Long Non-Coding RNA ANRIL Regulates Inflammatory Factor Expression in Ulcerative Colitis Via the miR-191-5p/SATB1 Axis.

Ulcerative colitis, an inflammatory bowel disease, manifests with symptoms such as abdominal pain, diarrhea, and mucopurulent feces. The long non-coding RNA (lncRNA) ANRIL exhibits significantly reduced expression in UC, yet its specific mechanism is unknown. This study revealed that ANRIL is involved in the progression of UC by inhibiting IL-6 and TNF-α via miR-191-5P/SATB1 axis. We found that in patients with UC, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were significantly overexpressed in inflamed colon sites, whereas ANRIL was significantly under-expressed and associated with disease severity. The downregulation of ANRIL resulted in the increased expression of IL-6 and TNF-α in LPS-treated FHCs. ANRIL directly targeted miR-191-5p, thereby inhibiting its expression and augmenting SATB1 expression. Moreover, overexpression of miR-191-5p abolished ANRIL-mediated inhibition of IL-6 and TNF-α production. Dual luciferase reporter assays revealed the specific binding of miR-191-5p to ANRIL and SATB1. Furthermore, the downregulation of ANRIL promoted DSS-induced colitis in mice. Together, we provide evidence that ANRIL plays a critical role in regulating IL-6 and TNF-α expression in UC by modulating the miR-191-5p/SATB1 axis. Our study provides novel insights into progression and molecular therapeutic strategies in UC.

Budesonide foam for mild to moderate distal ulcerative colitis: A systematic review and meta-analysis.

BACKGROUND AND AIM: Budesonide is a second-generation steroid with prominent topical effects and minimal systemic activity for patients with ulcerative colitis (UC). We perform a systematic review and meta-analysis of randomized placebo-controlled trials to assess the efficacy and safety of budesonide foam in mild-to-moderate distal UC. METHODS: Comprehensive searches were performed to identify all eligible studies. Outcome measures were clinical remission, endoscopic improvement, elimination of rectal bleeding, and adverse events. The risk ratio (RR) with 95% confidence interval (CI) was estimated for each outcome. All statistical analyses were performed in STATA 12.0. RESULTS: Three randomized placebo-controlled trials recruiting 711 patients with mild-to-moderate distal UC were included in this study. No significant bias and heterogeneity was identified. Pooled analyses showed that budesonide foam was significantly superior to placebo for induction of clinical remission (RR = 1.83, 95%CI: 1.41, 2.37; P < 0.001) and endoscopic improvement (RR = 1.44, 95%CI: 1.23, 1.68; P < 0.001), and eliminating rectal bleeding at week 2 (RR = 2.00, 95%CI: 1.50, 2.66; P < 0.001), week 4 (RR = 1.73, 95%CI: 1.42, 2.12; P < 0.001), and week 6 (RR = 1.76, 95%CI: 1.45, 2.14; P < 0.001). No statistically significant difference was observed in the incidence of treatment-related adverse events and therapeutic discontinuation because of adverse events between budesonide foam and placebo. CONCLUSIONS: Budesonide foam is well tolerated and superior to placebo in inducing clinical remission and endoscopic improvement, and eliminating rectal bleeding for mild-to-moderate distal UC.

Aquaporin 9 inhibits hepatocellular carcinoma through up-regulating FOXO1 expression.

Aquaporin 9 (AQP9) is the main aquaglyceroporin in the liver. Few studies have been performed regarding the role of AQP9 in liver cancer. Here we report AQP9 expression and function in liver cancer. We found that AQP9 mRNA and protein levels were reduced in human hepatocellular cancer compared to the para-tumor normal liver tissues. Human hepatoma cell line SMMC7721 expressed low basal levels of AQP9. When AQP9 was overexpressed in SMMC7721 cell line, cell proliferation was inhibited due to cell cycle arrest at G1 phase and increased apoptosis. At the molecular level, AQP9 overexpression decreased the protein levels of phosphatidylinositol-3-kinase (PI3K), leading to reduced phosphorylation of Akt. Subsequently, the protein levels of forkhead box protein O1 (FOXO1) were increased, resulting in down-regulation of proliferating cell nuclear antigen (PCNA) expression and up-regulation of caspase-3 expression. AQP9 overexpression inhibited growth of subcutaneously xenografted liver tumors in nude mice. These findings suggest that AQP9 expression is down-regulated in liver cancer compared to the normal liver tissue and restoration of AQP9 expression can inhibit development of liver cancer.

Efficacy and safety of transarterial chemoembolization plus sorafenib for early or intermediate stage hepatocellular carcinoma: A systematic review and meta-analysis of randomized controlled trials.

BACKGROUND AND AIM: The efficacy and safety of transarterial chemoembolization (TACE) plus sorafenib for patients with hepatocellular carcinoma (HCC) have been explored by many studies, but the results were controversial. Therefore, we performed this meta-analysis of high-quality randomized controlled trials to evaluate the efficacy and safety of TACE plus sorafenib versus TACE monotherapy in the early or intermediate stage HCC. METHODS: Multi-databases were systematically searched to identify all eligible literatures. The hazard ratio (HR) or risk ratio (RR) with 95% confidence intervals (95%CIs) for time to progression (TTP), overall survival (OS), objective response rate (ORR), disease control rate (DCR) and the incidence of treatment-related adverse events (AEs) were pooled using a fixed or random effect model in STATA 12.0. RESULTS: Four randomized controlled trials, including a total of 887 patients with early or intermediate stage HCC, were included in this meta-analysis. The pooled results showed that TACE plus sorafenib significantly improved TTP (HR=0.77, 95% CI: 0.64-0.92; P=0.005). Nevertheless, the OS (HR=0.97, 95% CI: 0.72-1.29; P=0.828), ORR (RR=1.20, 95% CI: 0.88-1.64; P=0.257) and DCR (RR=1.04, 95% CI: 0.90-1.02; P=0.568) were not improved. The incidence of treatment-related AEs was higher in the TACE plus sorafenib. CONCLUSIONS: Evidences from the meta-analysis of high-quality randomized controlled trials indicate that TACE plus sorafenib can significantly improve TTP but not OS, ORR and DCR in early or intermediate stage HCC. In addition, the combination therapy increases the adverse events which usually disturb the treatment progress and should be increased attention.

Aquaporin 9 is down-regulated in hepatocellular carcinoma and its over-expression suppresses hepatoma cell invasion through inhibiting epithelial-to-mesenchymal transition.

Aquaporin 9 (AQP9) is the main aquaglyceroporin in the liver. Few studies have been performed regarding the role of AQP9 in hepatocellular carcinoma (HCC). Here, we report the expression and function of AQP9 in HCC tissues and cell lines. We found that AQP9 mRNA and protein levels were down-regulated in HCC tissues and human hepatoma cell lines compared to the para-cancer normal liver tissues and normal hepatocyte line, respectively. In a human HCC SMMC-7721 cell line, over-expression of AQP9 suppressed cell invasion in vitro and xenograft tumor growth in vivo. AQP9 over-expression increased the expression of E-cadherin and decreased the expression of N-cadherin in SMMC-7721 cells and xenografted tumors, which was correlated with decreased levels of phosphoinositide 3-kinase (PI3K) and p-Akt. Conversely, using siRNA to knock down AQP9 over-expression could reverse the phenotype caused by AQP9 over-expression. Our findings suggest that AQP9 is down-regulated in hepatocellular carcinoma and its over-expression suppresses hepatoma cell invasion through inhibiting epithelial-to-mesenchymal transition.

Expression and clinical significance of aquaglyceroporins in human hepatocellular carcinoma.

Aquaglyceroporins (AQPs) are a subset of the aquaporin family, and are permeable to water and glycerol. The aim of the present study was to determine the expression and clinical significance of three AQPs, AQP3, 7 and 9 in hepatocellular carcinoma (HCC). Fresh HCC and adjacent non­tumorous liver tissues were collected from 68 patients diagnosed with HCC. The expression levels of AQP3, 7 and 9 were detected by reverse transcription­quantitative polymerase chain reaction, western blotting and immunohistochemical analysis. The association between the expression of AQPs and clinicopathological parameters of HCC were investigated. Compared with non­tumorous liver tissue, HCC tissues exhibited a significant (P<0.05) increase in the expression of AQP3 and a concomitant reduction in the expression levels of AQP7 and AQP9, at both the mRNA and protein levels. Immunohistochemistry revealed that AQP9 was dominantly localized on the plasma membrane of hepatocytes, while AQP3 and AQP7 exhibited a predominantly cytoplasmic and nuclear distribution. High expression of AQP3 was significantly (P<0.05) associated with low expression levels of AQP7 and AQP9. High expression of AQP3 was correlated with tumor grade (P=0.017), tumor stage (P=0.010) and lymphatic metastasis (P=0.031). Low expression of AQP7 was correlated with tumor grade (P=0.043). AQP3 was upregulated, and AQP7 and AQP9 were downregulated in HCC. A high expression of AQP3 and low expression of AQP7 was significantly associated with the aggressive features of HCC.

FOXO1-mediated epigenetic modifications are involved in the insulin-mediated repression of hepatocyte aquaporin 9 expression.

Aquaporin (AQP) 9 transports glycerol and water, and belongs to the aquaglyceroporin subfamily. Insulin acts as a negative regulator of AQP9, and FOXO1 has the ability to mediate the regulatory effects of insulin on target gene expression. The aim of the present study was to determine whether insulin­induced repression of AQP9 involved an epigenetic mechanism. HepG2 human hepatocyte cells were treated with 500 µM insulin for different durations. AQP9 mRNA expression levels were determined by quantitative polymerase chain reaction (qPCR), and histone H3 acetylation, phosphorylation and methylation at the insulin responsive element (IRE) of the AQP9 promoter was assessed using chromatin immunoprecipitation coupled with qPCR. The effects of lentiviral FOXO1 overexpression on AQP9 expression levels and H3 modifications at the AQP9 promoter were also determined. The insulin treatment resulted in a significant and time­dependent reduction in AQP9 mRNA expression levels in HepG2 cells, as compared with untreated cells (P<0.05). In the insulin­treated cells, the levels of H3 acetylation and phosphorylation were significantly reduced (P<0.05), but the level of H3 methylation was increased. Enforced expression of FOXO1 increased AQP9 mRNA and protein expression levels in HepG2 cells. Furthermore, FOXO1 overexpression promoted H3 acetylation and phosphorylation, and reduced H3 methylation at the IRE locus of the AQP9 promoter. These data provide, to the best of our knowledge, the first evidence that insulin­induced transcriptional suppression of AQP9 expression in hepatocytes involves FOXO1­mediated H3 modifications at the IRE locus in the promoter.

[Expression of aquaporin 3 and aquaporin 9 is regulated by oleic acid through the PI3K/Akt and p38 MAPK signaling pathways].

OBJECTIVE: To study the effect of oleic acid (OA) on expression of aquaglyceroporin genes, AQP3 and AQP9, in hepatocyte steatosis and to investigate the underlying molecular mechanisms using an in vitro system. METHODS: HepG2 cells were treated with OA at different concentration to establish in vitro models of nonalcoholic hepatocyte steatosis. The corresponding extents of hepatic steatosis modeling were assessed by oil red O staining and optical density (OD) measurements of the intracellular fat content. The model lines were then treated with inhibitors of the PI3K/Akt and p38 MAPK signaling pathway factors and effects on AQP3/9 expression was measured by real time RT-PCR and western blotting. RESULTS: The fat concentration, indicative of hepatic steatosis, increased in conjunction with increased concentrations of OA (0 less than 250 less than 500 mumol/L). OA exposure also down-regulated AQP3 mRNA and up-regulated AQP9 mRNA levels in a concentration-dependent manner. The most robust changes in expression occurred in response to the 500 mumol/L concentration of OA for both AQP3 (0.47+/-0.18; t = 4.5450, P less than 0.05) and AQP9 (1.57+/-0.21; t = 3.0306, P less than 0.05). Treatment with OA + PI3K pathway inhibitor (LY294004) significantly decreased AQP9 mRNA expression (4.55+/-0.62) as compared to the control group (1.00+/-0.10; t = 9.7909, P less than 0.01), that 500 mumol/L OA group (2.43+/-0.53; t = 4.5018, P less than 0.05), and the LY294002 group (1.90+/-0.16; t = 7.1683, P less than 0.01). Treatment with p38 MAPK pathway inhibitor (SB230580) significantly increased the OA-suppressed level of AQP3 mRNA to the level detected in the control group (1.27+/-0.11; t = 5.7455, P less than 0.01) and decreased the OA-stimulated AQP9 mRNA (0.38+/-0.09; t = 6.5727, P less than 0.01). No significant changes in mRNA expression of AQP3/9 were observed with inhibition of the ERK1/2 and JNK signal transduction pathways. The OA-induced changes in protein expression levels of AQR3 and AQP9 followed a similar trend of the genes. Finally, OA suppressed the level of phosphorylated Akt (from 0.21+/-0.02 to 0.13+/-0.03; t = 3.8431, P less than 0.05) but elevated the level of phosphorylated p38 (from 0.58+/-0.06 to 1.02+/-0.10; t = 12.5289, P less than 0.01). Again, OA treatment produced no significant affect on ERK1/2 and JNK phosphorylation. CONCLUSION: OA down-regulates AQP3 expression by stimulating the p38 MAPK signaling pathway, and up-regulates the AQP9 by blocking the PI3K/Akt pathway and activating the p38 MAPK signaling pathway.

Experimental studies and clinical experiences on treatment of secondary hypersplenism with extracorporeal high-intensity focused ultrasound.

The aim of this study is to investigate the efficacy and safety of extracorporeal high-intensity focused ultrasound (HIFU) in treatment of hypersplenism. Fifteen adult dogs, weighing 13-18 kg were divided into three groups: sham group, SVL group undergoing splenic vein ligation (SVL) after laparotomy, and SVL + HIFU group receiving SVL followed by extracorporeal HIFU. Pathologic and hematologic analyses were performed. We also reviewed the clinical data of 19 patients with secondary hypersplenism caused by liver cirrhosis or hepatocellular carcinoma who underwent extracorporeal HIFU. Extracorporeal HIFU significantly diminished the volume of the spleen of animals, coupled with occurrence of coagulation necrosis and fibrosis in the target area. Both platelet and red blood cell counts were significantly restored by HIFU intervention. Similarly, HIFU treatment improved the hematologic parameters in patients with hypersplenism, and no major complications were encountered. Extracorporeal HIFU intervention is effective and safe in managing secondary hypersplenism.

Oral immunization with recombinant Mycobacterium smegmatis expressing the outer membrane protein 26-kilodalton antigen confers prophylactic protection against Helicobacter pylori infection.

Helicobacter pylori infection is prevalent worldwide and results in chronic gastritis, which may lead to gastric mucosa-associated lymphoid tissue lymphoma and gastric cancer. We have previously reported that oral immunization with recombinant Mycobacterium smegmatis expressing the H. pylori outer membrane protein 26-kilodalton (Omp26) antigen affords therapeutic protection against H. pylori infection in mice. In the present study, we investigated the prophylactic effects of this vaccine candidate on H. pylori challenge in mice. We found that oral immunization with recombinant Mycobacterium Omp26 significantly reduced H. pylori colonization in the stomach compared to inoculation with wild-type M. smegmatis in control mice. Six of the recombinant Mycobacterium-immunized mice (60%) were completely protected from H. pylori infection. The severity of H. pylori-associated chronic gastritis assessed histologically was significantly milder in mice vaccinated with recombinant Mycobacterium than in control animals. Mice immunized with recombinant Mycobacterium showed enhanced antigen-specific lymphocyte proliferation and antibody responses. Moreover, immunization with recombinant Mycobacterium resulted in an increased expression of interleukin-2 and gamma interferon in the stomach and spleen, as determined by reverse transcription-PCR analysis. Our results collectively suggest that vaccination with recombinant Mycobacterium Omp26 confers prophylactic protection against H. pylori infection. The inhibition of H. pylori colonization is associated with the induction of antigen-specific humoral and cell-mediated immune responses.

[Effects of insulin and LY294002 inhibitors of PI3K on the regulations and expression of aquaporin 9 in normal liver cells].

OBJECTIVES: To explore the effects of insulin on the expression and the regulatory pathway of AQP9 in normal human liver cells. METHODS: Normal human liver cells L02 were cultured and treated with PI3K inhibitor LY294002, AKT inhibitor A-443654, MAPK inhibitors SB2030580 and insulin at different concentrations respectively. The AQP9 mRNA and protein expressions were detected with semi-quantitative RT-PCR and Western blot respectively. RESULTS: The insulin (100 nmol/L approximately 500 nmol/L) treatment decreased the expression of AQP9 in normal human liver cells (P less than 0.05) concentration dependently, and the expression of AQP9 began to reduce from 3 hours of insulin stimulation (P less than 0.05), especially at insulin treatment for 12 hours (P less than 0.05); Incubated with the selective inhibitor of PI3K (LY294002) and AKT (A-443654), the inhibitory effects of insulin on AQP9 expression decreased (P less than 0.05); but it did not change significantly by blocking the MAPK signaling pathway. CONCLUSION: The insulin treatment inhibited the expression of AQP9 and the PI3K/akt signal transduction pathway was involved in the mechanism.

[Efficacy of endoscopic variceal ligation and its correlation with liver function].

OBJECTIVE: To analyze the efficacy of endoscopic variceal ligation and its correlation with liver function. METHODS: 322 patients received EVL (endoscopic variceal ligation) and 34 patients with PDP (pericardial devascularization procedure) were retrospectively analyzed and divided into groups A, B and C. These patients were then subdivided into bleeding and non-bleeding subgroups according to Child-Pugh scores of liver function and history of upper gastrointestinal bleeding. The bleeding rate and mortality were contrasted between EVL and PDP. Liver function, Platelet count, leucocyte count and spleen thickness of before and after ligation were contrasted in EVL. RESULTS: The bleeding rate and mortality were 1.7%, 3.4%, 7.0%; 0%, 5.1%, 8.1% in EVL group and 9.1%, 14.3%, 100.0%; 0%, 9.5%, 50.0% in PDP group, respectively. Variceal obliteration needed means of 2.1+/-0.7, 3.1+/-0.8 and 4.2+/-1.2 sessions in A, B and C ligation groups, respectively (F = 41.2, P is less than 0.01). On subgroup analysis, the numbers of ligation session were 2.6+/-0.7, 3.2+/-0.9 and 4.3+/-1.1 in A, B and C bleeding subgroup (F = 39.3, P value is less than 0.01) and 2.0+/-0.6, 2.7+/-0.6, and 2.9+/-0.4 in A, B and C non-bleeding subgroup, respectively (F = 17.0, P value is less than 0.01). ALT, AST, Platelet count and leucocyte count reduced significantly, spleen thickness increased remarkably in bleeding subgroup after ligation. CONCLUSION: The efficacy of EVL was significantly negatively correlated with liver function and prior to pericardial devascularization procedure. EVL had no effect on liver function but might increase spleen thickness and aggravate hypersplenism. EVL was recommended especially for the bleeding liver cirrhosis patients with Child B and C scores.

Inhibition of hepatitis B virus replication by small interference RNA induces expression of MICA in HepG2.2.15 cells.

Hepatitis B virus (HBV) replicates in most tumor tissues of patients with HBV-associated hepatocellular carcinoma (HCC). In the present study, we have shown that the expression of HBV in the HCC cell lines, HepG2 and Huh7, down-regulated the expression of MHC class I-related molecule A (MICA), a ligand of the NKG2D receptor. Inhibition of HBV expression by small interference RNAs (siRNAs) in HepG2.2.15, a cell line that constitutively expresses HBV, induced up-regulation of MICA. The up-regulation of MICA increased the lysis of HepG2.2.15 cells by NK cells. Our results suggest that HBV compromises the innate immune system in HCC patients and that inhibition of HBV replication by siRNAs may enhance the antitumor immune response.

Angiotensin-converting enzyme inhibitors improve hepatic steatosis by modulating expression of tumour necrosis factor-alpha, interleukin-6 and adiponectin receptor-2 in rats with type 2 diabetes.

1. Angiotensin-converting enzyme inhibitors (ACEI) are hypotensive drugs that have been shown to prevent Type 2 diabetes mellitus (T2DM) in high-risk individuals. However, in T2DM, the effects of ACEI on hepatic steatosis are not known. The aim of the present study was to examine the effects of ACEI on changes in liver histology and hepatic mRNA expression of adipokines in rats with T2DM. 2. Thirty-six rats were divided into a normal control group, a T2DM group and a fosinopril-treated group. After six weeks of treatment with 5 mg/kg per day fosinopril, an ACEI, changes in liver histology, serum fasting glucose (FG), insulin, triglyceride (TG), total cholesterol (TC), alanine aminotransferase (ALT), aspartate aminotransferase (AST), tumour necrosis factor (TNF)-alpha, interleukin (IL)-6, adiponectin were evaluated, as was hepatic TNF-alpha, IL-6 and adiponectin receptor-2 (adipoR2) mRNA expression. 3. The degree of hepatic steatosis and inflammation, serum FG, insulin, TG, TC, ALT, TNF-alpha and IL-6 concentrations and hepatic TNF-alpha and IL-6 mRNA expression were significantly higher in rats with T2DM than in normal controls. Serum adiponectin concentrations and hepatic adipoR2 mRNA expression in rats with T2DM were significantly lower than in normal controls. Fosinopril significantly reduced the degree of hepatic steatosis, serum FG, insulin, ALT, TNF-alpha and IL-6 concentrations and hepatic TNF-alpha and IL-6 mRNA expression. Fosinopril significantly increased serum adiponectin concentrations and hepatic adipoR2 mRNA expression. 4. In conclusion, the ACEI improved insulin sensitivity and hepatic steatosis in rats with T2DM by increasing circulating adiponectin and hepatic adipoR2 levels, in addition to reducing pro-inflammatory cytokine levels in the circulation and liver.

[The effectiveness of endoscopic variceal ligation in patients with different grades of liver function].

OBJECTIVE: To evaluate the effectiveness of endoscopic variceal ligation (EVL) in patients with different grades of liver function. METHODS: MELD scores were determined for 156 patients before their EVL operations. After the EVL these patients were followed-up and their survival rates were analyzed. RESULTS: Fifty percent of the patients whose MELDs were less than or equal to 7 survived longer than 45 months after the EVL; in those with MELDs between 7 and 9, 50% of the patients survived 47.34 months; however, the figure for those whose MELD were more than 9 survived only 24.89 months. In the first two groups, 50% of the patient' survival duration was significantly longer than that of the third group. The difference was statistically significant. CONCLUSION: EVL becomes an effective clinical way to treat hemorrhage of esophagus varicose veins. The survival rate for this procedure is directly correlated with the liver function of the patient before the EVL.

[Evaluation of the effects of dense endoscopic ligation for bleeding esophageal varices].

OBJECTIVE: To evaluate the short-term and long-term effects of dense endoscopic variceal ligation (DEVL) for bleeding esophageal varices. METHODS: Patients with acute or with a history of esophageal variceal bleeding underwent regular DEVLs with a 2-3 week interval between 2 sessions until their varices disappeared at the lower 5-6 cm part of the esophagus. Follow-up study and gastroscopy were performed at 3, 6 and 12 months after the final DEVL in all patients. The results at 3 months were classified as short-term effects and those after 6 months as long-term ones. RESULTS: 126 patients underwent DEVLs with 403 sessions and 3641 ligations; each patient was ligated with a mean of 3.2 sessions and at 28.9 points. The cure rate of acute variceal bleeding was 100.0%; short-term rate of variceal eradication was 94.4% and variceal rebleeding occurred in 3.9% patients. After a mean of 22.3 months follow-up period, the recurrence of esophageal varices was observed in 11.9% patients, but the variceal rebleeding rate was only 3.2% and no patients died from it. CONCLUSION: DEVL was very useful and effective in both short-term and long-term variceal eradication and prevention of variceal rebleeding.

Rhein inhibits liver fibrosis induced by carbon tetrachloride in rats.

AIM: To investigate the effect of rhein on liver fibrosis induced by the exposure of carbon tetrachloride (CCl4)/ethanol in rats. METHODS: Male Wistar rats were divided into four study groups (n=10 each group): healthy controls, CCl4/ethanol-injured rats left untreated, and CCl4/ethanol-injured rats treated with rhein of low-dose (25 mg/kg) and high-dose (100 mg/kg). Rhein was given once a day since rat received CCl4/ethanol injury. After administration of rhein for 6 weeks rats were killed. The following parameters were determined: the activity of alanine aminotransferase (ALT), hyalauronic acid (HA) and procollagen type III (PC-III) concentrations in serum, liver malondialdehyde (MDA) level, the degree of liver fibrosis, and the expression of alpha-smooth muscle actin (alpha-SMA) and transforming growth factor-beta1 (TGF-beta1) in liver tissue. RESULTS: The treatment of rhein markedly reduced the ALT activity, HA and PC-III concentrations, and liver MDA level in CCl4/ethanol-injured rats (P<0.01). It also improved significantly histological changes of fibrosis and decreased the expression of alpha-SMA and TGF-beta1 in liver of these rats (P<0.05 or P<0.01). CONCLUSION: Rhein has protective effect on liver injury and can inhibit liver fibrosis induced by CCl4/ethanol in rats. The mechanisms possibly contribute to its action of antioxidant and anti-inflammatory activity, also associated with its effect of inhibiting TGF-beta1 and suppressing the activation of hepatic stellate cells.