Rat Model Of Hepatic Fibrosis Research Articles

Inhibitory effects of oxyresveratrol on ERK and Smad1/2 phosphorylation and HSC activation in preventing carbon tetrachloride-induced rat liver fibrosis

Oxyresveratrol (ORes, trans-2,4,3′,5′-tetrahydroxy stilbene) naturally exists in mulberry, grapes, peanuts and other plants. It belongs to stilbene polyphenolic family and has an extra hydroxyl group at 2-position comparing with resveratrol (Res). Hence, ORes has stronger antioxidant activity than resveratrol. In present study, we employed a rat hepatic fibrosis model induced by carbon tetrachloride (CCl4) and administrated ORes via gavage feeding to study the protective effects and potential mechanisms of ORes against hepatic fibrosis. We demonstrated that rat liver oxidative damage induced by CCl4 was significantly alleviated after ORes feeding. Furthermore, the mRNA transcription levels of α-smooth muscle actinn (α-SMA), desmin, and two MMPs (MMP2 and MMP9) were reduced and the expression levels of transforming growth factor β1 (TGF-β1), p- small mother against decapen-taplegic protein (Smad)1/2 and p-extracellular signal-regulated kinases (ERK)1/2 in the liver tissue down-regulated dramatically. In a parallel study with Res, ORes showed more efficacious protective effect than Res against rat liver fibrosis, which is attributed to extended conjugation system due to the extra hydroxyl group at 2-position on ORes making it more electron-rich and susceptible to oxidation than Res. Therefore, dietary consumption of mulberry and other fruits containing ORes may be beneficial in the prevention of liver fibrosis.

Preventive and therapeutic effects of Qiwei Yugan Granule on hepatic fibrosis in rats

Objective: To investigate the preventive and therapeutic effects of Qiwei Yugan Granule (QYG) on hepatic fibrosis in rats based on MMP-13/TIMP-1 imbalance. Methods: The rats were randomly divided into control group, model group, colchicine group (1.0×10-4 g/kg) and QYG treated groups (3.7, 7.4, 14.8 g/kg) (n=8). The rat model of hepatic fibrosis was established by injected with carbon tetrachloride subcutaneously for 4 weeks and treated with ethanol by gavage for 6 weeks. The effects of QYG on liver function, histopathology of liver, related indexes of serum liver fibrosis, and MMP-13, TIMP-1 in hepatic tissue were observed. Results: QYG at the doses of 14.8、7.4、3.7 g/kg could significantly decrease the serum levels of ALT, AST, HA, PCⅢ and C-Ⅳ, relieve the pathological changes of hepatic fibrosis, increase the activity of MMP-13, decrease the activity of TIMP-1 and alleviate the imbalance of MMP-13/TIMP-1. Among them, QYG had a certain trend of dose-effect relationship with TIMP-1 and MMP-13/TIMP-1 (P＜0.05, 0.01). Conclusion: QYG has the effect of preventing and treating liver fibrosis and one of mechanisms is to promote MMP-13/TIMP-1 to restore balance.

The small heterodimer partner inhibits activation of hepatic stellate cells via autophagy.

Hepatic fibrosis is a health concern worldwide, and it is of great importance to develop effective therapeutic targets. The small heterodimer partner (SHP) is a regulator of lipid and bile acid metabolism in the liver. The objective of this study was to investigate the contribution of SHP to hepatic fibrosis and the underlying mechanism. An in vivo rat model of hepatic fibrosis was created through treatment with carbon tetrachloride. We used arginine-glycine-aspartic acid-poly (ethylene glycol)-polyethyleneimine (RGD-PEG-PEI) for the specific transfer of SHP into hepatic stellate cells (HSC). The level of gene expression was detected using quantitative real-time polymerase chain reaction (qRT-PCR). The LX2 cell line was selected for the in vitro assay. Artificial activation of LX2 in vitro was conducted through treatment with platelet-derived growth factor-BB (PDGF-BB), and autophagy was activated using rapamycin. Gain and loss of function assays were performed using a SHP-expressing plasmid or siRNA-SHP. Both qRT-PCR and western blotting were utilized to detect the level of gene expression. RGD-PEG-PEI-mediated the specific transduction of SHP into HSC in the liver and effectively increased the expression of SHP in the rat liver. After treatment with RGD-PEG-PEI-SHP, downregulation of liver fibrosis-associated genes was observed. The results of the in vitro assay indicated that SHP attenuated the stimulating effect of PDGF-BB on the activation of LX2 cells. Overexpression of SHP leads to significant downregulation of HSC activation-associated molecular factors, including α-smooth muscle actin, tissue inhibitor of metalloproteinase-1, and type I collagen. Conversely, increased expression of these molecules could be observed following knockdown of SHP. Furthermore, SHP affected fibrosis by inhibiting autophagy activated through treatment with rapamycin in LX2 cells. Overexpression of SHP may prevent liver fibrogenesis through inhibition of autophagy in HSC. The SHP may prevent liver fibrogenesis through inhibition of autophagy in HSC. A SHP-targeting therapy-based anti-fibrosis strategy possesses potential for application to the treatment of liver fibrosis.

Tetramethylpyrazine ameliorates hepatic fibrosis through autophagy-mediated inflammation.

Imbalanced immune response and hepatic fibrosis are key factors related to the progression of chronic liver diseases. Tetramethylpyrazine (TMP), a natural alkaloid, has been widely used for treating liver injury. In this study, we explored the effect of TMP on hepatic fibrosis and the related mechanisms regulating autophagy. A rat model of hepatic fibrosis and a model using an hepatic stellate cell line (HSC-T6) were created using CCl4 and platelet-derived growth factor (PDGF). Staining with haematoxylin and eosin (HE), Masson's stain, and TUNEL were performed for pathological diagnosis. ELISA, Western blotting, and immunofluorescence analyses were conducted to determine the expression levels of the specific markers for fibrosis, autophagy, inflammation, and signalling pathways. TMP treatment significantly rescued pathological injury and hepatic fibrosis. It also alleviated imbalances in the immune system, accumulation of extracellular matrix, and autophagy signals in hepatic fibrosis. At the same time, we found that application of the autophagy inducer rapamycin enhanced the therapeutic effect of TMP, whereas the autophagy inhibitor 3-methyladenine, PI3K pathway inhibitor LY294002, and AKT pathway agonist SC79 did the opposite. TMP exerts therapeutic effects in hepatic fibrosis mainly through promoting autophagy to ameliorate inflammation by inhibiting the AKT-mTOR signalling pathway, providing a new perspective for the treatment of chronic liver diseases.

Effects of Carvedilol on the Expression of TLR4 and its Downstream Signaling Pathway in the Liver Tissues of Rats with Cholestatic Liver Fibrosis.

This study was designed to investigate the effects of carvedilol on the expression of TLR4 and its downstream signaling pathway in the liver tissues of rats with cholestatic liver fibrosis and provide experimental evidence for clinical treatment of liver fibrosis with carvedilol. A total of fifty male Sprague Dawley rats were randomly divided into five groups (10 rats per group): sham operation (SHAM) control group, bile duct ligation (BDL) model group, low-dose carvedilol treatment group (0.1mg·kg-1·d-1), medium-dose carvedilol treatment group (1mg·kg-1·d-1), and high-dose carvedilol treatment group (10mg·kg-1·d-1). Rat hepatic fibrosis model was established by applying BDL. Forty-eight hours after the operation, carvedilol was administered twice a day. The blood and liver were simultaneously collected under the aseptic condition for further detection in two weeks after the operation. The alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBil) and albumin (Alb) in serum were measured. HE and Masson staining were used to determine hepatic fibrosis degree. Hydroxyproline assay was employed to detect liver collagen synthesis. Western Blot was used to measure the expression of TLR4, NF-κB p65 and β-arrestin2 protein. Quantitative analysis of TLR4, MyD88, TNF-α and IL-6 mRNA was performed by Realtime-PCR. Compared with the SHAM group, the BDL group showed obvious liver injury, increased levels of inflammatory factors, and continued progression of liver fibrosis. The above changes in the BDL group were alleviated in the carvedilol treatment groups. The improvement effects augmented as dosages increased. In addition, compared with the BDL group, the reduction of the expressions of TLR4, MyD88 and NF-κB p65 in liver tissues and the increase of the expression of β -arrestin2 in the high-dose carvedilol group were more significant. Carvedilol can reduce the release of inflammatory mediators by downregulating TLR4 expression and inhibiting its downstream signaling pathway, thus playing a potential therapeutic role in cholestatic liver fibrosis.

WITHDRAWN: Effects of carvedilol on expression of TLR4 and its downstream signaling pathway in liver tissue of rats with cholestatic liver fibrosisjaundice.

This study was designed to investigate the effects of carvedilol on the expression of TLR4 and its downstream signaling pathway in liver tissue of rats with cholestatic liver fibrosis, and provided experimental evidence for clinical treatment of liver fibrosis with carvedilol.? A total of fifty male Sprague Dawley rats were randomly divided into five groups (10 rats per group): sham surgery control group, bile duct ligation (BDL) model group, low-dose carvedilol treatment group (0.1mgkg-1d-1), medium-dose carvedilol treatment group (1mgkg-1d-1), high-dose carvedilol treatment group (10mgkg-1d-1). Rat hepatic fibrosis model was established by applying BDL. Forty-eight hours after the operation, carvedilol was administered twice a day. The blood and liver were simultaneously collected under the aseptic condition for further detection in two weeks after operation.? Compared with the sham group, the BDL group showed obvious liver injury, increased levels of inflammatory factors, and continued progression of liver fibrosis. Carvedilol could alleviate the above changes. The improvement effects were augmenting as dosages increasing. In addition, compared with the BDL group, carvedilol can reduce the expressions of TLR4, MyD88 and NF-?B p65 in liver tissue and increase the expression of ?-arrestin2, and the effect in the high dose group was more obvious. Carvedilol can reduce the release of inflammatory mediators by down-regulating TLR4 expression and inhibiting its downstream signaling pathway, thus playing a therapeutic role in cholestatic liver fibrosis.

Danshensu, a novel indoleamine 2,3-dioxygenase1 inhibitor, exerts anti-hepatic fibrosis effects via inhibition of JAK2-STAT3 signaling

BackgroundIndoleamine 2,3-dioxygenase 1 (IDO1), an important intracellular rate-limiting enzyme in the development of Hepatic fibrosis (HF), and has been proposed as a hallmark of HF. Danshensu (DSS) is a major bioactive component that isolated from a edible traditional Chinese medicinal herb Salviae Miltiorrhizae Radix et Rhizoma (Danshen), while, the anti-HF mode and mechanism of action of DSS have not been fully elucidated. MethodsCarbon tetrachloride (CCl4)-induced rat HF model and TGF-β1-induced hepatic stellate cell (HSC) model were employed to assess the in vivo and in vitro anti-HF effects of DSS. HSC-T6 cells stably expressing IDO1, a constitutively active IDO1 mutant, was used to determine the role of JAK2-STAT3 signaling in the DSS's anti-HF effects. ResultsWe found that intragastric administration of DSS potently reduced fibrosis, inhibited IDO1 expression and STAT3 activity both in vitro and in vivo. Using molecular docking and molecular dynamics analysis, DSS was identified as a novel IDO1 inhibitor. Mechanistic studies indicated that DSS inhibited JAK2-STAT3 signaling, it reduced IDO1 expression, STAT3 phosphorylation and STAT3 nuclear localization. More importantly, overexpression of IDO1 diminished DSS's anti-HF effects. ConclusionOur findings provide a pharmacological justification for the clinical use of DSS in treating HF, and suggest that DSS has the potential to be developed as a modern alternative and/or complimentary agent for HF treatment and prevention.

Gas Chromatography-Mass Spectrometry (GC-MS) Analysis of the Volatile Oil of Cichorium Glandulosum Boiss et Huet and its Effects on Carbon Tetrachloride-Induced Liver Fibrosis in Rats.

BackgroundThis study aimed to use gas chromatography-mass spectrometry (GC-MS) to identify the chemical constituents of volatile oil extracted by steam distillation from Cichorium glandulosum Boiss et Huet (CG), a traditional Uyghur medicine, and to investigate its effects on carbon tetrachloride (CCl4)-induced hepatic fibrosis in rats.Material/MethodsSprague-Dawley rats (n=60) included six groups: the control group (n=10), untreated model group (n=10), the volatile oil of CG high-dose group (0.15 ml/kg) (n=10), the volatile oil of CG medium-dose group (0.10 ml/kg) (N=10), the volatile oil of CG low-dose group (0.05 ml/kg) (n=10), and the silybin-treated group (0.20 ml/kg) (n=10). Rats given the essential oil extract of CG by intragastric administration, and then subcutaneously injected with a solution of CCl4 in olive oil to create the rat model of hepatic fibrosis. Serum samples were analyzed for markers of liver function, including aspartate transaminase (AST), alanine transaminase (ALT), malondialdehyde (MDA), hydroxyproline (Hyp), γ-glutamyl transpeptidase (γ-GT), lactate dehydrogenase (LDH), alkaline phosphatase (ALP), and albumin (Alb). Histology and immunohistochemistry were performed on rat liver tissue.ResultsThirty-eight compounds were identified from the volatile oil of CG (total, 98.058%), with terpenoids, including citronellol, being the most abundant. In the animal model of liver fibrosis, all doses of volatile oil of CG significantly reduced the serum levels of AST, ALT, MDA, Hyp, γ-GT, LDH, ALP, and Alb.ConclusionsGC-MS identified the components of the volatile oil of CG, which included citronellol. Treatment with volatile oil of CG reduced liver fibrosis in a rat model.

LncRNA GAS5 restrains CCl4-induced hepatic fibrosis by targeting miR-23a through the PTEN/PI3K/Akt signaling pathway.

Hepatic fibrosis is chronic liver damage with many causes that has a relatively high death rate. The current study showed that long noncoding RNA (lncRNA) growth arrest-specific transcript 5 (GAS5), microRNA-23a (miR-23a), and phosphatase and tensin homolog (PTEN) play important roles in the pathological process of hepatic fibrosis but have a relatively unclear regulatory mechanism. This study aimed to investigate the roles of lncRNA GAS5, miR-23a, and PTEN in the pathological process of hepatic fibrosis and hepatic stellate cell (HSC) activation. We used carbon tetrachloride (CCl4) intraperitoneal injections to establish a rat hepatic fibrosis model and exogenous transforming growth factor-β1 to establish an HSC activation model. Quantitative RT-PCR, Western blot, dual-luciferase reporter system, and RNA pull-down assays were used to investigate which microRNAs and lncRNAs participate in the process of hepatic fibrosis and HSC activation. miR-23a expression increased significantly in hepatic fibrosis tissues and activated HSCs. miR-23a interaction with and degradation of PTEN further influenced the downstream signaling pathway phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin/Snail (PI3K/Akt/mTOR/Snail), causing E-cadherin expression levels to decrease and α-smooth muscle actin and collagen I expression levels to increase. lncRNA GAS5 can be used as a sponge platform for miR-23a to decrease miR-23a expression levels competitively. We revealed the role of the lncRNA GAS5/miR-23a/PTEN/PI3K/Akt/mTOR/Snail signaling pathway in hepatic fibrosis, providing molecular targets for the treatment of hepatic fibrosis. NEW & NOTEWORTHY This is the first study revealing that microRNA-23a (miR-23a) promotes hepatic fibrosis through the phosphatase and tensin homolog/phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin/Snail signaling pathway, and long noncoding RNA (lncRNA) growth arrest-specific transcript 5 (GAS5) can act as a sponge platform for miR-23a. Therefore, lncRNA GAS5/miR-23a may bring molecular targets for hepatic fibrosis therapy.

Effects of Ginsenoside Rg1 on JAK2/STAT3 Signaling Pathway in Rats with Hepatic Fibrosis Induced by Concanavalin a

Background: Ginsenoside Rg1, the active ingredient of natural plants, has been proved to have anti-liver fibrosis activity. However, the specific mechanism behind these effects has not been determined. This study aims to inverstigate the specific mechanism of the antihepatic fibrosis of ginsenoside Rg1. Methods: The rat model of hepatic fibrosis induced by Concanavalin A (Con A) were established and hepatic stellate cells were isolated. Then hepatic fibrosis and cell damage were evaluated and inverstigated the mechanism. Findings: We found that the anti-hepatic fibrosis effect of ginsenoside Rg1 was related to JAK2/STAT3 signaling pathway and IL-1β, IL-6, IL-10. In addition, the expression of IL-1β, IL-6, IL-10, α-SMA, TGF-β, CollaⅠ and CollaⅡ mRNA in rats treated with different doses of ginsenoside Rg1 for 8 weeks were decreased compared with model group, and mRNA expression of IL-10 was increased, which was associated with drug dosage. Forthermore, the expression of p-JAK2, p-STAT3, Caspase3, α-SMA, TGF-β, CollaⅠ and CollaⅡ protein decreased, and the degree of decrease was correlated with the dosage of administration. Similar results were detected by Immunohistochemistry. The expression of JAK2 and STAT3 protein were not changed significantly. Besides, flow cytometry revealed that ginsenoside Rg1 could promote hepatic stellate cell (HSC) apoptosis. Interpretation: TTaken together, our results indicate that the anti-fibrotic effect of ginsenoside Rg1 is related to JAK2/STAT3 signaling pathway and IL-1β, IL-6, IL-10, and ginsenoside Rg1 is a potential drug against liver fibrosis. Funding Statement: This study was supported by the National Science Foundation of China (grant 81270498) and the National Science Foundation of China (grant 81500473). Declaration of Interests: The authors declare no conflict of interest Ethics Approval Statement: During these studies, all animal procedures were conducted in accordance with the Anhui Medical University Policy on the Care, Welfare and Treatment of Laboratory Animals and were reviewed the Institutional Animal Care and Use Committee either at Anhui Medical University or by the ethical review process at the institution where the work was performed. The experimental protocol for this study was approved by the Anhui Medical University of Animal welfare, Ethics and Strategy and Anhui Province Key Laboratory of Major Autoimmune Diseases, Hefei, China.

Alteration of UDP-glucuronosyltransferase 1a1, 1a7 and P-glycoprotein expression in hepatic fibrosis rats and the impact on pharmacokinetics of puerarin

BackgroundPuerarin, derived from a traditional Chinese herb Pueraria lobata (Willd.) Ohwi which was distributed globally and planted in most parts of China, has been extensively applied in patients with cardiovascular diseases in China. Yet a considerable proportion of the patients were accompanied with liver illnesses simultaneously because of all sorts of reasons. Hypothesis/PurposeIt had been implied by some previous research that the absorption and the metabolism of puerarin were susceptible to liver issues due to changed P-gp and Ugt1a level, but pharmacokinetics of puerarin under such conditions were few concerned. Our study aimed to make sure whether and how much the behavior of puerarin in vivo was affected by hepatic diseases, and to explore the potential mechanisms. MethodsA CCl4 induced rat model of hepatic fibrosis (HF) was prepared and verified. Single low/high doses of oral and intravenous administration of puerarin to HF and normal rats were performed. Pharmacokinetics of puerarin were determined by a validated HPLC method. The expression of P-gp, Ugt1a1, and Ugt1a7 in both liver and intestines were determined by quantitative RT-PCR and Western blot analysis respectively. ResultsThe systemic exposure of puerarin in HF rats of experimental groups were found decreased remarkably except for that of the high dose intravenous group. Moreover, the expression of P-gp, Ugt1a1, and Ugt1a7 in liver and intestines of HF rats were figured out increased. ConclusionThe results indicated that the HF originated overexpression of Ugt1a1, Ugt1a7, and P-gp level played important roles in pharmacokinetics of puerarin, suggested the clinical regimen of puerarin based on normal populations might be inappropriate for patients with chronic liver diseases. It was implied drugs whose absorption or elimination were related to P-gp, Ugt1a1, or Ugt1a7 might also be affected by hepatic illnesses.

Inhibitory effect of Scutellariae Radix on hepatic fibrosis based on urinary metabonomic

Urinary metabolomics combined with histological progression were utilized to evaluate the therapeutic effect of Scutellariae Radix decoction and baicalin on hepatic fibrosis (HF) and explore their mechanisms, intervention targets and metabolic pathways. HF rat model was established through subcutaneous injection of CCl₄ for 8 weeks. Meanwhile, different doses of Scutellariae Radix decoction and baicalin were administered. Histomorphology of liver tissue was observed and scored by HE and Masson. Urinary metabonomic analysis based on UPLC-Q-TOF-MS was made for the changes of urinary potential biomarkers among different groups at different time points of HF. Finally, it was found that Scutellariae Radix decoction could improve HF by regulating L-tryptophan, 3-methyldioxyindole, 5-hydroxyindoleacetylglycine, kynurenic acid, 4-(2-amino-3-hydroxyphenyl)-2,4-dioxobutanoic acid, methylmalonic acid and L-leucine. However, baicalin could improve HF by regulating L-tryptophan, 3-methyldioxyindole, 5-hydroxyindoleacetylglycine, 4-(2-amino-3-hydroxyphenyl)-2,4-dioxobutanoic acid, kynurenic acid, and methylmalonic acid. These metabolites involved in tryptophan metabolism and valine, leucine and isoleucine degradation pathways. These results indicated that Scutellariae Radix had the multi-target and multi-pathway characteristics in the treatment of HF. Additionally, low-dose Scutellariae Radix decoction and baicalin are showed better efficacies, with no statistically significant difference between them in histomorphology.

Effect of miR-182 on hepatic fibrosis induced by Schistosomiasis japonica by targeting FOXO1 through PI3K/AKT signaling pathway.

The study aimed to investigate the impact of miR-182 and FOXO1 on S. japonica-induced hepatic fibrosis. Microarray analysis was performed to screen out differential expressed miRNAs and mRNAs. Rat hepatic fibrosis model and human hepatocellular cell line LX-2 were used to study the effect of miR-182 and FOXO1. qRT-PCR and Western blot were used to detect the expression of miR-182, FOXO1 or other fibrosis markers. The targeting relationship between FOXO1 and miR-182 was verified by luciferase reporter assay. Immunohistochemistry or immunofluorescence staining was conducted to detect FOXO1 or α-SMA in rat hepatic tissues. Cell viability and apoptosis were detected by MTT assay and flow cytometry. The expression of PI3K/AKT pathway-related proteins was detected by Western blot. miR-182 was highly expressed in liver fibrosis samples, and FOXO1 expression was negatively correlated with miR-182 expression. After transfection of miR-182, FOXO1 expression was down-regulated, with the results of LX-2 cells proliferation inhibition and apoptosis induction, as well as the aggravation of rat hepatic fibrosis. The expression of p-AKT/AKT and p-S6/S6 was increased, meaning that the PI3K/AKT signal pathway was activated. The results were reversed when treated with Wortmannin (PI3K inhibitor). After transfection of miR-182 inhibitor, FOXO1 expression was up-regulated, LX-2 cell proliferation was inhibited, and apoptosis rate was increased. High-expressed miR-182 and low-expressed FOXO1 promoted proliferation and inhibiting apoptosis on liver fibrosis cells, stimulating the development of S. japonica-induced hepatic fibrosis through feeding back to PI3K/AKT signaling pathway.

Effect of Dahuang Zhechong pills on long non-coding RNA growth arrest specific 5 in rat models of hepatic fibrosis

OBJECTIVETo investigate the involvement of growth arrest specific 5 (GAS5), a long non-coding RNA, in the anti-hepatic fibrosis process induced by Dahuang Zhechong pill (DHZCP) in rats. METHODSThirty adult rats were divided into three groups, including a control group, a CCl4-induced fibrosis group and a DHZCP-treated fibrosis group. Hematoxylin-eosin and Masson staining were used for histopathological study. Serum enzymes, cytokines and cell proliferation were assayed using commercially available kits. A GAS5 lenti-virus vector was constructed to further investigate the role of GAS5 in the anti-hepatic fibrosis effect of DHZCP in rats. RESULTSOur results revealed that the proliferation of hepatic stellate cells cultured in serum derived from rats treated with DHZCP was significantly decreased, compared with cells treated with serum from the untreated rats. DHZCP alleviated the CCl4-induced hepatic fibrosis. Additionally, DHZCP can restore the expression of GAS5, which was significantly decreased in the CCl4-induced group, and markedly suppress the expression of p-p38 and p-Erk induced by CCl4, but not p-Jnk. Cell proliferation was significantly arrested when cells overexpressed GAS5. Thus, DHZCP can inhibit the expression of p-p38 and p-Erk, while GAS5 can only inhibit the expression of p-Erk. CONCLUSIONSDHZCP can alleviate hepatic fibrosis by increasing the expression of GAS5 to suppress p-Erk and regulating other factors to inhibit p-p38.

Balancing Effect of Biejiajian Oral Liquid (鳖甲煎口服液) on ACE-Ang II-AT1R Axis and ACE2-Ang-(1–7)-Mas Axis in Rats with CCl4-Induced Hepatic Fibrosis

To explore the effect of Biejiajian Oral Liquid (, BOL) on CCl4-induced hepatic fibrosis in rats by detecting the changes in the levels of angiotensin II (Ang II), angiotensin-(1-7) [Ang-(1-7)], angiotensin-converting enzyme (ACE), ACE2, angiotensin II type 1 receptor (AT1R), Mas, etc. METHODS: A total of 180 Wistar rats were randomly divided into two groups by random digital table method: prevention experiment and treatment experiment. Each group was further subdivided into the following 6 subgroups: normal control group, model group, vitamin E [100 mg/(kg·d), VE] group, enalapril [10 mg/(kg·g), Ena] group, high-dosage [20 g/(kg·d)] BOL group, and low-dosage [10 g/(kg·d)] BOL group. The hepatic fibrosis rat model was established by subcutaneous injection of CCl4 for 6 weeks. Prevention experiment and treatment experiment were administered with specific drugs at different times. At the end of treatment experiment, the pathological changes of liver were observed after hematoxylin-eosin staining. The expressions of ingredients in renin-angiotensin-aldosterone system (RAAS) such as AngII, Ang-(1-7), ACE, ACE2, AT1R, Mas, renin, CYP11B2 and angen in liver were detected by enzyme linked immunosorbent assay, immunohistochemistry method or reverse transcription-polymerase chain reaction, respectively. The levels of AngII and Ang-(1-7) at the 6th week increased by 496.10% and 73.64%, respectively, compared with those at the 2nd week in the model group (P<0.01). With prevention or treatment with high-dosage BOL, there was an evident reduction of AngII level but an improvement of Ang-(1-7) level. Specifically, AngII level of high-dosage group decreased by 77.50% in prevention experiment (P=0.000) and by 76.93% in treatment experiment (P=0.002) compared with that in the model group. Ang-(1-7) level increased by 91.69% in prevention experiment (P=0.006) and by 70.77% in the treatment experiment (P=0.010) compared with that in the model group. The expression levels of mRNA of renin, ACE, CYP11B2, angen and AT1R decreased by 58.15%, 99.90%, 99.84%, 99.99% and 99.99% (all P<0.01), respectively. BOL could help resist liver fibrosis in rats by enhancing the level of each ingredient in ACE2-Ang-(1-7)-Mas axis, while decreasing the level of each ingredient in ACE-AngII-AT1R axis. To some extent, BOL could enhance the regulation of RAAS in rats with CCl4-induced hepatic fibrosis.

Sorafenib and fluvastatin synergistically alleviate hepatic fibrosis via inhibiting the TGFβ1/Smad3 pathway

BackgroundEffective strategies for the treatment of hepatic fibrosis are urgently in need. AimsTo investigate the effect of the co-treatment of sorafenib and fluvastatin on hepatic fibrosis and the underlying mechanisms. MethodsA diethylnitrosamine-induced hepatic fibrosis rat model was used to evaluate the anti-fibrosis effect. Epithelial mesenchymal transition (EMT) of hepatocytes and hepatic stellate cells (HSCs) in response to sorafenib and fluvastatin was explored. A co-treatment effect on TGFβ1 expression was explored in the Kupffer cells of rats. The effect of co-treatment on the regulation of the TGFβ1/Smad3 pathway was investigated in both L02 cells and LX-2 cells. ResultsSorafenib and fluvastatin synergistically reduced collagen content, α-SMA expression, lamin level, and hyaluronic acid level in the rat hepatic model. Combination treatment significantly inhibited the expression of mesenchymal markers and promoted the expression of epithelial markers in hepatocytes. Co-treatment statistically suppressed the production of TGFβ1 in Kupffer cells. Suppression of EMT in parallel with alleviated up-regulation of fibronectin and α-SMA expression was observed in TGFβ1-activated LX-2 cells. Mechanistically, sorafenib plus fluvastatin blocked the TGFβ1/Smad3 signaling pathway via inhibiting phosphorylation of TβR II in hepatocytes and HSCs. ConclusionsSorafenib and fluvastatin synergistically alleviated diethylnitrosamine-induced hepatic fibrosis in rats. Sorafenib plus fluvastatin may be a potential combination treatment for hepatic fibrotic diseases.

Arkadia protein expression is reduced in the liver during the progression of hepatic fibrosis.

Arkadia is able to degrade key signaling molecules in the transforming growth factor (TGF)-β1 signaling pathway; however, the expression of Arkadia in the liver during development and progression of TGF-β1/Smad signaling-regulated hepatic fibrosis remains to be elucidated. The present study aimed to examine Arkadia expression in the livers of two rat models of hepatic fibrosis induced by bile duct ligation and carbon tetrachloride intoxication, and in human liver samples from patients with hepatic fibrosis. Expression was analyzed by quantitative polymerase chain reaction, immunohistochemistry and western blot analysis. The results indicated that Arkadia was predominantly expressed in the cytoplasm of cholangiocytes and hepatocytes. The protein expression levels of Arkadia were significantly decreased in fibrotic livers, whereas the mRNA expression levels of Arkadia were significantly increased in fibrotic livers compared with in nonfibrotic livers. In conclusion, these data indicated that Arkadia may regulate the pathogenesis and progression of hepatic fibrosis.

Transforming growth factor-β1 small interfering RNA regulates platelet-derived growth factor and phosphorylated extracellular regulated protein kinase in rats with hepatic fibrosis: an experimental study

Objective: To investigate the impact of transforming growth factor-β1 (TGF-β1) silencing by small interfering RNA (siRNA) on the expression of platelet-derived growth factor-β (PDGF-BB) and its receptor (PDGF-βR) in rats with hepatic fibrosis. Methods: A total of 40 male Sprague-Dawley rats were randomly divided into normal control group, model group, TGF-β1 siRNA treatment group, and negative control group. All rats except those in the normal control group were given subcutaneous injection of 40% carbon tetrachloride to establish a rat model of hepatic fibrosis. The rats in the negative control group and the TGF-β1 siRNA treatment group were given tail vein injection of negative control plasmid or TGF-β1 siRNA plasmid twice a week at a dose of 0.25 mg/kg, and those in the normal control group and the model group were given the injection of sterile isotonic saline twice a week. The rats were sacrificed after 12 weeks and liver tissue samples were collected. Real-time PCR, immunohistochemistry, and Western blot were used to measure the expression of PDGF-BB, PDGF-βR, and phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2) in liver tissue. A one-way analysis of variance, the q test, and the Kruskal-Wallis test were used for statistical analysis based on data type. Results: Compared with the model group and the negative control group, the TGF-β1 siRNA treatment group had significantly inhibited mRNA and protein expression of PDGF-BB and PDGF-βR (F = 24.785 and 22.92, P < 0.01), as well as significantly inhibited expression of p-ERK1/2 (P < 0.05). Conclusion: Targeted TGF-β1 siRNA can effectively downregulate the expression of PDGF-BB, PDGF-βR, and p-ERK1/2 in liver tissue and thus help to improve hepatic fibrosis.

A Quantitative Description of Lipid and Extracellular Matrix Proteinaceous Fibers in Hepatic Fibrosis of a Rat Model by ImageJ using Nano-Images

We made a histological and ultrastructural studies using ImageJ freeware and Origin pro statistical analysis of the site of lipid accumulations and ECM pertinacious collagen fibers in a rat model of hepatic fibrosis using CCl4 and ethanol dual effect and using nano GTE as the main factor to diminishes both hepatic lipid and ECM fibers. This imageJ observation indicates and re-enforces the notion previously published by the same author that Nano GTE is a strong therapeutic agent to work against the oxidative effects of chemicals like CCl4 and ethanol.

Carvedilol Attenuates the Progression of Hepatic Fibrosis Induced by Bile Duct Ligation.

Background. The sympathetic nervous system (SNS) is responsible for hepatic stellate cells (HSCs) activation and the accumulation of collagen that occurs in hepatic fibrogenesis. Carvedilol has been widely used for the complication of hepatic cirrhosis in the clinic. Furthermore, it has powerful antioxidant properties. We assessed the potential antifibrotic effects of carvedilol and the underlying mechanisms that may further enhance its clinical benefits. Methods. Using a bile duct ligation rat model of hepatic fibrosis, we studied the effects of carvedilol on the fibrosis, collagen deposition, and oxidative stress based on histology, immunohistochemistry, western blot, and RT-PCR analyses. Results. Carvedilol attenuated liver fibrosis, as evidenced by reduced hydroxyproline content and the accumulation of collagen, downregulated TIMP-1 and TIMP-2, and upregulated MMP-13. MMP-2 was an exception, which was decreased after carvedilol treatment for 2 weeks and upregulated after carvedilol treatment for 4 weeks. Carvedilol reduced the activation of HSCs, decreased the induction of collagen, transforming growth factor-β1, and MDA content, and strengthened the SOD activity. The antifibrotic effects were augmented as dosages increased. Conclusions. The study indicates that carvedilol attenuated hepatic fibrosis in a dose-dependent manner. It can decrease collagen accumulation and HSCs activation by the amelioration of oxidative stress.