CCl4-induced Acute Liver Injury Research Articles

Screening for the protective effect target of deproteinized extract of calf blood and its mechanisms in mice with CCl4-induced acute liver injury.

Liver injury is a common pathological basis of various liver diseases, and long-term liver injury is often an important initiation factor leading to liver fibrosis and even liver cirrhosis and hepatocellular carcinoma (HCC). It has been reported that deproteinized extract of calf blood (DECB) can inhibit the replication of hepatitis B virus and confers a protective effect on the liver after traumatic liver injury. However, few studies on the regulatory factors and mechanisms of DECB have been reported. In this current study, an acute mouse liver injury model was established with carbon tetrachloride (CCl4). The differentially expressed genes and related cell signal transduction pathways were screened using mRNA expression microarray. STEM software V1.3.6 was used for clustering gene functions, and the DAVID and KEGG databases were applied for the analysis. A total of 1355 differentially expressed genes were selected, among which nine were validated by RT-qPCR. The results showed that the Fas, IL1b, Pik3r1, Pik3r5, Traf2, Traf2, Csf2rb2, Map3k14, Pik3cd and Ppp3cc genes were involved in the regulation of DECB in an acute mouse liver injury model. Targets of the protective effects of DECB and its related mechanisms were found in mice with acute liver injury induced by carbon tetrachloride, which may provide an important theoretical basis for further DECB research.

Protective effect of human serum amyloid P on CCl4-induced acute liver injury in mice.

Human serum amyloid P (hSAP), a member of the pentraxin family, inhibits the activation of fibrocytes in culture and inhibits experimental renal, lung, skin and cardiac fibrosis. As hepatic inflammation is one of the causes of liver fibrosis, in the present study, we investigated the hepatoprotective effects of hSAP against carbon tetrachloride (CCl4)-induced liver injury. Our data indicated that hSAP attenuated hepatic histopathological abnormalities and significantly decreased inflammatory cell infiltration and pro-inflammatory factor expression. Moreover, CCl4-induced apoptosis in the mouse liver was inhibited by hSAP, as measured by terminal-deoxynucleotidyl transferase mediated nick-end labeling (TUNEL) assay and cleaved caspase-3 expression. hSAP significantly restored the expression of B cell lymphoma/leukemia (Bcl)-2 and suppressed the expression of Bcl-2-associated X protein (Bax) in vivo. The number of hepatocytes in early apoptosis stained with Annexin V was significantly reduced by 28–30% in the hSAP treatment group compared with the CCl4 group, and the expression of Bcl-2 was increased, whereas the expression of Bax and cleaved caspase-3 were significantly inhibited in the hSAP pre-treatment group compared with the CCl4 group. hSAP administration also inhibited the migration and activation of hepatic stellate cells (HSCs) in CCl4-injured liver and suppressed the activation of isolated primary HSCs induced by transforming growth factor (TGF)-β1 in vitro. Collectively, these findings suggest that hSAP exerts a protective effect againts CCl4-induced hepatic injury by suppressing the inflammatory response and hepatocyte apoptosis, potentially by inhibiting HSC activation.

Methane alleviates carbon tetrachloride induced liver injury in mice: anti-inflammatory action demonstrated by increased PI3K/Akt/GSK-3β-mediated IL-10 expression.

The inflammatory response plays an important role in carbon tetrachloride (CCl4)-induced acute liver injury and methane has been shown to exert beneficial effects on inflammation-associated diseases. Thus, we investigated the potential protective effects of methane-rich saline (MS) on CCl4-induced acute liver injury and explored the underlying mechanism. A CCl4-induced acute liver injury model was established by injection of CCl4 (0.6ml/kg, ip) in mice followed by treatment with MS (16ml/kg, ip), 24h later. All groups of mice were sacrificed and blood and liver tissues were collected. Serum aminotransferase, necrotic areas, and inflammatory cell infiltration in liver slices were enhanced after CCl4 treatment but decreased with MS treatment. IL-6, TNF-α, IL-1β, IFN-γ, ICAM-1, CXCL1, MPO, NF-κB p65, ERK, JNK, and MAPK P38, expression in serum or liver homogenate were greater after CCl4 treatment but comparatively less after MS treatment. Only IL-10 increased after MS treatment. Anti-IL10 blockade (1.5mg/kg) restored MS-mediated attenuated phosphorylation of NF-ĸbB/MAPK and the protective effect of MS was abolished for all indices examined. The PI3K inhibitor, wortmannin had the same effects on MS as anti-IL-10 antibody. MS also induced phosphorylation of GSK-3β and AKT in CCl4-treated mice. After pre-treatment with wortmannin (0.7mg/kg), phosphorylation of GSK-3β and AKT proteins were reduced compared to its solvent control group-DMSO-treated animals. Thus, the data provide evidence that MS may activate the PI3K-AKT-GSK-3β pathway to induce IL-10 expression and produce anti-inflammatory effects via the NF-κB and MAPK pathways. The findings provide a new pharmacological strategy for management of inflammatory response after acute liver injury.

Hepatoprotective effect of the ethanol extract of Polygonum orientale on carbon tetrachloride-induced acute liver injury in mice

Polygonum orientale L. (Polygonaceae) fruits have various medicinal uses, but their hepatoprotective effects have not yet been studied. This study investigated the hepatoprotective activity of the ethanolic extract of P. orientale (POE) fruits against carbon tetrachloride (CCl4)-induced acute liver injury (ALI). Mice were pretreated with POE (0.1, 0.5, and 1.0 g/kg) or silymarin (0.2 g/kg) for 5 consecutive days and administered a dose of 0.175% CCl4 (ip) on the 5th day to induce ALI. Blood and liver samples were collected to measure antioxidative activity and cytokines. The bioactive components of POE were identified through high-performance liquid chromatography (HPLC). Acute toxicity testing indicated that the LD50 of POE exceeded 10 g/kg in mice. Mice pretreated with POE (0.5, 1.0 g/kg) experienced a significant reduction in their serum aspartate aminotransferase (AST), serum alanine aminotransferase (ALT), and alkaline phosphatase (ALP) levels and reduction in the extent of liver lesions. POE reduced the malondialdehyde (MDA), nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) levels, and increased the activity of superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione reductase (GRd) in liver. HPLC revealed peaks at 11.28, 19.55, and 39.40 min for protocatechuic acid, taxifolin, and quercetin, respectively. In summary, the hepatoprotective effect of POE against CCl4-induced ALI was seemingly associated with its antioxidant and anti-proinflammatory activities.

Editor's Highlight: Farnesoid X Receptor Protects Against Low-Dose Carbon Tetrachloride-Induced Liver Injury Through the Taurocholate-JNK Pathway.

Hepatotoxicity is of major concern for humans exposed to industrial chemicals and drugs. Disruption of farnesoid X receptor (FXR), a master regulator of bile acid (BA) metabolism, enhanced the sensitivity to liver injury in mice after toxicant exposure, but the precise mechanism remains unclear. In this study, the interconnection between BA metabolism, FXR, and chemically induced hepatotoxicity was investigated using metabolomics, Fxr-null mice (Fxr-/-) and hepatocytes, and recombinant adenoviruses. A single low-dose intraperitoneal injection of carbon tetrachloride (CCl4), an inducer of acute hepatitis in mice, resulted in more severe hepatocyte damage and higher induction of pro-inflammatory mediators, such as chemokine (C-C motif) ligand 2 (Ccl2), in Fxr-/-. Serum metabolomics analysis revealed marked increases in circulating taurocholate (TCA) and tauro-β-muricholate (T-β-MCA) in these mice, and forced expression of bile salt export protein (BSEP) by recombinant adenovirus in Fxr-/- ameliorated CCl4-induced liver damage. Treatment of Fxr-null hepatocytes with TCA, but not T-β-MCA, significantly increased c-Jun-N-terminal kinase (JNK) activation and Ccl2 mRNA levels, and up-regulation of Ccl2 mRNA was attenuated by co-treatment with a JNK inhibitor SP600125, indicating that TCA directly amplifies hepatocyte inflammatory signaling mainly mediated by JNK under FXR-deficiency. Additionally, pretreatment with SP600125 or restoration of FXR expression in liver by use of recombinant adenovirus, attenuated CCl4-induced liver injury. Collectively, these results suggest that the TCA-JNK axis is likely associated with increased susceptibility to CCl4-induced acute liver injury in Fxr-/-, and provide clues to the mechanism by which FXR and its downstream gene targets, such as BSEP, protects against chemically induced hepatotoxicity.

Protective effects of five different types of Dendrobium on CCl4-induced liver injury in mice

This study aims to investigate the protective effect of Dendrobium huoshanense, D.officinale(Huoshan), D.officinale(Yunnan), D.moniliforme and D. henanense on CCl4-induced hepatic damage in mice. C57BL/6 mice were randomly divided into control group, model group, high-dose(7.5 g•kg⁻¹) and low-dose (1.25 g•kg⁻¹) groups of the five Dendrobium. Each group was intragastrically administered with drugs for 2 weeks. The control group was intraperitoneally injected with Olive oil solution, while the other groups were intraperitoneally given 0.5%CCl4combined with Olive oil solution 2 h later after the last administration. Subsequently, ALT and AST activities in serum, SOD activities and MDA contents in liver tissues were determined in all groups 16 h later after administration. The liver index was calculated, and hepatic histopathological examination was performed. The mRNA expressions of IL-1β, IL-6 and TNF-α were analyzed by Real-time PCR. Compared with the CCl4 model group, the activities of ALT and AST in serum decreased significantly in the five different Dendrobium groups. Meanwhile, in liver tissues, the levels of MDA reduced obviously, while the SOD activities markedly increased. Furthermore, liver tissue damage induced by CCl4 was ameliorated according to the histopathological examination. IL-1β, IL-6 and TNF-α mRNA expressions in D.huoshanense-treated liver tissues were significantly decreased. In conclusion, the five different Dendrobium groups showed hepatoprotective effects on CCl4-induced acute liver injury in mice. However, there were differences among Dendrobium of different types and origins. The protect effect of D.huoshanense is the most obvious, and the order of the protective effect of the other Dendrobium from high to low is D.officinale(Yunnan), D. officinale(Huoshan), D.henanense and D.moniliforme. The differences between the different types of Dendrobium might be related to their chemical components.

Hesperetin derivative-14 alleviates inflammation by activating PPAR-γ in mice with CCl4-induced acute liver injury and LPS-treated RAW264.7 cells

Hesperetin is a flavanone glycoside compound naturally occurring in the fruit peel of Citrusaurantium L. (Rutaceae). Previous studies revealed that hesperetin possesses various pharmacological effects, including anti-inflammation, anti-tumor, anti-oxidant and neuroprotective properties. Hesperetin derivative-14 (HD-14) is a derivative of hesperetin improved in water solubility and bioavailability. In this study, we indicated that HD-14 (2μM) significantly attenuated inflammation in LPS-treated RAW264.7 cells, besides, HD-14 (100mg/kg) exhibited hepato-protective effects and anti-inflammatory effects on C57BL/6J mice with CCl4-induced acute liver injury. In addition, it was demonstrated that HD-14 dramatically up-regulated the expression of PPAR-γ in vivo and in vitro. Interestingly, over-expression of PPAR-γ had anti-inflammatory effects on the expressions of TNF-α, IL-6, and IL-1β, whereas, knockdown of PPAR-γ with small interfering RNA had pro-inflammatory effects in LPS-treated RAW264.7 cells. Thus, our findings demonstrated that HD-14 alleviated inflammation by activating PPAR-γ expression at least in part. Further studies founded that HD-14 remarkably inhibited the expression of p-JAK1 and p-STAT1 through up-regulating PPAR-γ. Together, these results suggested that HD-14 served as an activator of PPAR-γ and the JAK1/STAT1 signaling pathway may be involved in the progress of inflammation. Collectively, HD-14 may be utilized as a potential anti-inflammation monomeric compound in the treatment of acute liver injury.

Tyrosine kinase inhibitor BIBF1120 ameliorates inflammation, angiogenesis and fibrosis in CCl4-induced liver fibrogenesis mouse model

Hepatic fibrosis, a progressive chronic disease mainly caused by hepatitis viral infections, alcohol abuse or metabolic syndrome leading to liver dysfunction and is the growing cause of mortality worldwide. Tyrosine kinase inhibitor BIBF1120 (Nintedanib) has been evaluated in clinical trials for idiopathic pulmonary fibrosis and advanced Hepatocellular carcinoma, but has not been explored for liver fibrosis yet. In this study, we aimed to investigate the therapeutic effects and mechanism of BIBF1120 in liver fibrogenesis. The effects of BIBF1120 were evaluated in TGFβ-activated mouse 3T3 fibroblasts, LX2 cells, primary human hepatic stellate cells (HSCs) and CCl4-induced liver fibrogenesis mouse model. Fibroblasts-conditioned medium studies were performed to assess the paracrine effects on macrophages and endothelial cells. In-vitro in TGFβ-activated fibroblasts, BIBF1120 significantly inhibited expression of major fibrotic parameters, wound-healing and contractility. In vivo in CCl4-induced acute liver injury model, post-disease BIBF1120 administration significantly attenuated collagen accumulation and HSC activation. Interestingly, BIBF1120 drastically inhibited intrahepatic inflammation and angiogenesis. To further elucidate the mechanism of action, 3T3-conditioned medium studies demonstrated increased 3T3-mediated macrophage chemotaxis and endothelial cells tube formation and activation, which was significantly decreased by BIBF1120. These results suggests that BIBF1120 can be a potential therapeutic approach for the treatment of liver fibrosis.

Hepatoprotective effect of Coreopsis tinctoria flowers against carbon tetrachloride-induced liver damage in mice

BackgroundCoreopsis tinctoria is a traditional remedy for the management of various diseases including hepatitis. The hepatoprotective role of the plant is not scientifically explored till now. This study was designed to investigate the hepatoprotective potentials of the ethanol extract from C. tinctoria (CTEtOH) using an animal model of carbon tetrachloride (CCl4)-induced acute liver injury.MethodsCTEtOH (0.5 and 1.0 g/kg) and silymarin (200 mg/kg) were administered to the experimental mice for 7 days followed by 0.2% CCl4 (10 mL/kg of body weight (bw), ip), then all mice were sacrificed after 24 h. The serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were measured. Histological analysis of liver was performed. The tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), nitric oxide (NO), malondialdehyde (MDA), and antioxidant enzymatic activities were also measured..ResultsThe results revealed that the serum ALT and AST levels significantly decreased after treatment with CTEtOH. Moreover, histological analyses indicated that CTEtOH (0.5 and 1.0 g/kg) and silymarin reduced the extent of CCl4-induced liver lesions. CTEtOH (0.5 and 1.0 g/kg) reduced the levels of malondialdehyde, nitric oxide, and proinflammatory cytokines (TNF-α and IL-1β). Furthermore, CTEtOH (1.0 g/kg) reduced the level of IL-6. The activities of antioxidant enzymes, namely superoxide dismutase and glutathione reductase, significantly increased after treatment with CTEtOH (0.5 and 1.0 g/kg) and that of glutathione peroxidase increased after treatment with 1.0 g/kg of CTEtOH.ConclusionsThese results demonstrate the hepatoprotective effect of CTEtOH against CCl4-induced acute liver injury in mice, and the underlying hepatoprotective mechanisms are associated with antioxidant and antiproinflammatory activities.

Obeticholic acid protects against carbon tetrachloride-induced acute liver injury and inflammation

The farnesoid X receptor (FXR) is a ligand-activated transcription factor that plays important roles in regulating bile acid homeostasis. The aim of the present study was to investigate the effects of obeticholic acid (OCA), a novel synthetic FXR agonist, carbon tetrachloride (CCl4)-induced acute liver injury. Mice were intraperitoneally injected with CCl4 (0.15ml/kg). In CCl4+OCA group, mice were orally with OCA (5mg/kg) 48, 24 and 1h before CCl4. As expected, hepatic FXR was activated by OCA. Interestingly, OCA pretreatment alleviated CCl4-induced elevation of serum ALT and hepatic necrosis. Moreover, OCA pretreatment inhibited CCl4-induced hepatocyte apoptosis. Additional experiment showed that OCA inhibits CCl4-induced hepatic chemokine gene Mcp-1, Mip-2 and Kc. Moreover, OCA inhibits CCl4-induced hepatic pro-inflammatory gene Tnf-α and Il-1β. By contrast, OCA pretreatment elevated hepatic anti-inflammatory gene Il-4. Further analysis showed that OCA pretreatment inhibited hepatic IκB phosphorylation and blocked nuclear translocation of NF-κB p65 and p50 subunits during CCl4-induced acute liver injury. In addition, OCA pretreatment inhibited hepatic Akt, ERK and p38 phosphorylation in CCl4-induced acute liver injury. These results suggest that OCA protects against CCl4-induced acute liver injury and inflammation. Synthetic FXR agonists may be effective antidotes for hepatic inflammation during acute liver injury.

Augmenter of liver regeneration promotes the proliferation of HL-7702 cells in carbon tetrachloride-induced acute liver injury via increasing autophagy

Objective: To investigate the protective effect of augmenter of liver regeneration (ALR) against acute liver injury and related mechanisms. Methods: HL-7702 cells were divided into normal control group, carbon tetrachloride (CCl4)-induced acute liver injury group, ALR+CCl4 intervention group, 3-methyladenine (3-MA)+CCl4 intervention group, and ALR+3-MA+CCl4 intervention group. The ALR+CCl4 and ALR+3-MA+CCl4 intervention groups were transfected with ALR plasmids at 8 hours before CCl4 treatment. All groups except the normal control group were treated with CCl4, and 30 minutes later, the 3-MA+CCl4 and ALR+3-MA+CCl4 intervention groups were treated with 3-MA. The cells were collected at 24 hours after CCl4 treatment. The HL-7702 cells and supernatant were collected to measure the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) (IU/L). Western blot was used to measure the levels of ALR, cyclin D, cyclin E, proliferating cell nuclear antigen (PCNA), autophagy-related gene 7 (Atg7), and autophagy genes LC3, p62, and Beclin-1. Quantitative real-time PCR was used to measure the mRNA expression of ALR. A one-way analysis of variance was used for comparison of means between any two groups. Results: The ALR+CCl4 intervention group had significant increases in the protein and mRNA expression of ALR compared with the acute liver injury group (both P < 0.05). The CCl4-induced acute liver injury group had significant increases in the protein and mRNA expression of ALR compared with the normal control group (both P < 0.05). Compared with the CCl4-induced acute liver injury group, the ALR+CCl4 intervention group had significant reductions in ALT (0.73±0.17 IU/L vs 1.43±0.38 IU/L, P < 0.05) and AST (19.85±1.83 IU/L vs 56.73±6.25 IU/L, P < 0.05) in supernatant, significantly increased expression of cyclin D, cyclin E, PCNA, LC3, Atg7, and Beclin-1 in hepatocytes, and significantly reduced expression of p62, which suggested that ALR protected the liver against acute liver injury, promoted the regeneration of hepatocytes, and enhanced the autophagy of hepatocytes. The ALR+3-MA+CCl4 intervention group had a significant reduction in the expression of regeneration-associated proteins compared with the ALR+CCl4 intervention group, while there was no significant difference between the ALR+3-MA+CCl4 intervention group and 3-MA+CCl4 intervention group, which suggested that after the inhibition of autophagy, there were significant reductions in the regeneration of hepatocytes and liver regeneration promoted by ALR. Conclusion: ALR can promote the regeneration of hepatocytes in liver parenchyma, which is achieved by the regulation of autophagy.

Pequi (Caryocar brasiliense Camb.) almond oil attenuates carbon tetrachloride-induced acute hepatic injury in rats: Antioxidant and anti-inflammatory effects

Carbon tetrachloride (CCl4) is a potent hepatotoxin, capable of generating free radicals that lead to oxidative stress and the inflammation process. Pequi almond oil (PAO) has been reported to possess unsaturated fatty acid and antioxidant compounds related to beneficial effects on oxidation and inflammatory conditions. The present study was undertaken to evaluate the hepatoprotective effects of handmade and coldpressed PAO on CCl4-induced acute liver injury. The possible mechanisms underlying the effect on liver injury enzymes, histopathological parameters, lipid profile, lipid peroxidation, and antioxidant and detoxification defense systems, as well as inflammatory parameters, were determined. Rats treated with PAO (3 or 6 mL/kg) for 21 days before CCl4 induction (3 mL/kg, 70%) showed significantly decreased levels of alanine aminotransferase and aspartate aminotransferase, milder hepatic lesions and higher levels of serum high-density lipoprotein compared to CCl4 group. Moreover, PAO enhanced antioxidant capacity by increasing hepatic glutathione peroxidase and glutathione reductase enzyme activities, as well as reducing circulating concentrations of leptin and inflammatory mediators such as interleukin-6, leukotrienes -4 and -5 and the tumor necrosis factor receptor. In summary, PAO, especially cold-pressed oil, attenuated the CCl4-induced alterations in serum and hepatic tissue in rats due to its antioxidant and anti-inflammatory properties.

Protective effect of rhamnopyranosyl vanilloyl isolated from &lt;i&gt;Scrophularia ningpoensis&lt;/i&gt; Hemsl (Scrophulariaceae) root against acute liver injury in mice

Purpose : To investigate the protective effect of rhamnopyranosyl vanilloyl (RV) from Scrophularia ningpoensis root against tetrachloromethane (CCl4)-induced acute liver injury (ALI) in mice. Methods : RV was isolated from S. ningpoensis by column chromatography. ALI model of mice was established by intraperitoneal injection of CCl4. Liver index, liver function indices, as well as serum alanine transaminase (ALT), aspartate aminotransferase (AST) and total bilirubin (TBIL) were evaluated. Lipid peroxidation (LPO)-related indices, including malonaldehyde (MDA), glutathione (GSH), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px). Apoptotic proteins (Bcl-2, Bax and caspase-3) in liver tissue were determined by enzyme-linked immunosorbent assay (ELISA) and Western blot. Results : After treatment with RV (10, 20 or 40 mg/kg), liver index (5.65 - 5.21 vs. 6.68 %), ALT (90.18 - 79.68 vs. 112.47 U/L), AST (64.44 - 57.63 vs. 75.41 U/L) and TBIL (2.68 - 1.95 vs. 3.21 U/L) activities, as well as MDA (3.58 - 2.88 vs. 4.13 μmol/g), Bax and caspase-3 levels significantly (p < 0.05 or 0.01) decreased, compared with those in control group. After treatment with RV (10, 20 or 40 mg/kg), GSH (16.58 - 22.14 vs. 12.34 μmol/g), Bcl-2, SOD (86.45 - 107.61 vs. 68.43 U/mg) and GSH-Px (295.64 - 329.47 vs. 268.49 U/mg) levels or activities significantly (p < 0.05 or 0.01) increased, compared with those in control group. Conclusion : RV has protective effect against CCl4-induced ALI in mice, and the mechanisms involve the inhibition of LPO and apoptosis in liver cells. Thus, RV is a potential drug for the treatment of liver injury Keywords : Rhamnopyranosyl vanilloyl, Scrophularia ningpoensis, Liver injury, Protective effect, Lipid peroxidation, Apoptosis

Serum metabonomics study of the hepatoprotective effect of Corydalis saxicola Bunting on carbon tetrachloride-induced acute hepatotoxicity in rats by 1H NMR analysis

Corydalis saxicola Bunting (CS), a traditional Chinese folk medicine, has been effectively used for treating liver disease in Zhuang nationality in South China. However, the exact hepatoprotective mechanism of CS was still looking forward to further elucidation by far. In present work, metabonomic study of biochemical changes in the serum of carbon tetrachloride (CCl4)-induced acute liver injury rats after CS treatment were performed using proton nuclear magnetic resonance (1H-NMR) analysis. Metabolic profiling by means of principal components analysis (PCA) and partial least squares-discriminate analysis (PLS-DA) indicated that the metabolic perturbation caused by CCl4 was reduced by CS treatment. A total of 9 metabolites including isoleucine (1), lactate (2), alanine (3), glutamine (4), acetone (5), succinate (6), phosphocholine (7), d-glucose (8) and glycerol (9) were considered as potential biomarkers involved in the development of CCl4-induced acute liver injury. According to pathway analysis by metabolites identified and correlation network construction by Pearson’s correlation coefficency matrix, alanine, aspartate and glutamate metabolism and glycerolipid metabolism were recognized as the most influenced metabolic pathways associated with CCl4 injury. As a result, notably, deviations of metabolites 1, 3, 4, 7 and 9 in the process of CCl4-induced acute liver injury were improved by CS treatment, which suggested that CS mediated synergistically abnormalities of the metabolic pathways, composed of alanine, aspartate and glutamate metabolism and glycerolipid metabolism. In this study, it was the first report to investigate the hepatoprotective effect of the CS based on metabonomics strategy, which may be a potentially powerful tool to interpret the action mechanism of traditional Chinese folk medicines.

Involvement of TGF-β1/Smad3 Signaling in Carbon Tetrachloride-Induced Acute Liver Injury in Mice.

Transforming growth factor-beta1 (TGF-β1) is a major factor in pathogenesis of chronic hepatic injury. Carbon tetrachloride (CCl4) is a liver toxicant, and CCl4-induced liver injury in mouse is a classical animal model of chemical liver injury. However, it is still unclear whether TGF-β1 is involved in the process of CCl4-induced acute chemical liver injury. The present study aimed to evaluate the role of TGF-β1 and its signaling molecule Smad3 in the acute liver injury induce by CCl4. The results showed that CCl4 induced acute liver injury in mice effectively confirmed by H&E staining of liver tissues, and levels of not only liver injury markers serum ALT and AST, but also serum TGF-β1 were elevated significantly in CCl4-treated mice, compared with the control mice treated with olive oil. Our data further revealed that TGF-β1 levels in hepatic tissue homogenate increased significantly, and type II receptor of TGF-β (TβRII) and signaling molecules Smad2, 3, mRNA expressions and Smad3 and phospho-Smad3 protein levels also increased obviously in livers of CCl4-treated mice. To clarify the effect of the elevated TGF-β1/Smad3 signaling on CCl4-induced acute liver injury, Smad3 in mouse liver was overexpressed in vivo by tail vein injection of Smad3-expressing plasmids. Upon CCl4 treatment, Smad3-overexpressing mice showed more severe liver injury identified by H&E staining of liver tissues and higher serum ALT and AST levels. Simultaneously, we found that Smad3-overexpressing mice treated with CCl4 showed more macrophages and neutrophils infiltration in liver and inflammatory cytokines IL-1β and IL-6 levels increment in serum when compared with those in control mice treated with CCl4. Moreover, the results showed that the apoptosis of hepatocytes increased significantly, and apoptosis-associated proteins Bax, cytochrome C and the cleaved caspase 3 expressions were up-regulated in CCl4-treated Smad3-overexpressing mice as well. These results suggested that TGF-β1/Smad3 signaling was activated during CCl4-induced acute liver injury in mice, and Smad3 overexpression aggravated acute liver injury by promoting inflammatory cells infiltration, inflammatory cytokines release and hepatocytes apoptosis. In conclusion, the activation of TGF-β signaling contributes to the CCl4-induced acute liver injury. Thus, TGF-β1/Smad3 may serve as a potential target for acute liver injury therapy.

Protective Role of Grape Seed Proanthocyanidins Against Ccl4 Induced Acute Liver Injury in Mice.

BackgroundWe investigated the effect of grape seed proanthocyanidins (GSPs) on carbon tetrachloride (CCl4)-induced acute liver injury.Material/MethodsSixty SPF KM mice were randomly divided into 6 groups: the control group, CCl4-model group, bifendate group (DDB group), and low-, moderate-, and high-dose GSP groups. The following parameters were measured: serum levels of alanine aminotransferase (ALT); aspartate aminotransferase (AST); tumor necrosis factor (TNF)-α; interleukin-6 (IL-6); high-mobility group box (HMGB)-1; body weight; liver, spleen, and thymus indexes; superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activity; HMGB1 mRNA; malondialdehyde (MDA) content; hepatocyte proliferation; and changes in liver histology.ResultsCompared to the CCl4-model group, decreases in liver index and increases in thymus index significantly increased SOD and GSH-Px activities and reduced MDA content, and higher hepatocyte proliferative activity was found in all GSP dose groups and the DDB group (all P<0.001). Compared with the CCl4-model group, serum TNF-α and IL-6 levels and HMGB 1 mRNA and protein expressions decreased significantly in the high GSP dose group (all P<0.05).ConclusionsOur results provide strong evidence that administration of GSPs might confer significant protection against CCl4-induced acute liver injury in mice.

Protective effect of wedelolactone against CCl4-induced acute liver injury in mice

Eclipta, a traditional Chinese medicine, has been used to treat liver disease for centuries. However, the chemical basis and biological mechanisms of Eclipta remain elusive. The current study aims to investigate the hepatoprotective effect of wedelolactone (WEL), a major coumarin in Eclipta, using C57BL/6 mice with carbon tetrachloride CCl4-induced acute liver injury (ALI). Our data showed that WEL markedly decreased the CCl4-induced elevation of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, and improved hepatic histopathology changes. WEL also significantly decreased the content of MDA in liver tissues, meanwhile increased the activities of antioxidant enzymes SOD and GSH-Px. In addition, WEL reduced the protein expression of TNF-α, IL-1β and IL-6, as well as mRNA expression. Western blot results revealed that WEL repressed phosphorylation of extracellular signal-regulated kinase (ERK) and translocation of NF-κB p65 from cytoplasm to nucleus and enhanced the phosphorylation of c-Jun. N-terminal kinase (JNK). Moreover, results showed that WEL significantly inhibited CCl4-induced hepatocytes apoptosis, markedly suppressed the down-regulation of Bax and active Caspase-3 expression and accelerated the expression of Bcl-2. Overall, the findings indicate that WEL exhibits a protective effect against CCl4-induced ALI in mice by enhancing the antioxidative defense system, suppressing the inflammatory response and cell apoptosis of liver.

Hepatoprotective Effects of Rubus idaeus L. Leaves against CCl4-Induced Liver Injury via Antioxidant, Anti-inflammatory and Antiapoptotic Mechanisms

Aims: To investigate the hepatoprotective effects of red raspberry (Rubus idaeus L.) leaves against oxidative stress, inflammation and apoptosis induced by carbon tetrachloride (CCl4) in rats. Study Design and Methodology: Forty rats were divided into 5 groups, 1) normal control; 2) Raspberry (Rsp) control (100 mg/kg); 3) CCl4 control; 4) and 5) CCl4 groups pre-treated with 50 and 100 mg/kg of Rsp leaves, respectively, once daily for 10 days. Liver injury was induced on the 11th day in groups 3, 4 and 5 by intraperitoneal injection of CCl4 (1.0 ml/kg) in 50% olive oil (1:1). Serum and liver were separated and prepared for biochemical and histological assay. Antioxidant, inflammatory and apoptotic markers were evaluated. Results: CCl4-induced liver damage was manifested by increased activities of serum marker enzymes, elevated levels of lipid peroxidation and by decreased hepatic antioxidants including reduced glutathione, glutathione peroxidase, superoxide dismutase and catalase. CCl4 also resulted in elevated levels of hepatic inflammatory mediators including tumor necrosis factor-α (TNFα), interleukin -6 (IL-6), IL-1β and nitric oxide (NO) as well as upregulation of apoptotic markers including caspase-3, FasL and Bax. Hepatic levels of Bcl2, an anti-apoptotic factor, was Original Research Article Attia; EJMP, 14(1): 1-16, 2016; Article no.EJMP.25300 2 decreased by CCl4. Pretreatment with Rsp leaves significantly ameliorated the elevation in serum liver enzymes and prevented lipid peroxidation and the depletion of the antioxidants. In addition, Rsp leaves exhibited anti-inflammatory effects via attenuation of increased hepatic levels of TNF-α, Il-6, IL-1β and NO. Additionally, CCl4-Induced apoptosis in the liver cell was attenuated by Rsp pretreatment. With regard to histological examination, Rsp leaves significantly alleviated degeneration and necrosis of hepatocytes, accompanied by decreased inflammatory cell infiltration. Conclusion: These results suggest that Rsp leaves have protective effects against CCl4-induced acute liver injury, and this protection is likely due to antioxidant, anti-inflammatory and antiapoptotic mechanisms.

Resolvin D1 attenuates CCl4-induced acute liver injury involving up-regulation of HO-1 in mice

Acute hepatic failure involves in excessive oxidative stress and inflammatory responses, leading to a high mortality due to lacking effective therapy. Resolvin D1 (RvD1), an endogenous lipid mediator derived from polyunsaturated fatty acids, has been shown anti-inflammatory and anti-oxidative actions, however, whether RvD1 has protective effects on hepatic failure remains elusive. In this study, the roles and molecular mechanisms of RvD1 were explored in carbon tetrachloride (CCl4)-induced acute liver injury. Our results showed that RvD1 protected mice against CCl4-induced hepatic damage, as evaluated by reduced aminotransferase activities and malondialdehyde content, elevated glutathione and superoxide dismutase activities, and alleviated hepatic pathological damage. Moreover, RvD1 significantly attenuated serum tumor necrosis factor-α and interleukin-6 levels as well as hepatic myeloperoxidase activity, whereas enhanced serum IL-10 level in CCl4-administered mice. Further, RvD1 markedly up-regulated the expression and activity of heme oxygenase-1 (HO-1). However, inhibition of HO-1 activity reversed the protective effects of RvD1 on CCl4-induced liver injury. These results suggest that RvD1 could effectively prevent CCl4-induced liver injury by inhibition of oxidative stress and inflammation, and the underlying mechanism may be related to up-regulation of HO-1.

Glycoprotein Nonmetastatic Melanoma B (Gpnmb)-Positive Macrophages Contribute to the Balance between Fibrosis and Fibrolysis during the Repair of Acute Liver Injury in Mice.

Background and aimsGlycoprotein nonmetastatic melanoma B (Gpnmb), a transmembrane glycoprotein that is expressed in macrophages, negatively regulates inflammation. We have reported that Gpnmb is strongly expressed in the livers of rats fed a choline-deficient, L-amino acid-defined (CDAA) diet. However, the role of macrophage-expressed Gpnmb in liver injury is still unknown. This study aimed to clarify the characteristics of infiltrating macrophages that express Gpnmb, and the involvement of Gpnmb in the repair process in response to liver injury.MethodsC57BL/6J, DBA/2J [DBA] and DBA/2J-Gpnmb+ [DBA-g+] mice were treated with a single intraperitoneal injection of carbon tetrachloride (CCl4) at a dose of 1.0 mL/kg body weight. Mice were sacrificed at predetermined time points, followed by measurement of serum alanine aminotransferase (ALT) levels and histological examination. Expression of Gpnmb, pro-/anti-inflammatory cytokines, and profibrotic/antifibrotic factors were examined by quantitative RT-PCR and/or Western blotting. Immunohistochemistry, fluorescent immunostaining and flow cytometry were used to determine the expression of Gpnmb, CD68, CD11b and α-SMA, phagocytic activity, and the presence of apoptotic bodies. We used quantitative RT-PCR and ELISA to examine TGF-β and MMP-13 expression and the concentrations and supernatants of isolated infiltrating hepatic macrophages transfected with siGpnmb.ResultsIn C57BL/6J mice, serum ALT levels increased at two days after CCl4 injection and decreased at four days. Gpnmb expression in the liver was stimulated four days after CCl4 injection. Histological examination and flow cytometry showed that Gpnmb-positive cells were almost positive for CD68-positive macrophages, contained engulfed apoptotic bodies and exhibited enhanced phagocytic activity. Isolated infiltrating hepatic macrophages transfected with siGpnmb showed high MMP-13 secretion. There was no significant difference in the magnitude of CCl4-induced liver injury between DBA-g+ and DBA mice. However, hepatic MMP-13 expression, as well as α-SMA expression and collagen production, increased significantly in DBA-g+ compared with DBA mice.ConclusionsGpnmb-positive macrophages infiltrate the liver during the recovery phase of CCl4–induced acute liver injury and contribute to the balance between fibrosis and fibrolysis in the repair process following acute liver injury.