CCl4-induced Liver Fibrosis In Rats Research Articles

Protective effects of selenium-glutathione-enriched probiotics on CCl4-induced liver fibrosis

Hepatic fibrosis is a common pathological basis of liver cirrhosis and hepatocellular carcinomas. So, prevention and treatment of liver fibrosis is one of the crucial therapeutic goals in hepatology. Organic selenium, glutathione or probiotics supplementation could ameliorate hepatic fibrosis, respectively. The purpose of this study is to develop a novel selenium-glutathione-enriched probiotics (SGP) and to investigate its protective effect on CCl4-induced liver fibrosis in rats. Yeast strains with the high-yield glutathione were isolated and identified by analysis of 26S ribosomal DNA sequences. The fermentation parameters of SGP were optimized through single-factor, Plackett–Burman (PB) design and response surface methodology (RSM). The final SGP contained 38.4 μg/g of organic selenium, 34.1 mg/g of intracellular glutathione, approximately 1×1010 CFU/g live Saccharomyces cerevisiae and 1×1012 CFU/g live Lactobacillus acidophilus. SGP had better protective effects on liver fibrosis than selenium, glutathione or probiotics, respectively. The hepatic silent information regulator 1 (SIRT1) level was down-regulated and oxidative stress, endoplasmic reticulum (ER) stress, inflammation and phosphorylated MAPK was increased in CCl4-treated rats. However, SGP can significantly reverse these changes caused by CCl4. Our findings suggest that SGP was effective in attenuating liver fibrosis by the activation of SIRT1 signaling and attenuating hepatic oxidative stress, ER stress, inflammation and MAPK signaling.

Asiatic acid attenuates CCl4-induced liver fibrosis in rats by regulating the PI3K/AKT/mTOR and Bcl-2/Bax signaling pathways

Liver fibrosis is a major pathological feature of chronic liver diseases, and effective therapies are limited at present. Asiatic acid (AA) is a triterpenoid isolated from Centella asiatica, which exhibits efficient anti-inflammatory and anti-oxidative activities. In this study, we attempted to evaluate the potential therapeutic effect of AA on CCl4-induced liver fibrosis in rats and to investigate the underlying molecular mechanisms. Liver fibrosis-related indexes including body weight, biochemical parameters, histological changes, the mRNA expression levels of inflammatory cytokines and biomarkers, and changes in the expression of related proteins in liver tissue were assessed. The results showed that AA treatment effectively ameliorated CCl4-induced liver injury and fibrosis. Mechanistically, AA treatment attenuated CCl4-induced oxidative stress, inflammation, and hepatocyte apoptosis and regulated the Bcl-2/Bax signaling pathway in the liver. Additionally, we demonstrated that AA also inhibited hepatic stellate cell activation and extra cellular matrix (ECM) synthesis by regulating the PI3K/AKT/mTOR signaling pathway. In conclusion, these findings suggest that AA prevents the progression of liver fibrosis through multiple mechanisms and indicate that AA might be used for the treatment of liver fibrosis in the future.

Transplantation of human matrix metalloproteinase-1 gene-modified bone marrow-derived mesenchymal stem cell attenuates CCL4-induced liver fibrosis in rats.

It has been reported that bone marrow-derived mesenchymal stem cells (BMSCs) alleviated liver fibrosis. We investigated whether BMSCs transfected with human matrix metalloproteinase 1 (BMSCs/MMP1) would improve their therapeutic effect in liver fibrosis induced by carbon tetrachloride (CCl4) in rats. BMSCs were transfected with an adenovirus carrying enhanced green fluorescence protein (GFP) and human MMP1 gene. BMSCs or BMSCs/MMP1 were directly injected into fibrotic rats via the tail vein. GFP-labeled cells appeared in the fibrotic liver after BMSC transplantation. The expression of BMSCs/MMP1 elevated levels of MMP1 in vitro. Although BMSC administration reduced liver fibrosis, transplantation of BMSCs/MMP1 enhanced the reduction of liver fibrosis to a higher level. Treatment with BMSCs/MMP1 not only decreased collagen content but also suppressed activation of hepatic stellate cells (HSCs) in fibrotic liver, which led to subsequent improvement of both liver injury and fibrosis. Treatment with BMSCs/MMP1 resulted in an improved therapeutic effect compared with BMSCs alone, which is probably because of the sustainably expressed MMP1 level in the liver. BMSCs/MMP1 transplantation not only improved biochemical parameters but also attenuated progression of liver fibrosis, suggesting that BMSCs may be a potential cell source in preventing liver fibrosis and MMP1 gene may enhance the anti-fibrotic effect of BMSCs.

The antifibrotic effect of Zilla spinosa extracts targeting apoptosis in CCl4-induced liver damage in rats

Background/aim Liver fibrosis and its end-stage cirrhosis are the main reasons of morbidity and mortality all over the world. The current study aimed to evaluate the efficacy of Zilla spinosa (Z. spinosa) on CCl4-induced liver fibrosis, apoptosis, and oxidative stresses in rats. Materials and methods Extract of aerial part of Z. spinosa was used in this study. Thirty male Sprague‑Dawley rats were enrolled in this study and divided into five groups (six each): group 1 served as control and groups 2–5 were treated with CCl4 (1 ml/kg intraperitoneal twice a week for 8 weeks), where group 2 served as a control positive, group 3 received silymarin (50 mg/kg) daily, and groups 4 and 5 were administrated with Z. spinosa (100 and 200 mg/kg, respectively) daily for 8 weeks. At the end of each experiment, liver function tests were analyzed in serum, whereas malondialdehyde (MDA), Nitric oxide (NO), Glutathione (GSH), and hydroxyproline (HA) were analyzed in liver tissues. Liver fibrosis was confirmed histopathologically, and collagen content, caspase-3, and α-smooth muscle actin (α-SMA) were assayed immunhistochemically. Results Alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, total bilirubin, MDA, NO, and HA levels were increased (P Conclusion The present study clarified that Z. spinosa extract has antioxidant and antiapoptotic properties in CCl4-induced liver fibrosis in rats, and may be able to exert a therapeutic effect on developing hepatic fibrosis; moreover, high dose of 200 mg/kg appeared to be more potent than low dose (100 mg/kg).

Flavanones from Sedum sarmentosum Bunge Alleviate CCl4-Induced Liver Fibrosis in Rats by Targeting TGF-β1/TβR/Smad Pathway In Turn Inhibiting Epithelial Mesenchymal Transition.

Objective The aim of the study is to evaluate the therapeutic effects of flavanones from Sedum sarmentosum Bunge (FSSB) on CCl4-induced liver fibrosis in rats and the underlying mechanisms of action. Methods An experimental model of liver fibrosis was established by subcutaneous injection of rats with CCl4 (40% v/v, 3 ml/kg) twice per week for six weeks. FSSB (100, 200, and 400 mg/kg) was intragastrically administered once per day consecutively for five weeks. Results Our results showed that FSSB significantly attenuated CCl4-induced liver fibrosis as evidenced by reducing the elevated levels of serum biochemical indexes and improving the histological changes, including decreasing the elevation in serum alanine transaminase (ALT), aspartate transaminase (AST), hyaluronic acid (HA), and laminin (LN) level, reducing infiltration of inflammatory cells and collagen fibers in liver tissue. In addition, compared to the model group, FSSB markedly downregulated the protein and mRNA expression of TGF-β1, TGF-β1 receptors I and II (TβRI and TβRII), Smad2, Smad3, and Vimentin in liver tissue, at the mean time upregulating the expression of Smad7 and E-cadherin. Conclusions The results suggest that FSSB alleviated CCl4-induced liver fibrosis probably through inhibition of TGF-β/TβR/Smad pathway in turn inhibiting epithelial mesenchymal transition.

In vivo hepatic differentiation potential of human umbilical cord-derived mesenchymal stem cells: Therapeutic effect on liver fibrosis/cirrhosis.

AIMTo investigate the hepatic differentiation potential of human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) and to evaluate their therapeutic effect on liver fibrosis/cirrhosis.METHODSA CCl4-induced liver fibrotic/cirrhotic rat model was used to assess the effect of hUC-MSCs. Histopathology was assessed by hematoxylin and eosin (H&E), Masson trichrome and Sirius red staining. The liver biochemical profile was measured using a Beckman Coulter analyzer. Expression analysis was performed using immunofluorescent staining, immunohistochemistry, Western blot, and real-time PCR.RESULTSWe demonstrated that the infused hUC-MSCs could differentiate into hepatocytes in vivo. Functionally, the transplantation of hUC-MSCs to CCl4-treated rats improved liver transaminases and synthetic function, reduced liver histopathology and reversed hepatobiliary fibrosis. The reversal of hepatobiliary fibrosis was likely due to the reduced activation state of hepatic stellate cells, decreased collagen deposition, and enhanced extracellular matrix remodeling via the up-regulation of MMP-13 and down-regulation of TIMP-1.CONCLUSIONTransplanted hUC-MSCs could differentiate into functional hepatocytes that improved both the biochemical and histopathologic changes in a CCl4-induced rat liver fibrosis model. hUC-MSCs may offer therapeutic opportunities for treating hepatobiliary diseases, including cirrhosis.

Lycium barbarum polysaccharides improve CCl4-induced liver fibrosis, inflammatory response and TLRs/NF-kB signaling pathway expression in wistar rats

Lycium barbarum polysaccharides (LBPs) have multiple biological and pharmacological functions, including antioxidant, anti-inflammatory and anticancer activities. This research was conducted to evaluate whether LBPs could alleviate carbon tetrachloride (CCl4)-induced liver fibrosis and the underlying signaling pathway mechanism. Fifty male wistar rats were randomly allocated to five groups (n=10): control, CCl4 and CCl4 with 400, 800 or 1600mg/kg LBPs, respectively. Each wistar rat from each group was used for blood and tissue collections at the end of experiment. The results showed that CCl4 induced liver fibrosis as demonstrated by increasing histopathological damage, α-smooth muscle actin expression, aspartate transaminase activities, alkaline phosphatase activities and alanine aminotransferase activities. LBPs supplementation alleviated CCl4-induced liver fibrosis as demonstrated by reversing the above parameters. In addition, CCl4 treatment induced the oxidative injury, increased the mRNA levels of tumor necrosis factor-α, monocyte chemoattractant protein-1 and interleukin-1β, and up-regulated the protein expressions of toll-like receptor 4 (TLR4), TLR2, myeloid differentiation factor 88, nuclear factor-kappa B (NF-kB) and p-p65. LBPs supplementation alleviated CCl4-induced oxidative injury, inflammatory response and TLRs/NF-kB signaling pathway expression by reversing the above some parameters. These results suggest that the alleviating effects of LBPs on CCl4-induced liver fibrosis in wistar rats may be through inhibiting the TLRs/NF-kB signaling pathway expression.

Protective effect of ethyl acetate fraction of Drynaria quercifolia against CCl4 induced rat liver fibrosis via Nrf2/ARE and NFκB signalling pathway

Ethnopharmacological relevanceDrynaria quercifolia rhizome is traditionally used as hepatoprotective drug especially in chronic jaundice. Aim of the studyThe present study was undertaken to scientifically evaluate the efficacy of D. quercifolia rhizome against liver fibrosis. Materials and methodsD. quercifolia rhizome crude extract (DQ) and its fractions of hexane (HDQ), ethyl acetate (EDQ), butanol (BDQ) were evaluated in vitro using primary hepatocytes and RAW 264.7 cells. In vivo anti-liver fibrotic activity of EDQ was assessed using CCl4 induced liver fibrosis in Wistar rats and serum biochemical parameters (AST, ALT, ALP, SB, cholesterol), MDA, PT, INR, GSH, SOD, CAT, liver glycogen, serum albumin levels were monitored. qRT-PCR analysis of TNF-α, COX-2, iNOS were performed. ELISA method was used to estimate TNF-α, COX-1 & 2. Histopathological studies like H & E, Masson’s trichrome, immunohistochemistry staining for α-SMA, TIMP-1, Nrf2 were conducted. LC-Q-TOF-MS analysis of EDQ was conducted. ResultsIn vitro activity guided fractionation of D. quercifolia revealed EDQ as active fraction when compared to other extracts. EDQ treatment significantly inhibited the expression of α-SMA, TIMP-1, COX-2, TNF-α, iNOS and increased the levels of Nrf2 in rat liver fibrosis. LC-Q-TOF-MS analysis of EDQ confirmed the presence of naringin and naringenin. ConclusionThe anti-liver fibrotic activity of EDQ is via inhibition of NFκB signalling pathway, antioxidant response through Nrf2 activation and further inhibition of HSC activation.

Chinese herbal formula Fuzheng Huayu alleviates CCl4-induced liver fibrosis in rats: a transcriptomic and proteomic analysis.

Liver fibrosis is a consequence of chronic liver disease that can progress to liver cirrhosis or even hepatocarcinoma. Fuzheng Huayu (FZHY), a Chinese herbal formula, has been shown to exert anti-fibrotic effects. To better understand the molecular mechanisms underlying the anti-fibrotic effects of FZHY, we analyzed transcriptomic and proteomic combination profiles in CCl4-induced liver fibrosis in rats, which were treated with extracted FZHY powder (0.35 g·kg-1·d-1, ig) for 3 weeks. We showed that FZHY administration significantly improved liver function, alleviated hepatic inflammatory and fibrotic changes, and decreased the hydroxyproline content in the livers of CCl4-treated rats. When their liver tissues were examined using microarray and iTRAQ, we found 255 differentially expressed genes (fold change ≥1.5, P<0.05) and 499 differentially expressed proteins (fold change ≥1.2, P<0.05) in the FZHY and model groups. Functional annotation with DAVID (The Database for Annotation, Visualization and Integrated Discovery) showed that 15 enriched gene ontology terms, including drug metabolic process, response to extracellular stimulus, response to vitamins, arachidonic acid metabolic process, response to wounding, and oxidation reduction might be involved in the anti-fibrotic effects of FZHY; whereas KEGG pathway analysis revealed that eight enriched pathways, including arachidonic acid metabolism, retinol metabolism, metabolism of xenobiotics by cytochrome P450, and drug metabolism might also be involved. Moreover, the protein-protein interaction network demonstrated that 10 core genes/proteins overlapped, with Ugt2a3, Cyp2b1 and Cyp3a18 in retinol metabolism pathway overlapped to a higher degree. Compared to the model rats, the livers of FZHY-treated rats had significantly higher mRNA and protein expression levels of Ugt2a3, Cyp2b1 and Cyp3a18. Furthermore, the concentration of retinoic acid was significantly higher in the FZHY-treated rats compared with the model rats. The results suggest that the anti-fibrotic effects of FZHY emerge through multiple targets, multiple functions, and multiple pathways, including FZHY-regulated retinol metabolism, xenobiotic metabolism by cytochrome P450, and drug metabolism through up-regulated Ugt2a3, Cyp2b1, and Cyp3a18. These genes may play important anti-fibrotic roles in FZHY-treated rats.

Systemic inhibition of BMP1-3 decreases progression of CCl4-induced liver fibrosis in rats

Liver fibrosis is a progressive pathological process resulting in an accumulation of excess extracellular matrix proteins. We discovered that bone morphogenetic protein 1-3 (BMP1-3), an isoform of the metalloproteinase Bmp1 gene, circulates in the plasma of healthy volunteers and its neutralization decreases the progression of chronic kidney disease in 5/6 nephrectomized rats. Here, we investigated the potential role of BMP1-3 in a chronic liver disease. Rats with carbon tetrachloride (CCl4)-induced liver fibrosis were treated with monoclonal anti-BMP1-3 antibodies. Treatment with anti-BMP1-3 antibodies dose-dependently lowered the amount of collagen type I, downregulated the expression of Tgfb1, Itgb6, Col1a1, and Acta2 and upregulated the expression of Ctgf, Itgb1, and Dcn. Mehanistically, BMP1-3 inhibition decreased the plasma levels of transforming growth factor beta 1(TGFβ1) by prevention of its activation and lowered the prodecorin production further suppressing the TGFβ1 profibrotic effect. Our results suggest that BMP1-3 inhibitors have significant potential for decreasing the progression of fibrosis in liver cirrhosis.

CAFFEINEAS A PROMISING ANTIFBROTIC AGENT AGAINST CCL4-INDUCED LIVER FIBROSIS

Objective: Hepatic fibrosis is a wound-healing process in the liver with chronic injury and is characterized by an excess production and deposition of extracellular matrix (ECM) components. Our aim is to investigate the hepatoprotective effects of caffeine against CCL4-induced liver fibrosis in rats and to be compared with the reference standard N-acetylcysteine (NAC).Methods: Rats were divided into 5 groups (n=8), the 1st group served as normal control, the 2nd group received corn oil, the 3rd group is a fibrosis control and the remaining two groups received in addition to CCL4, NAC (150 mg/kg/day) as a reference treatment and caffeine (15 mg/kg/day). At the end of experimental period the following parameters were measured, specific fibrosis biomarkers [hepatic transforming growth factor β1(TGF-β1) and hepatic hydroxyproline (HYP)], liver function biomarkers [serum alanine transaminase (ALT), aspartate transaminase (AST), total bilirubin and albumin], lipid profile [serum triglycerides, total cholesterol, low density lipoprotein cholesterol (LDL-Ch) and high density lipoprotein cholesterol (HDL-Ch)], inflammatory biomarkers [serum tumor necrosis factor alpha (TNF-α), hepatic myeloperoxidase (MPO)], oxidative stress biomarkers [hepatic malondialdehyde (MDA), glutathione (GSH) and catalase (CAT)], relative liver body weight and liver histopathological study.Results: Caffeine significantly improved all the aforementioned biochemical parameters and liver sections obtained from this group showed prominent histopathological improvement.Conclusion: Caffeine hepatoprotective effects may be due to modulation of the fibrous scar formation, Improvement of liver function, Anti-inflammatory and antioxidant potentials.

Effect of ginseng extract on the TGF-\u03b21 signaling pathway in CCl4-induced liver fibrosis in rats

BackgroundLiver diseases are major global health problems. Ginseng extract has antioxidant, immune-modulatory and anti-inflammatory activities. This study investigated the effect of ginseng extract on carbon tetrachloride (CCl4)-induced liver fibrosis in rats.MethodsMale Wistar rats were divided into four groups: control group, ginseng group, CCl4 group and CCl4 + ginseng group. Liver injury was induced by the intraperitoneal (I.P) injection of 3 ml/kg CCl4 (30% in olive oil) weekly for 8 weeks. The control group was I.P injected with olive oil. The expression of genes encoding transforming growth factor beta (TGF-β), type I TGF-β receptor (TβR-1), type II TGF-β receptor (TβR-II), mothers against decapentaplegic homolog 2 (Smad2), Smad3, Smad4, matrix metalloproteinase 2 (MMP2), MMP9, tissue inhibitor matrix metalloproteinase-1 (TIMP-1), Collagen 1a2 (Col1a2), Collagen 3a1 (Col3a1), interleukin-8 (IL-8) and interleukin -10 (IL-10) were measured by real-time PCR.ResultsTreatment with ginseng extract decreased hepatic fat deposition and lowered hepatic reticular fiber accumulation compared with the CCl4 group. The CCl4 group showed a significant increase in hepatotoxicity biomarkers and up-regulation of the expression of genes encoding TGF-β, TβR-I, TβR-II, MMP2, MMP9, Smad-2,-3, -4, and IL-8 compared with the control group. However, CCl4 administration resulted in the significant down-regulation of IL-10 mRNA expression compared with the control group. Interestingly, ginseng extract supplementation completely reversed the biochemical markers of hepatotoxicity and the gene expression alterations induced by CCl4.Conclusionginseng extract had an anti‐fibrosis effect via the regulation of the TGF‐β1/Smad signaling pathway in the CCl4‐induced liver fibrosis model. The major target was the inhibition of the expression of TGF‐β1, Smad2, and Smad3.

Dimethyl α-ketoglutarate reduces CCl4-induced liver fibrosis through inhibition of autophagy in hepatic stellate cells

Sustained activation of hepatic stellate cells (HSCs) leads to liver fibrosis. Autophagy fuels the activation of HSCs by generation of ATP. Our previous research demonstrated an inhibitory effect of dimethyl α-ketoglutarate (DMKG) on HSCs activation in vitro. In the current study, we demonstrated that DMKG reduced CCl4-induced liver fibrosis in Wistar rats. Then, with the use of the HSC-T6 cell lines and double immunofluorescent staining of liver sections, we showed that the anti-fibrotic effect occurred through the inhibition of the autophagy of HSCs. Both experiments showed that DMKG could inhibit autophagy and activation of HSCs, and that the activation of HSCs was down-regulated with autophagy. In addition, we showed that DMKG could lead to lipid droplet accumulation and decrease cellular ATP content in HSCs. Furthermore, the mechanism of how DMKG inhibited autophagy of HSCs was explored in vitro with the use of c646 (a competitive inhibitor of acetyl-coenzyme A which binds to the acetyltransferase EP300) and lipoic acid (an alternative acetyl-coenzyme A -replenishing agent to DMKG), and showed that both acetyl-coenzyme A and EP300 were involved.Collectively, our study investigated the possible role of DMKG in preventing liver fibrosis and HSCs activation. We showed that DMKG may be a potential therapeutic agent for the treatment of liver fibrosis.

Activation of the miR-34a/SIRT1/p53 Signaling Pathway Contributes to the Progress of Liver Fibrosis via Inducing Apoptosis in Hepatocytes but Not in HSCs.

Liver fibrosis results from a sustained wound healing response to chronic liver injury, and the activation of nonparenchymal hepatic stellate cells (HSCs) is the pivotal process. MicroRNA-34a (miR-34a) is the direct target gene of p53 and activates p53 through sirtuin 1 (SIRT1) simultaneously. The miR-34a/SIRT1/p53 signaling pathway thus forms a positive feedback loop wherein p53 induces miR-34a and miR-34a activates p53 by inhibiting SIRT1, playing an important role in cell proliferation and apoptosis. miR-34a expression has been found to be increased in animal models or in human patients with different liver diseases, including liver fibrosis. However, the exact role of this classical miR-34a/SIRT1/p53 signaling pathway in liver fibrosis remains unclear. In the present study, using a CCl4-induced rat liver fibrosis model, we found that the miR-34a/SIRT1/p53 signaling pathway was activated and could be inhibited by SIRT1 activator SRT1720. Further studies showed that the miR-34a/SIRT1/p53 signaling pathway was activated in hepatocytes but not in HSCs. The activation of this pathway in hepatocytes resulted in the apoptosis of hepatocytes and thus activated HSCs. Our data indicate that the miR-34a/SIRT1/p53 signaling pathway might be a promising therapeutic target for liver fibrosis.

Metabolic Profile Changes of CCl₄-Liver Fibrosis and Inhibitory Effects of Jiaqi Ganxian Granule.

Jiaqi Ganxian Granule (JGG) is a famous traditional Chinese medicine, which has been long used in clinical practice for treating liver fibrosis. However, the mechanism underlying its anti-hepatic fibrosis is still not clear. In this study, an Ultra-Performance Liquid Chromatography-Time-Of-Flight Mass Spectrometry (UPLC-TOF-MS)-based metabolomics strategy was used to profile the metabolic characteristic of serum obtained from a carbon tetrachloride (CCl4)-induced hepatic fibrosis model in Sprague-Dawley (SD) rats with JGG treatment. Through Principal Component Analysis (PCA) and Partial Least Square Discriminant Analysis (PLS-DA), it was shown that metabolic perturbations induced by CCl4 were inhibited after treatment of JGG, for 17 different metabolites related to CCl4. Among these compounds, the change tendency of eight potential drug targets was restored after the intervention with JGG. The current study indicates that JGG has a significant anti-fibrosis effect on CCl4-induced liver fibrosis in rats, which might be by regulating the dysfunction of sphingolipid metabolism, glycerophospholipid metabolism, N-acylethanolamine biosynthesis, fat digestion and absorption, while glycerophospholipid metabolism played vital roles in the inhibitory effects of JGG on hepatic fibrosis according to Metabolic Pathway Analysis (MetPA). Our findings indicated that the metabolomics approach may provide a useful tool for exploring potential biomarkers involved in hepatic fibrosis and elucidate the mechanisms underlying the action of therapies used in traditional Chinese medicine.

Protective effect and mechanism of action of mulberry marc anthocyanins on carbon tetrachloride-induced liver fibrosis in rats

Mulberry fruit is widely consumed either fresh or in processed food products such as juice, mulberry jelly and jam. Conversely, mulberry marc, the solid component after juicing, is primarily a waste product or used in animal fodder, despite being rich in anthocyanins. Anthocyanins have been reported to have hepatoprotective, hypolipidaemic and antioxidant activities. In this study, the protective effect of mulberry marc anthocyanins (MMA) on carbon tetrachloride (CCl4)-induced liver fibrosis in rats was investigated. Male Sprague Dawley rats were injected with CCl4 to induce liver fibrosis. Compared with the model group, the contents of alanine aminotransferase, aspartate amino transferase, hyaluronidase acid, hydroxyproline and collagen type-III in the MMA 200 mg/kg group were decreased. Moreover, pathological examination and immunohistochemical showed that CCl4-induced liver fibrosis was alleviated by administration of MMA. These results demonstrate that MMA has a potent protective effect on CCl4-induced liver fibrosis in rats.

Gingerol protects against experimental liver fibrosis in rats via suppression of pro-inflammatory and profibrogenic mediators.

6-Gingerol (Gin) is known to possess hepatoprotective effects. Liver fibrosis is a major health concern that results in significant morbidity and mortality. There is no FDA-approved medication for liver fibrosis. The present work aimed at exploring the beneficial effects of Gin against liver fibrosis in rats. Experimental fibrosis was induced by challenging animals with CCl4 for 6weeks. Gin significantly ameliorated the increase in serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, albumin, total cholesterol (TC) and triglyceride (TG) concentrations, and liver index. These effects were confirmed by light and electron microscopic examinations. The antifibrotic effects were confirmed by examining Masson trichrome-stained liver sections which indicated reduced collagen deposition in Gin-treated animals. Further, Gin administration hampered alpha-smooth muscle actin (α-SMA) expression and significantly reduced hepatic content of transforming growth factor-beta (TGF-β). Also, Gin elicited profound antioxidant actions as indicated by preventing reduced glutathione (GSH) depletion and lipid peroxide accumulation. The observed antifibrotic activities involved decreased production of nuclear factor κB (NF-κB), tumor necrosis factor alpha (TNF-α), expression of toll-like receptor 4 (TLR4), intercellular adhesion molecule (ICAM), and vascular cell adhesion molecule (VCAM). Involvement of Gin anti-inflammatory activity was verified by the decreased expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in livers of animals treated with Gin. Thus, it can be concluded that Gin protects against CCl4-induced liver fibrosis in rats. This can be ascribed, at least partly, to its antioxidant, anti-inflammatory effects as well as the inhibition of NF-κB/TLR-4 expression.

Hepatocyte Growth Factor Mediates the Antifibrogenic Action of Ocimum bacilicum Essential Oil against CCl4-Induced Liver Fibrosis in Rats.

The current investigation aimed to evaluate the antifibrogenic potential of Ocimum basilicum essential oil (OBE) and further to explore some of its underlying mechanisms. Three groups of rats were used: group I (control), group II (CCl4 model) and group III (OBE-treated) received CCl4 and OBE 2 weeks after the start of CCl4 administration. Oxidative damage was assessed by the measurement of MDA, NO, SOD, CAT, GSH and total antioxidant capacity (TAC). Liver fibrosis was assessed histopathologically by Masson’s trichrome staining and α-smooth muscle actin (α-SMA) immunostaining. Expression of hepatocyte growth factor (HGF) and cytochrome P450 (CYP2EI isoform) was estimated using real-time PCR and immunohistochemistry. OBE successfully attenuated liver injury, as shown by histopathology, decreased serum transaminases and improved oxidative status of the liver. Reduced collagen deposition and α-SMA immuopositive cells indicated an abrogation of hepatic stellate cell activation by OBE. Furthermore, OBE was highly effective in stimulating HGF mRNA and protein expression and inhibiting CCl4-induced CYP2E1 down-regulation. The mechanism of antifibrogenic action of OBE is hypothesized to proceed via scavenging free radicals and activating liver regeneration by induction of HGF. These data suggest the use of OBE as a complementary treatment in liver fibrosis.

MicroRNA-17-5p activates hepatic stellate cells through targeting of Smad7

A considerable amount of research has focused on the roles of microRNAs (miRNA) in the pathophysiology of liver fibrosis in view of their regulatory effects on hepatic stellate cell (HSC) functions, including proliferation, differentiation, and apoptosis. Recently, miR-17-5p was shown to promote cell proliferation and migration in liver. Transforming growth factor-β1 (TGF-β1) has been characterized as the master fibrogenic cytokine that stimulates HSC activation and promotes progression of liver fibrosis. The issue of whether miR-17-5p plays a role in TGF-β1-induced hepatic fibrogenesis remains to be established. In this study, we demonstrated a dose-/time-dependent increase in miR-17-5p expression in TGF-β1-treated HSCs. Enhanced miR-17-5p expression was additionally observed in CCl4-induced rat liver fibrosis. Inhibition of miR-17-5p led to suppression of HSC proliferation induced by TGF-β1 without affecting cellular apoptosis. Notably, miR-17-5p was significantly associated with TGF-β1-induced expression of type I collagen and α-SMA in HSC. Furthermore, Smad7, a negative regulator of the TGF-β/Smad pathway, was confirmed as a direct target of miR-17-5p. Serum miR-17-5p levels were significantly higher in patients with cirrhosis, compared to healthy controls. Our results collectively indicate that miR-17-5p promotes HSC proliferation and activation, at least in part, via reduction of Smad7, supporting its potential utility as a novel therapeutic target for liver fibrosis.

MicroRNA-370 Attenuates Hepatic Fibrogenesis by Targeting Smoothened.

Recent research shows that abnormal expression of microRNA plays an important role in the process of hepatic fibrosis . miR-370 has been reported to be involved in liver function and is suppressed during hepatic carcinogenesis. The aim of this study was to investigate the role of miR-370 in hepatic fibrosis. The expression levels of miR-370 in rat fibrotic livers and activated hepatic stellate cells (HSCs) were evaluated by quantitative real-time PCR. The effect of miR-370 on the activation of HSCs was analyzed by flow cytometric analyses, real-time PCR and Western blot. Adenovirus carrying miR-370 was injected through the tail vein to access the effect of miR-370 on hepatic fibrosis induced by CCl4 in rats. The downstream targets of miR-370 were predicted by the Target Scan database and verified by luciferase assays, real-time PCR and Western blot in HSCs and were further confirmed by immunohistochemistry in vivo. Real-time PCR showed that miR-370 expression was significantly reduced in rat fibrotic livers and TGFβ1-stimulated HSCs. Overexpression of miR-370 inhibited the proliferation of HSC-T6 cells via inducing cell apoptosis and suppressed the activation of HSCs. Upregulation of miR-370 obviously attenuated the CCl4-induced liver fibrosis in rats. miR-370 was directly bound to the 3'UTR of Smoothened (SMO) and suppressed the expression of SMO in HSCs and fibrotic livers. Our study demonstrated that miR-370 plays an inhibitory role in hepatic fibrogenesis by targeting SMO. Restoration of miR-370 may have beneficial effects on the treatment of liver fibrosis.