OBJECTIVETo explore the expression of integrin alpha 6 in hepatic sinusoidal capillaration.

METHODSThe rat hepatic fibrosis model was established by injection of carbon tetrachloride subcutaneously. Then the expression of laminin and integrin alpha 6 subunit was observed by immunohistochemistry and dot immuno-blotting.

RESULTSWe observed sinusoidal capillaration formed by deposition of laminin along sinusoids in Disse interspace by immunohistochemistry staining. In normal rat the expression of integrin alpha 6 was restricted to portal vascular endothelial cells and bile duct epithelial cell membranes. No expression was observed in sinusoidal endothelial cell membranes. When capillaration integrin alpha 6 was detected in a continuous pattern along the sinusoids, the content of integrin alpha 6 was significantly higher in fibrotic liver tissues than in normal liver tissues as measured by dot immuno-blotting (P<0.05).

CONCLUSIONSDuring fibrogenesis, laminin continuously accumulate in liver tissues and form basement membrane resulting in sinusoidal capillaration, and then induce the expression of integrin alpha 6 on SEC membranes.

Animals ; Antigens, CD ; biosynthesis ; Carbon Tetrachloride ; Immunoblotting ; Immunohistochemistry ; Integrin alpha6 ; Laminin ; metabolism ; Liver ; metabolism ; pathology ; Liver Cirrhosis, Experimental ; chemically induced ; metabolism ; pathology ; Male ; Rats ; Rats, Wistar ; Time Factors

Animals ; Carbon Tetrachloride ; H(+)-K(+)-Exchanging ATPase ; metabolism ; Liver ; metabolism ; pathology ; Liver Cirrhosis, Experimental ; chemically induced ; metabolism ; pathology ; Male ; RNA, Messenger ; Rats ; Rats, Sprague-Dawley

BACKGROUND/AIMS: Oxidative stress is one of the important underlying mechanisms of hepatic fibrosis. DA-9601, the ethanol extracts of Artemisia asiatica, has been reported to possess strong antioxidative and cytoprotective actions. We tried to evaluate whether antioxidant can ameliorate dimethylnitrosamine (DMN)-induced hepatic fibrosis. METHODS: Rat hepatic fibrosis was induced by intraperitoneal administrations of 10 mg DMN six times. Additionally, rats of one group were started daily with DA-9601 30 mg/kg containing diets and another group was fed a pellet diet containing DA-9601 100 mg/kg. The immunohistochemical studies for collagen, alpha-smooth muscle actin (alpha-SMA), and fibronectin, the measurements of hepatic malondialdehyde (MDA) and collagens, and the changes of liver function profiles were performed. Hepatic stellate cells (HSC) were isolated and in vitro effects of DA-9601 on HSC activations were measured. RESULTS: DA-9601 significantly attenuated the loss of body weights (p<0.05), the reduction of liver wet weights (p<0.05), and the elevation of liver enzymes provoked by DMN administrations. DMN injections caused the severe fibrosis of portal tract, hepatic inflammation, and significant oxidative damages, but DA-9601 treatment significantly reduced the mean scores of hepatic fibrosis, the amounts of hepatic collagens, and hepatic MDA levels. The prominent decreases in the expressions of collagens type I and III, alpha-SMA, and fibronectin or hepatic inflammations were observed in DA-9601-treated groups dose-dependently and similar efficacy was also proven in in vitro HSC experiment. CONCLUSIONS: DA-9601 effectively protected rat liver tissues against the DMN-induced hepatic fibrosis. Antioxidant could be considered as a supplementary therapeutic for alleviating the hepatic fibrosis.
Animals ; Antioxidants/*pharmacology ; Artemisia ; Dimethylnitrosamine ; English Abstract ; Immunohistochemistry ; Liver/drug effects/metabolism/pathology ; Liver Cirrhosis, Experimental/chemically induced/metabolism/*pathology ; Plant Extracts/*pharmacology ; Rats ; Rats, Sprague-Dawley

To investigate the influence of bear bile on rat hepatocarcinoma induced by diethylnitrosamine (DEN), a total of 40 rats were randomly divided into 4 groups: normal control group, model group, and two bear bile treatment groups. The rat liver cancer model was induced by breeding with water containing 100 mg x L(-1) DEN for 14 weeks. The rats of the bear bile groups received bear bile powder (200 or 400 mg x kg(-1)) orally 5 times per week for 18 weeks. The general condition and the body weight of rats were examined every day. After 18 weeks the activities of serum alanine transaminase (ALT), aspartate transaminase (AST) and total bilirubin (TBIL) were detected. Meanwhile, the pathological changes of liver tissues were observed after H&E staining. The expression of proliferative cell nuclear antigen (PCNA) and a-smooth muscle actin (alpha-SMA) in liver tissue were detected by immunohistochemical method. After 4 weeks the body weights of rats in normal group were significantly more than that in other groups (P < 0.05); and that in the two bile groups was significantly more than that in the model group. Compared with normal group, the level of serum glutamic-pyruvic transaminase and total bilirubin increased significantly in other groups; compared with model group, these two indexes decreased significantly in two bile groups. Hepatocellular carcinoma occurred in all rats except for normal group; there were classic cirrhosis and cancer in model group while there were mild cirrhosis and high differentiation in two bile groups. There were almost no expressions of PCNA and alpha-SMA in normal group while there were high expressions in model group; the two bile groups had some expressions but were inferior to the model group, and alpha-SMA reduced markedly. It indicated that bear bile restrained the development of liver cancer during DEN inducing rat hepatocarcinoma, which may be related to its depressing hepatic stellate cell activation and relieving hepatic lesion and cirrhosis.
Actins ; metabolism ; Alanine Transaminase ; blood ; Animals ; Antineoplastic Agents ; pharmacology ; Aspartate Aminotransferases ; blood ; Bile ; chemistry ; Bilirubin ; blood ; Body Weight ; drug effects ; Carcinoma, Hepatocellular ; blood ; chemically induced ; pathology ; Diethylnitrosamine ; Liver ; metabolism ; pathology ; Liver Cirrhosis ; chemically induced ; pathology ; Liver Neoplasms, Experimental ; blood ; chemically induced ; pathology ; Male ; Powders ; pharmacology ; Proliferating Cell Nuclear Antigen ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Ursidae

OBJECTIVETo investigate the anti-fibrogenesis property of intraportal vein injection of small interfering RNA targeting connective tissue growth factor (CTGF) in a rat model of liver fibrosis and its effect on the accumulation of extracellular matrix (ECM).

METHODSThirty male rats were randomly divided into five groups. Some rats received CCl4 subcutaneously every three days for 6 consecutive weeks, and in the meantime they also received either siRNA targeting CTGF (preventive group), saline (model group) or siRNA (siRNA control group) by intraportal vein injections. Other rats received CCl4 by subcutaneous injection for 2 weeks, followed by CCl4 and CTGF siRNA intraportal vein injection for 4 more weeks (as treatment group). The expressions of CTGF and type I and III collagen genes were detected by means of reverse transcription-polymerase chain reaction (RT-PCR) and/or Western blot respectively. Hepatic histology was evaluated by HE and Sirius red stained sections. The collagen staining areas were measured quantitatively using a computer-aided manipulator with slight modifications. Serum procollagen type III and hyaluronic acid were determined by radioimmunoassay.

RESULTSSix weeks after CCl4 injection, prominent upregulation of gene expressions of CTGF, type I and III collagen, and laminin in saline or siRNA-treated rat livers were observed. The expressions of CTGF at mRNA and protein level and type I and III collagen at mRNA level were markedly reduced in rats with CTGF siRNA treated for four or six weeks. Expressions of CTGF at mRNA and protein levels decreased by 76%+/-8%, 80%+/-3% (F = 68.630) and 95%+/-2%, 93%+/-3% (F = 21.234, P < 0.01); type I and III collagen and laminin at mRNA levels decreased by 74%+/-8%, 78%+/-8%, 31%+/-7% and 57%+/-6%, 59%+/-10%, 43%+/-9% (F = 24.219, 16.315, 9.716, P < 0.01) compared with rats in the model group at 72 h. The CTGF siRNA treatment markedly reduced serum levels of procollagen type III and hyaluronic acid and the degrees of liver fibrosis.

CONCLUSIONIntraportal vein siRNA injection targeting CTGF could significantly inhibit CTGF gene expression in rats, thereby attenuating liver fibrosis by reducing ECM accumulation.

Animals ; Carbon Tetrachloride ; Connective Tissue Growth Factor ; metabolism ; Extracellular Matrix ; metabolism ; Gene Silencing ; Liver ; metabolism ; pathology ; Liver Cirrhosis, Experimental ; chemically induced ; metabolism ; pathology ; Male ; RNA, Small Interfering ; Rats ; Rats, Sprague-Dawley

To study the effects of Smad4 on liver fibrosis and hepatocarcinogenesis in mice treated with CCl(4)/ethanol. The wild-type mice (Smad4 +/+) and the Smad4 knockout mice (Smad4 +/-) were injected subcutaneously with carbon tetrachloride(CCl(4))/ethanol twice a week for twenty weeks. The expression of Smad4, TGFbeta1, Smad2, Smad3, Smad6, TIMP1, MMP2 and MMP9 was detected by RT-PCR. In the cirrhotic liver, the expression of Smad4 mRNA was significantly higher than that in the normal liver. Comparing with wild-type mice (Smad4 +/+), the TGFbeta1-Smad4 signaling was markedly attenuated in the Smad4 knockout mice (Smad4 +/-). After induction by CCl(4)/ethanol, the hepatic fibrosis in the Smad4 knockout mice (Smad4 +/-) was obviously alleviated compared with the wild-type mice (Smad4 +/+), and the incidence rate of hepatocarcinogenesis of the former was also lower than that of the latter(32.0% vs 41.9%). These results indicate that knocking out Smad4 can delay the progression of liver fibrosis and liver cancer.
Animals ; Carbon Tetrachloride ; administration & dosage ; Disease Models, Animal ; Ethanol ; administration & dosage ; Female ; Liver Cirrhosis, Experimental ; chemically induced ; metabolism ; pathology ; Liver Neoplasms, Experimental ; chemically induced ; metabolism ; pathology ; Male ; Mice ; Mice, Knockout ; RNA, Messenger ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; Smad Proteins ; genetics ; metabolism ; Smad4 Protein ; genetics ; metabolism ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; metabolism ; Transforming Growth Factor beta1 ; genetics ; metabolism

Animals ; Antioxidants ; therapeutic use ; Catalase ; metabolism ; Drugs, Chinese Herbal ; therapeutic use ; Liver ; chemistry ; pathology ; Liver Cirrhosis, Experimental ; chemically induced ; drug therapy ; metabolism ; Male ; Phytotherapy ; Rats ; Rats, Wistar ; Serum ; Superoxide Dismutase ; metabolism ; Swine

OBJECTIVETo study the changes of advanced glycation end products (AGEs) in different phases of a rat liver fibrosis model induced by CCl4, and the interventional effect of aminoguanidin (AG).

METHODSFifty-four SD rats were divided into three groups: a control group, a CCl4 model group and an intervention group. Their blood serum AGEs and hyaluronic acid (HA) and AGEs in their liver homogenates were measured. These measurements were correlatively assessed to the degrees of liver fibrosis at different phases of the rat model before and after the intervention with aminoguanidin.

RESULTSThe content of AGEs in their blood sera and liver homogenates, and the level of blood serum HA, and the score of liver fibrosis degree at week 12 in our rat liver fibrosis mode groups were significantly higher than those in the control group (P < 0.01). In the intervention group with aminoguanidin, these figures were lower than those in the liver fibrosis model group (P < 0.05). The content of AGEs in their blood sera and liver homogenates had a linear correlation with the level of HA in their blood sera.

CONCLUSIONThe contents of AGEs in their blood sera and liver homogenates were increased in the late phase of our rat liver fibrosis model. To some extent, the level of AGEs may reflect the fibrosis degree of the rat livers. Aminoguanidin has an interventional effect in our CCl4 induced rat liver fibrosis model.

Animals ; Carbon Tetrachloride ; Carbon Tetrachloride Poisoning ; Glycation End Products, Advanced ; metabolism ; Guanidines ; therapeutic use ; Liver ; metabolism ; pathology ; Liver Cirrhosis, Experimental ; chemically induced ; drug therapy ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate the recipe-based pathogenesis and effects of Xiayuxue Decoction (XD), Yinchenhao Decoction (YcD), Yiguanjian Decoction (YgD) and Huangqi Decoction (HD) on carbon tetrachloride (CCl4) induced liver cirrhosis formation in rats on the basis of the recognition of basic pathogenesis of liver cirrhosis in TCM and train of thoughts of detecting the TCM syndrome by recipe.

METHODSModel rats of liver cirrhosis were established by subcutaneous injecting of 100% CCl4 3ml/kg followed by 50% CCl4 olive solution 2ml/kg, twice a week for 12 weeks. They were randomly divided into the model group, the XD treated group, the YcD treated group, the YgD treated group and the HD treated group. Rats in the three treated group received the treatment starting from the 9th week of modeling with the corresponding decoctions. All animals were sacrificed by the end of the 12th week, and their hepatic function, liver pathological changes and hydroxyproline (Hyp) content of hepatic tissue were detected.

RESULTS(1) Typical chronic liver injury and fibrosis became evident in the model rat at the 8th week and cirrhosis came into being at the 12th week. (2) Compared with the rats in the model group, hepatic pathological changes were alleviated significantly, content of Hyp in hepatic tissue was decreased markedly and hepatic function improved remarkably in the XD group and YgD group. The improvement in the XD group was superior to that in the YgD group, while the serum albumin level elevated more significant in the YgD group.

CONCLUSIONThe main pathological changes during CCl4 induced liver cirrhosis formation in rats is the rapid hyperplasia of hepatic fibrous connective tissue and obstruction of collaterals by blood stasis, thus induced reconstruction of the tissue structure, which could be treated with XD effectively, while the severe injury of liver parenchyma in this phase is another pathological change of Gan-yin deficiency syndrome, which could be effectively treated with YgD by its Yin-nourishing action.

Animals ; Carbon Tetrachloride ; Diagnosis, Differential ; Drugs, Chinese Herbal ; therapeutic use ; Hydroxyproline ; metabolism ; Liver ; drug effects ; metabolism ; pathology ; Liver Cirrhosis, Experimental ; chemically induced ; diagnosis ; drug therapy ; Male ; Medicine, Chinese Traditional ; Phytotherapy ; Random Allocation ; Rats ; Rats, Wistar ; Yin Deficiency ; drug therapy

OBJECTIVETo study the inhibitory effect of flavonoids from Glycyrrhiza uralensis on thioacetamide-induced chonic hepatic fibrosis in rats and the effect on the protein expressions of transforming growth factor-β1 (TGF-β1) and Caspase-3 in livers.

METHODMale Sprague-Dawley rats were randomly divided into totally seven groups: the normal control group, the model group, LF groups s (400, 200, 100, 50 mg · kg(-1) · d(-1)) and the silymarin positive control group (30 mg · kg(-1) · d(-1)). The hepatic fibrosis model was induced in the rats through intraperitoneal injection with 3% thioacetamide (TAA) at a dose of 150 mg · kg(-1) body weight twice a week for 12 weeks. During the course, the control group and the model group were orally administered with saline (1 mL · kg(-1) · d(-1)). After the modeling and drug intervention, the pathologic changes and fibrosis in liver tissues were observed by HE staining and Masson's Trichrome staining. The serum alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP) and liver hydroxyproline (HYP) contents were assayed by biochemical process. The serum hyaluronic acid (HA) was assessed by radioimmunoassay. In addition, the protein expressions of liver TGF-β1 and Caspase-3 were examined by immunohistochemical method. The mRNA expression of TGF-β1 in hepatic tissues was examined by quantitative Real-time PCR analysis.

RESULTCompared with the model group, flavonoids can protect the integrity of the structure of liver tissues, significantly reduce the hepatic cell degeneration and necrosis and the proliferation of fibrous tissues, notably reduce the serum AST, ALT, ALP and HA and HYP in hepatic tissues and down-regulate the protein expressions of liver TGF-β1 and Caspase-3 and the mRNA expression of TGF-β1 in hepatic tissues.

CONCLUSIONThe licorice flavonoids can resist the thioacetamide-induced hepatic fibrosis in rats. Its mechanism may be related to the down-regulation of the protein expressions of TGF-β1 and Caspase-3.

Animals ; Caspase 3 ; analysis ; Flavonoids ; pharmacology ; Glycyrrhiza uralensis ; chemistry ; Hyaluronic Acid ; blood ; Liver ; pathology ; Liver Cirrhosis, Experimental ; chemically induced ; metabolism ; prevention & control ; Male ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Thioacetamide ; Transforming Growth Factor beta1 ; analysis ; genetics