BACKGROUND/AIMS: The Wnt/beta-catenin signaling pathway has been reported to play an important role in liver fibrosis. This study was designed to investigate whether mesoderm-specific transcript homologue (Mest), a strong negative regulator of Wnt/beta-catenin signaling, could inhibit liver fibrosis. METHODS: pcDNA-Mest was transfected into hepatic stellate cells (HSCs) and rats. Rats were randomly divided into four groups: normal group (normal saline), treatment group (pcDNA-Mest+CCl4), control group (pcDNA-neo+CCl4), and model group (normal saline+CCl4). Changes in liver pathology were evaluated by hematoxylin and eosin and Masson's trichrome staining. The levels of alanine transaminase, aspartate transaminase, lactic dehygrogenase, hyaluronic acid, and laminin in the serum and hydroxyproline in the liver were detected by biochemical examination and radioimmunoassay, respectively. The expression and distribution of beta-catenin, alpha-smooth muscle actin (alpha-SMA), Smad3, and tissue inhibitor of metalloproteinase type I were determined, and the viability of the HSCs was tested. RESULTS: Our data demonstrate that Mest alleviated CCl4-induced collagen deposition in liver tissue and improved the condition of the liver in rats. Mest also significantly reduced the expression and distribution of beta-catenin, alpha-SMA and Smad3 both in vivo and in vitro, in addition to the viability of HSCs in vitro. CONCLUSIONS: We found that Mest attenuates liver fibrosis by repressing beta-catenin expression, which provides a new therapeutic approach for treating liver fibrosis.
Animals ; Carbon Tetrachloride/toxicity ; Cells, Cultured ; Hepatic Stellate Cells/physiology ; Liver Cirrhosis, Experimental/*physiopathology ; Male ; Proteins/*physiology ; Random Allocation ; Rats, Wistar ; Transfection ; Wnt Signaling Pathway/*physiology ; beta Catenin/metabolism

Magnesium lithospermate B (MLB) is one of the major active components of Salvia miltiorrhizae. The anti-oxidative effects of Salvia miltiorrhizae have been previously reported. The aim of this study was to investigate the effect of purified MLB on hepatic fibrosis in rats and on the fibrogenic responses in hepatic stellate cells (HSCs). Hepatic fibrosis was induced in rats by intraperitoneal thioacetamide (TAA) injections over a period of 8 or 12 weeks. MLB was orally administered daily by gavage tube. Serum AST and ALT levels in TAA + MLB group were significantly lower than those in TAA only group at week 8. Hepatic fibrosis was significantly attenuated in TAA + MLB group than in TAA only group at week 8 or 12. Activation of HSCs was also decreased in TAA + MLB group as compared to TAA only group. Hepatic mRNA expression of alpha-smooth muscle actin (alpha-SMA), TGF-beta1, and collagen alpha1(I) was significantly decreased in TAA + MLB group as compared to TAA only group. Incubation with HSCs and MLB (> or =100 microM) for up to 48 h showed no cytotoxicity. MLB suppressed PDGF-induced HSC proliferation. MLB inhibited NF-kappaB transcriptional activation and monocyte chemotactic protein 1 (MCP-1) production in HSCs. MLB strongly suppressed H2O2-induced reactive oxygen species (ROS) generation in HSCs, and MLB inhibited type I collagen secretion in HSCs. We concluded that MLB has potent antifibrotic effect in TAA-treated cirrhotic rats, and inhibits fibrogenic responses in HSCs. These data suggest that MLB has potential as a novel therapy for hepatic fibrosis.
Actins/genetics/metabolism ; Animals ; Antioxidants/*administration & dosage ; Cell Proliferation/drug effects ; Collagen Type I/genetics/metabolism ; Drugs, Chinese Herbal/*administration & dosage ; Fibrosis/prevention & control ; Hepatic Stellate Cells/*drug effects/metabolism/pathology ; Liver/*drug effects/metabolism/pathology ; Liver Cirrhosis, Experimental/chemically induced/*drug therapy/physiopathology ; Male ; NF-kappa B/metabolism ; Rats ; Rats, Sprague-Dawley ; Reactive Oxygen Species/metabolism ; Salvia miltiorrhiza/immunology ; Thioacetamide/administration & dosage ; Transcriptional Activation/drug effects

To investigate the role of heme oxygenase(HO), a catalyzing enzyme of heme to produce CO, in modulation of systemic circulation in CCl4-induced cirrhotic rats. Saline(vehicle) and ZnPP were s.c. injected into the posterior necks of rats respectively and the rats were then anesthetized by pentobarbital sodium in four hours. Mean arterial pressure (MAP, kPa), heart rate (HR, b/min) and portal pressure (PP, cm/H2O) were measured by indwelling catheter. Plasma CO was determined by Chalmers method. Heme oxygenase acivity was determined by the rate of bilirubin formation. The cirrhotic rats showed significant hyperdynamic circulation indicated by decreased mean arterial pressure [MAP, (15.6+/-1.7) vs (18.9+/-0.9) kPa, t = 4.52, P less than 0.01] and increased portal pressure [PP, (16.7+/-0.8) vs (8.8+/-0.3) cm H2O, t = 23.10, P less than 0.01] as compared to normal control rats(NS). ZnPP could cause a significant increase in MAP [(17.3+/-1.5) vs (15.6+/-1.7) kPa, t = 2.18, P less than 0.05] and significant decrease in PP [(13.2+/-0.7) vs (16.7+/-0.8) cm H2O, t = 8.53, P less than 0.01] in cirrhotic rats. The cirrhotic group presented a significant increase in plasma CO [(18.0+/-1.9) vs (10.4+/-1.3)mumol/L, t = 8.42, P less than 0.01] and HO activity in the spleens [(11.1+/-0.9) vs (6.5+/-0.9) nmol bilirubin/mg protein/h, t = 9.28, P less than 0.01] and intestines [(2.5+/-0.1) vs. (1.3+/-0.2) nmol bilirubin/mg protein/h, t = 15.1, P less than 0.01]. ZnPP could cause significant decreases in plasma CO and HO activity in liver, spleen and intestine of both control and cirrhotic rats. HO-CO system activation may be an important reason for the hemodynamic disturbance of liver cirrhosis.
Animals ; Carbon Monoxide ; metabolism ; Heme Oxygenase (Decyclizing) ; metabolism ; Hemodynamics ; Liver ; physiopathology ; Liver Cirrhosis, Experimental ; metabolism ; physiopathology ; Male ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo observe the occurrence and progression of liver fibrosis induced by pig serum exposure and bile duct ligation, and analyze the relationship between hepatic inflammation and liver fibrosis.

METHODSChronically immune-mediated liver fibrosis was induced in rats by weekly injection of pig serum (IPS) into the peritoneal cavity at 3 ml/kg for 12 weeks. Cholestatic fibrosis was induced by common bile duct ligation (BDL). The Knodell score was used to evaluate the histological changes in the liver, and immunohistochemistry was performed using anti-SMA, anti-ED1, anti-CK7, and anti-CD45 antibodies. Quantitative real time PCR (qPCR) analysis was employed to quantify the mRNA expression of the genes related to inflammation, including interleukin-1beta (IL-1beta), IL-6, monocyte chemotactic protein-1, tumor necrosis factor-alpha, regulated upon activation normal T cell expressed and secreted (RANTES), transforming growth factor-beta, platelet-derived growth factor A, as well as the genes associated with fibrogenesis, namely collagen 1, alphaSMA, MMP-9 and TIMP-1.

RESULTSKnodell scores for periportal necrosis, intralobular degeneration and focal necrosis, and portal inflammation were all significantly higher in the BDL group than in the IPS group (P<0.01), whereas the scores for fibrosis was higher in the IPS group (P<0.05). Immunohistochemistry showed obvious inflammation with numerous alphaSMA-positive cells in the liver of the rats in BDL group; the liver of the rats in IPS group showed numerous alphaSMA-positive myofibroblasts with limited inflammatory cell infiltration. qPCR demonstrated a significant up-regulation of the genes related to extracellular matrix remodeling such as collagen 1 (P<0.01), alphaSMA (P<0.01), MMP-9 (P<0.01) and TIMP-1 (P<0.01) in the rat liver in IPS group compared with those in the normal control group, and the mRNA expressions of the inflammation-related cytokines, except for RANTES, were comparable with those in the control. In contrast, the BDL group showed a significant up-regulation of all the pro-inflammatory genes examined with also increased expression of the fibrogenesis-related genes (P<0.05).

CONCLUSIONLiver fibrosis induced by IPS is characterized by active ECM remodeling in the absence of obvious inflammation, indicating that chronic development of liver fibrosis can be independent of active hepatic inflammation. BDL-induced liver fibrosis highlights obvious inflammation and fibrous proliferation in the liver.

Animals ; Bile Ducts ; surgery ; Cholestasis ; complications ; physiopathology ; Ligation ; Liver Cirrhosis, Experimental ; etiology ; immunology ; pathology ; Male ; Rats ; Rats, Inbred F344 ; Serum ; immunology ; Swine

OBJECTIVETo investigate the effects of Fuzheng Huayu recipe and Huangqi tang on DMN-induced experimental liver cirrhosis in rats and explore the therapeutic characteristics of Buxu herbals on liver cirrhosis.

METHODLiver cirrhosis in rats was induced by intraperitoneally injection of DMN for 4 weeks. Cirrhotic rats were randomly divided into 4 groups: model group, and Fuzheng Huayu recipe group, Huangqi tang group, Fuzheng Huayu recipe combined with Huangqi Tang group. The rats in treatment groups were orally administered with Fuzheng Huayu recipe, Huangqi tang, Fuzheng Huayu recipe combined with Huangqi tang (1:1), respectively. Normal and model control rats were given the equivalent normal saline. The body weight, liver weight and spleen weight were observed when rats were sacrificed. Liver histology was examined by HE staining and Sirius red staining. The liver function parameters including ALT, T. Bil and Alb were determined. The SOD activity and MDA content in liver tissues were also measured. Hepatic hydroxyproline (Hyp) content was determined by Jamall's method. The expression of alpha-SMA was determined by both immunohistochemistry staining and western blot method.

RESULTCompared with normal rats, the serum ALT and T. Bil levels in model rats increased obviously, by contrast, the serum Alb level decreased. There was a significant decline of SOD activity in model rat liver tissue, while the content of MDA and Hyp increased remarkably. A severe deterioration of liver architecture, infiltration of inflammatory cells and deposition of collagen were observed in model rat liver tissue. The expression of alpha-SMA also increased significantly. Compared with model rats, the liver function, lipid peroxidation parameters, Hyp content and liver histology were all improved in the 3 treatment groups. The combined group is better than any single-use group in decreasing collagen deposition and expression of alpha-SMA.

CONCLUSIONFuzheng Huayu recipe, Huangqi tang, Fuzheng Huayu recipe combined with Huangqi tang can attenuate liver fibrosis in DMN induced rats. Fuzheng Huayu recipe combined with Huangqi tang is better than that using alone in decreasing collagen deposition. The mechanism is partially due to the better effect of Fuzheng Huayu recipe combined with Huangqi tang on inhibiting activated HSC.

Animals ; Body Weight ; drug effects ; Dimethylnitrosamine ; adverse effects ; Disease Models, Animal ; Drugs, Chinese Herbal ; administration & dosage ; Humans ; Liver ; drug effects ; enzymology ; Liver Cirrhosis, Experimental ; chemically induced ; drug therapy ; enzymology ; physiopathology ; Organ Size ; drug effects ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Superoxide Dismutase ; metabolism

Animals ; Colon ; blood supply ; pathology ; Colonic Diseases ; drug therapy ; etiology ; physiopathology ; Hemodynamics ; Hepatic Veins ; pathology ; physiopathology ; Hypertension, Portal ; complications ; physiopathology ; Intestinal Mucosa ; blood supply ; drug effects ; pathology ; Liver Cirrhosis, Experimental ; complications ; Losartan ; administration & dosage ; therapeutic use ; Male ; Microscopy ; Portal Pressure ; drug effects ; Random Allocation ; Rats ; Rats, Wistar

OBJECTIVETo explore the mechanism of Danggui Buxue Decoction (, DBD) on the liver fifibrosis related to hepatic lipid peroxidation and matrix metalloproteinases (MMP) -2/9 activities.

METHODSThe liver fifibrosis in 28 rats was induced by an injection of carbon tetrachloride (CCl(4)) and fed with high lipid and low protein diet for 6 weeks, the model rats were randomly divided into the model group and DBD treated group, 14 in each group, and another 10 rats as the normal group were observed as well. Rats in the DBD group were administered with DBD at the dose of 6 g/kg body weight for 6 weeks since CCl(4) intoxication. The hepatic inflammation and fibrosis were examined with HE and Sirius red stain. The liver function including serum alanine aminotransamine (ALT), aspartate transamine (AST), albumin (Alb) and total bilirubin (TBIL), liver triglyceride (TG) and malondialdehyde (MDA) contents, superoxide dismutase (SOD) activity were assayed. Hepatic hydroxyproline (Hyp) content was detected with Jamall's method. The alpha-SMA expression was analyzed by immunohistochemistry and the Western blot. Liver MMP-2 mRNA was analyzed with Real-time PCR, and MMP-2/9 activities were measured with gelatin zymography and in situ zymography.

RESULTSCompared with the normal group, the levels of ALT, AST and TBIL, the content of Hyp, TG and MDA were remarkably increased, the Alb content and SOD activity were signifificantly decreased in the model group (P<0.05), and higher levels of MMP-2 mRNA and MMP-2/9 activities (P<0.01), the hepatic fatty degeneration and collagen accumulation and fifibrosis at liver were observed. Compared with the model control, DBD group showed slighter hepatic fatty degeneration and collagen deposition, and had lower levels of ALT, AST and TBIL activities, lower contents of MDA, TG and Hyp, but higher SOD level and Alb content (P<0.05), and DBD also down-regulated MMP-2 mRNA expression and decreased MMP-2/9 activities in the fifibrotic livers (P<0.01).

CONCLUSIONThe action of DBD against liver fibrosis is related to prevent lipid peroxidation and inhibit MMP-2/9 activities in the fibrotic livers.

Animals ; Carbon Tetrachloride ; toxicity ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Hepatic Stellate Cells ; drug effects ; Lipid Peroxidation ; drug effects ; Liver ; drug effects ; pathology ; physiopathology ; Liver Cirrhosis, Experimental ; drug therapy ; metabolism ; pathology ; Male ; Matrix Metalloproteinase 2 ; genetics ; metabolism ; Matrix Metalloproteinase 9 ; genetics ; metabolism ; Matrix Metalloproteinase Inhibitors ; Rats ; Rats, Wistar

OBJECTIVETo evaluate the effect of Ginkgo biloba extract (EGb) on hepatic microcirculation and portal hypertension in CCl4 treated rats.

METHODSTwenty-five male Wistar rats were divided into a blank, a CCl4 treated and a CCl4 plus EGb treated group, and all were treated for 10 weeks. The free portal vein pressures were measured through catheterizations. Hepatic sinusoidal endothelial cells and other parameters of hepatic microcirculation were studied with transmission electron microscopy. The amounts of malondialdehyde (MDA), endothelin (ET-1), platelet-activating factor (PAF), nitric oxide (NO), cNOS and iNOS in the liver tissues were determined.

RESULTSThe portal vein pressure of the CCl4 plus EGb treated group was (7.4 +/- 0.6) mm Hg while the pressure of the CCl4 treated group was (8.7 +/- 0.8) mm Hg. Aggregation of blood cells or microthrombosis in hepatic sinusoids, deposition of collagen in hepatic sinusoids and spaces of Disse, injury of endothelial cells and capillarization of hepatic sinusoid were significantly milder in the EGb group. The amounts of MDA, ET-1, PAF, NO and iNOS were markedly lower in the CCl4 plus EGb treated group than in the CCl4 treated group.

CONCLUSIONThe results demonstrated that EGb can decrease the portal vein pressure and improve hepatic microcirculation in CCl4 treated rats. The mechanisms of this effect may involve its inhibition on ET-1, PAF, lipid peroxidation, and down regulation of the hepatic iNOS and NO expressions.

Animals ; Ginkgo biloba ; Hepatic Veins ; pathology ; Hypertension, Portal ; drug therapy ; physiopathology ; Liver Cirrhosis, Experimental ; chemically induced ; drug therapy ; pathology ; physiopathology ; Male ; Microcirculation ; drug effects ; Plant Extracts ; pharmacology ; Rats ; Rats, Wistar

OBJECTIVEThe predictive value of postoperative hepatic function evaluated by liver functional imaging combined with single photon emission computed tomography (SPECT) technique was appraised in the present study.

METHODSTwenty New Zealand white rabbits were divided randomly into two groups, including the Hepatic Fibrosis with Carcinoma Group (FC-Group, n = 10) and the Control Group (C-Group, n = 10). All the rabbits underwent the resection of outer-right lobe of the liver. The whole hepatic function indexes, such as HCI(5), HLI(5) and Ex(15), and the hepatic function remnant indexes, including HCI(5P), HLI(5P) and Ex(15P), were calculated by 99mTc-EHIDA liver imaging.

RESULTSEx(15) of FC-Group was lower than that of C-Group (P < 0.05). HCI(5) and HLI(5) of FC-Group had the trends to increase compared with the C-Group. Ex(15) was positively correlated with ALB, and negatively correlated with TBil and GGT (P < 0.05). HCI(5) had a positive correlation with CHE (P < 0.05), while HLI(5) had a negative correlation with A/G (P < 0.05). HLI(5P) had the negative correlation with postoperative A/G (P < 0.05), and Ex(15P) had the negative correlation with postoperative TBil and GGT (P < 0.05).

CONCLUSIONSThis study has established a method of 3-D liver function evaluation system on an animal model. Among the indexes, Ex(15) can exactly represent the whole liver function while Ex(15P) and HLI(5P) can predict the liver function after the liver resection. The results may help the future clinical use of this technique to evaluate the risk of operation.

Animals ; Hepatectomy ; Liver ; diagnostic imaging ; physiopathology ; surgery ; Liver Cirrhosis, Experimental ; diagnostic imaging ; physiopathology ; surgery ; Liver Function Tests ; Liver Neoplasms, Experimental ; diagnostic imaging ; physiopathology ; surgery ; Postoperative Period ; Rabbits ; Tomography, Emission-Computed, Single-Photon

Animals ; Liver ; pathology ; Liver Cirrhosis, Experimental ; metabolism ; pathology ; physiopathology ; Male ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptor, Platelet-Derived Growth Factor beta ; metabolism