotensin II T1 receptor blocker (ARB) withpartial peroxisome proliferator-activated receptor gamma (PPARgamma) agonist properties.Telmisartan had a significant antifibrotic and hepatoprotective effects in Ccl4-induced hepaticfibrosis in rats (Awara et al. J. Egypt. Soc. Pharmacol. Exp, Ther. 2008; 29(1): 175-200).Also, telmisartan showed a significantly lower fibrosis in hypertensive patients with NASH(Georgescu et al. Worl J. Gastroenterol. 2009; 15(8): 942-954). The aim of this study wasto evaluate the antifibrotic effect of telmisartan compared to losartan in patients with chronichepatitis C (HCV) treated with pegylated interferon and ribavirin. Methods: Sixty patientswere randomized into 3 groups. Telmisartan (40mg/day) or Losartan (50mg/day) was givento HCV patients treated with pegylated interferon α2a (180μg/week) and ribavirin (15mg/kg) for 8 weeks. Control group received pegylated interferon and ribavirin only. Bloodsamples were collected before treatment and after 8 weeks for measurement of ALT, AST,bilirubin, creatinine and CBC. Urinary excretion of hydroxyproline was also estimated.Serum samples were stored for measurement of alpha 2 macroglobulin, metalloproteinaseII and TGF-beta before and after 8 weeks of therapy. Results: All treated patients showeda significant decrease in ALT serum levels. Telmisartan treated patients showed a significantreduction in AST compared to losartan and control groups (p<0.05). Also, telmisartan treatedpatients had a 30.83% decrease in the urinary excretion of hydroxyproline compared tobaseline. This was significant when compared with the other groups (p<0.05). Telmisartanand losartan treated patients showed significantly less leucopenia and thrombocytopenia incomparison with the control group (p<0.05). Conclusions: In our study, telmisartan had ahepatoprotective and anti-fibrotic effects in HCV patients under treatment with pegylated

S-837 AASLD Abstracts

interferon and ribavirin. In addition, both telmisartan and losartan had a secondary beneficialeffect against hematopoietic toxicity of interferon.

T1960

Leptin Activates Hedgehog Pathway Signaling and Promotes MyofibroblasticTransition and Accumulation in Rat Hepatic Stellate CellsSteve S. Choi, Wing-Kin Syn, Gamze F. Karaca, Alessia Omenetti, Xiaoling Wang,Youngmi Jung, Vanessa S. Teaberry, Cynthia A. Moylan, Anna Mae Diehl

Background: Nonalcoholic fatty liver disease (NAFLD) is strongly associated with obesityand the metabolic syndrome. In NAFLD, cirrhosis develops when nonalcoholic fatty liver(steatosis) progresses to NASH and is heralded by hepatic accumulation of myofibroblasts.Hepatic stellate cells (HSC) are a major source of myofibroblasts in injured livers and theobesity-related factor, leptin, is known to promote growth of cultured myofibroblasts andliver fibrosis. Leptin and Hedgehog (Hh) are key mediators of MF-HSC accumulation, andit is unclear how leptin and Hh interact to regulate MF-HSC accumulation. Two events arenecessary for MF-HSC to accumulate in damaged livers: transition of resident, quiescentHSC to MF-HSC, and expansion of MF-HSC numbers. We evaluated the hypothesis thatleptin promotes the activation of the Hh pathway, thereby stimulating signals that promotetransition of Q-HSC into MF-HSC and enhance accumulation of MF-HSC. Methods: Freshly-isolated, primary rat HSC were cultured for 7 days to induce transition into MF-HSC, thentreated with vehicle, leptin (100 ng/mL) ± cyclopamine (5 μM, Hh pathway inhibitor) ortomatidine (an inactive cyclopamine analog), or inhibitors of a key HSC survival pathway(PI3-K/AKT). After 48 h,MF-HSCwere harvested formRNA and protein analysis by quantitat-ive, real-time RT-PCR and immunoblot. Results: As expected, HSC expression of myofibrobl-astic genes increased during culture activation. During culture, expression of Hhip (endogen-ous Hh inhibitor) decreased while sonic hedgehog (ligand) and Gli2 (target gene) expressionwas induced. Adding leptin resulted in a 2-fold increase in expression of myofibroblasticgenes (αSMA andCol1α1), while increasing Shh andGli2 expression by 2-fold and decreasingHhip expression by 50%. Treatment with cyclopamine decreased myofibroblastic geneexpression by 50% and blocked Hh pathway expression compared to controls. Inhibitionof PI3-K/AKT pathway attenuated the effects of leptin, while cyclopamine treatment blockedthe profibrogenic effects of leptin. Interestingly, leptin treatment promoted changes in geneexpression associated with epithelial-to-mesenchymal transitions (EMT), a contributor toMF-HSC transition. Expression of BMP-7, an EMT inhibitor, decreased by 50% whileexpression of snail, an EMT inducer, increased 2-fold. Hh pathway inhibition reversed theseeffects. Conclusions: These findings suggest that the profibrogenic effect of leptin resultsfrom activation of the Hh signaling pathway to promoteMF-HSC transition and accumulationand have important implications for the pathogenesis of cirrhosis in NAFLD.

T1961

The Role of Matrix Metalloproteinase 2 (MMP2) in the EMT-Like Changes ofHepatocytesJenny Chen, Joy X. Jiang, Nobuko Serizawa, Natalie Torok

BACKGROUND: Hepatocytes have been shown to contribute to liver fibrosis via epithelialmesencyhmal transition (EMT). However, the microenvironment that plays a role in thesechanges is less defined. Activated hepatic stellate cells (HSC) are located in close proximityto hepatocytes and known to secrete mediators (e.g. TGF-β, MMPs) which may induceEMT-like responses of hepatocytes. Previously we have shown that phagocytosis of apoptoticbodies (AB) induces TGF-β1 expression in HSC. Our HYPOTEHSIS is that in addition toTGF-β1, activated HSC secrete MMP2 which plays a role in the EMT-like changes inhepatocytes. METHODS: To establish the effect of TGF-β, primary rat hepatocytes (24 hoursafter isolation) were treated with recombinant TGF-β1. Immunocytochemistry and real-timepolymerase chain reaction (RT-PCR) were performed to examine the expression of albumin,E-cadherin, and Snail. To study if the expression of MMP2 was upregulated in primary HSCfollowing their activation, the cells were exposed to AB and RT-PCR was performed. Toassess if active MMP2 induced EMT of hepatocytes, primary rat hepatocytes were treatedwith active MMP2 and the expression of albumin, E-cadherin and Snail were evaluated byRT-PCR. To study the causal relationship between MMP2 activity and EMT-like changes inhepatocytes, a co-culture of primary hepatocytes and phagocytosis-activated primary HSCcultured in the presence or absence of an MMP2 inhibitor was established in a Transwellsystem. RESULTS: Primary hepatocytes exposed to TGF-β exhibited fibroblast-like changeswith loss of cell polarity, a decrease in E-cadherin and albumin staining, a decrease in E-cadherin and albumin expression, and an increase in Snail expression by RT-PCR. Exposinghepatocytes to active MMP2 induced similar changes with a significant decrease in E-cadherin(p=0.02) and albumin (p=0.03) expression and an increase in Snail expression. Phagocytosisof AB induced a significant upregulation of MMP2 in HSC (p=0.01). In the co-culturesystem, E-cadherin expression decreased significantly when hepatocytes were cultured withphagocytosing HSC (p=0.01), while Snail expression was increased. When using the MMP2inhibitor, Snail expression decreased (p=0.03) suggesting that MMP2 may play a role in theEMT of hepatocytes. Further studies are required to delineate the exact contribution of TGF-β and MMP2 to the morphological changes of hepatocytes. CONCLUSIONS: TGF-β andMMP2 secreted from active HSC may induce EMT-like changes in hepatocytes duringliver fibrogenesis.

T1962

PPARγ Reverses Hepatic Nutritional Fibrosis in Mice and Induces HepaticStellate Cell Apoptosis Through Intrinsic Apoptotic PathwayJun Yu, Sui Zhang, Eagle SH Chu, Minnie Y. Go, Rebecca HY Lau, Junhong Zhao,Chung-Wah Wu, Terence CW Poon, Joseph J. Sung

Background and aims: PPARγ ligand prevented the development of fibrosing steatohepatitisin mice. The role of PPARγ as a potential therapeutic target for this disorder remainsdetermined. We evaluated the role of PPARγ on established fibrosing steatohepatitis in mice

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