CCl4-treated Rats Research Articles

Role of N-Acetylcysteine and Coenzyme Q10 in the Amelioration of Myocardial Energy Expenditure and Oxidative Stress, Induced by Carbon Tetrachloride Intoxication in Rats.

This study is designed to evaluate the potential impact of N-acetyl cysteine (NAC) and coenzyme Q10 (CoQ10) each alone or in combination against carbon tetrachloride (CCl4)-induced cardiac damage in rats. Animals were treated with CCl4 in single intraperitoneal dose of 1 mL/Kg body weight; CCl4-intoxicated animals were pretreated with 20 mg/kg/d NAC or pretreated with 200 mg/kg/d CoQ10 or NAC and CoQ10 with the same previously mentioned doses. Carbon tetrachloride–intoxicated rats showed a significant elevation in nitric oxide and lipid peroxides and downregulation in reduced glutathione level and calcium adenosine triphosphatase. Cardiac glycolytic enzymes levels such as lactate dehydrogenase, phosphofructokinase, and hexokinase were declined coupled with a reduction in glucose content after CCl4 treatment. Moreover, myocardial hydroxyproline level was significantly increased after CCl4-treatment indicating accumulation of interstitial collagen. N-acetyl cysteine and/or CoQ10 effectively alleviated the disturbances in myocardial oxidative stress and antioxidant markers. These antioxidants effectively upregulated the reduction in cardiac energetic biomarkers due to CCl4 treatment. N-acetyl cysteine and/or CoQ10 significantly decreased hydroxyproline level compared to that of CCl4-treated rats. The current data showed that the aforementioned antioxidants have a remarkable cardioprotective effect, suggesting that they may be useful as prophylactic agents against the detrimental effects of cardiotoxins.

Carbon tetrachloride administration induces the expression of hypoxia inducible factor-1alpha in rat liver

Background: There is now increasing evidence that HIF-1 is also responsive to a variety of non-hypoxic stimuli. However, the mechanisms by which these non-hypoxic stimuli induce HIF-1α are not completely known, yet, although some evidence points to a role of ROS as messengers regulating HIF activity.
 Objective: To determine the expression of HIF-1α in liver rat tissue induced by carbon tetrachloride under normoxic conditions, with or without N-acetylcysteine protection.
 Methods: Twenty-five male Sprague-Dawley rats were divided into 5 groups: normal control rats, normal rats orally administered with coconut oil (1 mL/200 g body weight) for 1 day, rats orally administered with CCl4 (0.55 mg/g body weight) for 1 day, rats injected i.v. with NAC (0.15 mg/g body weight) for 8 days and then orally administered with CCl4 (0.55 mg/g body weight) for 1 day, rats orally administered with CCl4 (0.55 mg/g body weight) for 1 day and then injected i.v. with NAC (0.15 mg/g body weight) for 2 days. The expression of HIF-1α mRNA was measured by real-time RT-PCR using the Livak method. The expression of HIF-1α protein was measured by ELISA assay.
 Results: The highest HIF-1α mRNA and protein expression found in the group treated by CCl4 and then was gradually lowered in the pre-NAC group, post-NAC group, control group, and last, in the oil group.
 Conclusion: Our study shows the effect of CCl4-treated rats under normoxic conditions increased the mRNA and protein HIF-1α. NAC post-treatment provide a better protective effect compared with NAC pre-treatment

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.

High resolution UPLC-MS/MS profiling of polyphenolics in the methanol extract of Syzygium samarangense leaves and its hepatoprotective activity in rats with CCl4-induced hepatic damage

Oxidative stress plays a crucial role in the development of several liver diseases. Many natural polyphenols can attenuate oxidative stress and liver injury. In this study, a phytochemical profiling of a methanol extract from leaves of Syzygium samarangense revealed 92 compounds belonging to flavonoids, phenolic acids, condensed tannins, and ellagitannins. The S. samarangense extract exhibited a noticeable antioxidant activity with an EC50 of 5.80 μg/mL measured by DPPH scavenging capacity assay, 2632 Trolox equivalents, 10 mM Fe2+ equivalents/mg of samples by TEAC and FRAP assays, respectively. The total phenolic content was 419 mg gallic acid equivalent GAE/g extract. In a cell-based model (HaCaT cells), the extract completely inhibited ROS production induced by UVA, and prevented GSH-depletion and p38 phosphorylation. In addition, the extract exhibited a substantial antioxidant and hepatoprotective activities in CCl4-treated rats, with an increase in GSH (reduced glutathione) and SOD (superoxide dismutase) activities by 84.75 and 26.27%, respectively, and a decrease of 19.08, 63.05, 52.21, 37.00, 13.26, and 15.15% in MDA, ALT, AST, TB (total bilirubin), TC (total cholesterol), and TG (total glycerides), respectively. These results were confirmed by histopathological analyses. We believe that Syzygium samarangense is a good candidate for further evaluation as an antioxidant and liver protecting drug.

Fresh Bitter Melon Fruit (Momordica charantia) Attenuated Oxidative Stress, Fibrosis and Renal Injury in Carbon Tetrachloride Treated Rats

Chemical or drug-induced kidney damage is increasing every year and the end-stage renal disease is becoming a burden for health care system of many countries. Oxidative stress may be a crucial pathway for the development of end-stage renal disease. Thus, natural antioxidant or plant-based therapy would be a better alternative to protect renal function against chemical-induced renal damage. To determine these aspects we evaluated renoprotective effects of M. charantia in carbon tetrachloride administered rats. A 10% w/w mixture of fresh fruits of M. charantia was given with the chow food every day to CCl4 treated rats. After fourteen days, all animals were sacrificed and the kidneys were examined to observe the possible protective effects of M. charantia against CCl4 induced toxicity. The CCl4 treated rats showed increased oxidative stress parameters and decreased antioxidant enzymes activities. Supplementation of 10% w/w M. charantia fruits in CCl4 administered rats prevented the oxidative stress and restored the antioxidant enzymes activities. M. charantia fruits supplementation also prevented the rise of uric acid and creatinine concentration in plasma of CCl4 treated rats. Furthermore, histological studies showed that supplementation of 10% w/w M. charantia fruits prevented the collagen deposition, immune cell migration and iron deposition in kidney sections of CCl4 treated rats. The results of this study revealed that the fruits of M. charantia may protect oxidative stress-mediated damage in kidneys due to CCl4 administration, which is mediated probably via the restoration of anti-oxidant enzyme functions.Dhaka Univ. J. Pharm. Sci. 16(2): 205-214, 2017 (December)

Protective effects of Mentha piperita L. leaf essential oil against CCl4 induced hepatic oxidative damage and renal failure in rats

BackgroundMentha piperita L. is a flowering plant belonging to the Lamiaceae family. Mentha plants constitute one of the main valuable sources of essential oil used in foods and for medicinal purposes.MethodsThe present study aimed to investigate the composition and in vitro antioxidant activity of Mentha piperita leaf essential oil (MpEO). A single dose of CCl4 was used to induce oxidative stress in rats, which was demonstrated by a significant rise of serum enzyme markers. MpEO was administrated for 7 consecutive days (5, 15, 40 mg/kg body weight) to Wistar rats prior to CCl4 treatment and the effects on serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), lactate dehydrogenase (LDH), and γ -glutamyl transpeptidase (γ-GT) levels, as well as the liver and kidney superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) activity and thiobarbituric acid reactive substances (TBARS) levels were evaluated. In addition, histopathological examinations of livers and kidneys was performed.ResultsThe in vitro antioxidant activity of MpEO was lower than that of silymarin. Pretreatment of animals with MpEO at a dose of 5 mg/kg did not have a significant effect on ALT, AST, ALP, LDH, γGT, urea or creatinine levels in CCl4-induced stress. Whereas pretreatment with MpEO at doses of 15 and 40 mg/kg prior to CCl4, significantly reduced stress parameters (ALT, AST, ALP, LDH, γGT, urea and creatinine) compared to the CCl4-only group. Moreover, a significant reduction in hepatic and kidney lipid peroxidation (TBARS) and an increase in antioxidant enzymes SOD, CAT and GPx was also observed after treatment with MpEO (40 mg/kg) compared to CCl4-treated rats. Furthermore, pretreatment with MpEO at 40 mg/kg can also markedly ameliorate the histopathological hepatic and kidney lesions induced by administration of CCl4.ConclusionsWe could demonstrate with this study that MpEO protects liver and kidney from CCl4-induced oxidative stress and thus substantiate the beneficial effects attributed traditionally to this plant.

Hepatoprotective Effect of Eriobotrya japonica Leaf Extract and Its Various Fractions against Carbon Tetra Chloride Induced Hepatotoxicity in Rats.

Eriobotrya japonica is traditionally used as an antipyretic, digestive, and diuretic agent. Its flowers possess free radical–scavenging, antioxidative, and hepatoprotective effects. We investigated the hepatoprotective potential of E. japonica leaf extract and its various fractions against hepatotoxicity in rats. Liver injury was stimulated by the oral administration of carbon tetrachloride (CCl4; 2.5 mL/kg b.wt.). Male albino rats (n = 55) were distributed arbitrarily into 11 groups: Group I, normal control group; Group II, CCl4 (positive control group); Group III, CCl4 + silymarin; Groups IV and V, CCl4 + two doses of 250 and 500 mg/kg of the 80% methanolic extract of E. japonica leaves, respectively; Groups VI and VII, CCl4 + 250 mg/kg and 500 mg/kg of the ethyl acetate fraction, respectively; Groups VIII and IX, CCl4 + 250 and 500 mg/kg of the butanol fraction, respectively; and Groups X and XI, CCl4 + 250 and 500 mg/kg of the aqueous fraction of E. Japonica leaves, respectively. CCl4-treated rats that were given 250 or 500 mg/kg of the methanol extract of E. Japonica leaves, or its ethyl acetate, butanol, or aqueous fractions, had significantly lower levels of biochemical parameters such as alanine aminotransferase, aspartate transaminase, alkaline phosphate, total protein, gamma-glutamyl transferase, and bilirubin levels than those of the CCl4 positive group. However, the extract and fractions did not significantly affect lipid profiles. Thus, we conclude that Eriobotrya leaf extract and its fractions have a hepatoprotective effect against CCl4-induced hepatotoxicity in rats.

Lyophilized Extract of the Horse Mushroom, Agaricus arvensis (Agaricomycetes), Delays Erythrocyte Hemolysis and Stabilizes Some Blood Parameters in Carbon Tetrachloride-Intoxicated Rats.

This study evaluates the protective effects of a lyophilized extract of the horse mushroom, Agaricus arvensis, on hematological parameters and its antioxidative role against carbon tetrachloride (CCl4)-induced oxidative stress in rats. The protective effects of the A. arvensis extract were evaluated by comparing hematological parameters, erythrocyte fragility, and components of the antioxidant defense system (e.g., reduced glutathione [GSH] level; glutathione S-transferase, catalase, glutathione peroxidase [GPx], and superoxide dismutase activities; and malondialdehyde content in erythrocytes). The 4 experimental groups included the controls (group I), CCl4-treated rats (group II), rats treated with CCl4 + 100 mg/kg A. arvensis extract (group III), and those treated with CCl4 + 500 mg/kg A. arvensis extract (group IV). Erythrocyte hemolysis in 0.6% NaCl was lower in groups I, III, and IV (at 38%, 22%, and 34%, respectively) than in group II. GSH levels in groups I, III, and IV were higher by 54%, 12%, and 11%, respectively, than that in group II. GPx enzyme activity in group II was significantly lower than the activity in groups I, III, and IV (by 138%, 70%, and 64%, respectively). In addition, administration of the A. arvensis extract resolved the dangerous effects of CCl4 on some hematological parameters such as red blood cells, hemoglobin, hematocrit, white blood cells, platelet count, plateletcrit, and red cell distribution width. We conclude that the A. arvensis extract has therapeutic effects against CCl4-induced oxidative stress, reduces erythrocyte fragility, and improves a number of hematological parameters in rats.

Swertiamarin attenuates carbon tetrachloride (CCl4)-induced liver injury and inflammation in rats by regulating the TLR4 signaling pathway

The aim of the present study is to illustrate the effects of swertiamarin (STM), a natural iridoid from herbal medicines, on hepatic inflammation induced by carbon tetrachloride (CCl4) in rats. Male Sprague Dawley rats were exposed to CCl4 with or without STM co-administration for 8 weeks. Our results revealed that STM administration (100 and 200 mg/kg b.w.) significantly attenuated inflammation in livers of CCl4-treated rats. STM remarkably reduced the production of interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α), macrophage inflammatory protein-1a (MIP-1α), and monocyte chemotactic protein-1 (MCP-1) in liver tissue of CCl4-treated rats. In addition, STM treatment downregulated connective tissue growth factor (CTGF) and ser307pIRS-1 expression, which was induced by CCl4 exposure. In the process of exploring the anti-inflammatory mechanisms of STM action, we demonstrated that STM significantly inhibited Toll-like receptor 4 (TLR4) and nuclear factor kappa B (NF-κB) p65 expression in the liver. In conclusion, these results suggested that the inhibition of CCl4-induced inflammation by STM was, at least in part, due to its regulation of the TLR4 /NF-κB signaling pathway.

Sildenafil, a phosphodiesterase-5 inhibitor, offers protection against carbon tetrachloride-induced hepatotoxicity in rat.

Elevation of phosphodiesterase-5 (PDE5) activity converts cyclic guanosine monophosphate (cGMP) to 5'-GMP, a mechanism that could be associated with drug-mediated hepatotoxicity. This study investigated whether selective inhibition of PDE5 by sildenafil could offer protection against hepatotoxicity induced by carbon tetrachloride (CCl4). CCl4 (0.5 mL/kg) was administered intraperitoneally to induce hepatotoxicity. The control group received normal saline. Sildenafil (5 mg, 10 mg, and 20 mg/kg, p.o.) was administered to CCl4-treated rats. CCl4 significantly increased the serum levels of gamma glutamyl transferase (γ-GT), alkaline phosphatase (ALP), aspartate aminotransferase (AST), and alanine aminotransferase (ALT) and reduced total protein (TP) (p<0.05). Pretreatment with sildenafil moderately reduced ALP, AST, and ALT activities with modest increase in TP level. CCl4-induced changes in the antioxidant status of the liver were significantly improved by sildenafil, especially at the lowest dose of 5 mg/kg by elevating the levels of reduced glutathione (GSH), glutathione peroxidase (GPx), catalase (CAT), superoxide dismutase (SOD), and glutathione-S-transferase (GST) and preventing lipid peroxidation (p<0.05). Sildenafil did not significantly alter the total cholesterol and triglyceride levels. However, high-density lipoprotein (HDL) level was significantly increased by sildenafil (p<0.05). The results from this study suggest that sildenafil, when used at low doses, may be a useful pharmacological protective agent against CCl4-induced hepatotoxicity.

Hepatoprotective effect of methyl ferulic acid against carbon tetrachloride-induced acute liver injury in rats.

The present study aimed to investigate the hepatoprotective effects of methyl ferulic acid (MFA) against oxidative stress and apoptosis in acute liver injury induced by carbon tetrachloride (CCl4) in rats, as well as the underlying mechanisms. Sprague Dawley rats were treated with CCl4 after oral administration of MFA (25, 50, and 100 mg/kg) or dimethyl diphenyl bicarboxylate (200 mg/kg) for 7 days. The hepatoprotective effects of MFA were determined by analyzing serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities as well as changes of oxidant parameters. Histopathological analysis was performed to determine the degree of hepatic injury. The mechanisms were investigated by detecting the levels of NADPH oxidase (NOX) trans-membrane subunit NOX4, its ligand p22phox, as well as caspase3, cleaved caspase3, B-cell lymphoma (Bcl)-2, Bcl-2-associated X protein (Bax), tumor necrosis factor (TNF)-α, interleukin (IL)-1, reactive oxygen species (ROS), thiobarbituric acid-reactive substances (TBARS), total anti-oxidant capacity (TAC), phosphorylated J-Jun N-terminal kinase (p-JNK) and p-p38 mitogen-activated protein kinase (MAPK) using semi-quantitative polymerase chain reaction, western blot analysis and colorimetric assays. MFA treatment significantly decreased serum enzymatic activities of ALT and AST. MFA markedly increased activities of liver superoxide dismutase, catalase and glutathione peroxidase, and reduced the malondialdehyde concentration. Histopathological examination demonstrated that MFA reduced lipid degeneration, cytoplasmic vacuolization, necrosis and inflammatory cell infiltration in the liversof CCl4-treated rats. MFA treatment markedly inhibited the expression of inflammatory factors TNF-α and IL-1β. Mechanistic study revealed that MFA decreased the TAC and the levels of ROS and TBARS. Furthermore, MFA treatment led to a reduction of the mRNA and protein expression of NOX4 and p22phox, as well as the protein levels of caspase3, cleaved caspase-3 and Bax, and an upregulation of p-JNK, p-p38 MAPK and Bcl-2 proteins in the liver. The present study demonstrated that MFA has hepatoprotective effects against CCl4-induced acute liver damage. MFA has anti-oxidant, anti-inflammatory and anti-apoptotic activities and was able to modulate the NOX4/p22phox/ROS-JNK/p38 MAPK signaling pathway.

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.

White butterfly (Clerodendrum volubile) leaf extract protects against carbon tetrachloride-induced hepatotoxicity in rats

Currently, there is increasing attention towards flavonoids and phenolic compounds of plant origin because of their association with decrease in the incidence of cardiovascular diseases and different types of cancer. The present study investigates the protective effect of Clerodendrum volubile (C. volubile) methanolic extracts against carbon tetrachloride (CCl4)-induced hepatotoxicity in rats. Control rats (group I) received olive oil (1 mL/kg, i.p.), group II received CCl4 in olive oil (1 ml/kg, i.p.) to induce hepatotoxicity, groups III, IV and V were pretreated with leaf extract of C. volubile at 125 mg/kg, 250 mg/kg and 500 mg/kg body weight respectively for 14 days prior to CCl4 administration, group VI received vitamin E (100 mg/kg, p.o.) as standard antioxidant to compare with antioxidant effects of the extract. CCl4 hepatotoxicity, characterized by significant (P < 0.05) increase in the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP), hepatic degeneration, and inflammation was attenuated by C. volubile methanolic extracts. The serum lipid parameters which include high density lipoprotein (HDL) and low density lipoprotein (LDL) were significantly (P < 0.05) decreased, and increased respectively by CCl4. Methanolic extracts of C. volubile significantly prevented the decrease in the level of HDL and the increase in LDL in a dose-dependent manner (P < 0.05). Decrease in total protein induced by CCl4 was moderately increased following administration of methanolic extracts of C. volubile. Lipid peroxidation was significantly (P < 0.05) reduced while the reduced glutathione (GSH) level and the activities of hepatic antioxidant enzymes (catalase, superoxide dismutase, and glutathione peroxidase) were significantly elevated by C. volubile extract in the CCl4-treated rats. Our findings indicate that C. volubile extract has a significant protective effect against CCl4-induced hepatotoxicity in rats which may be due to its antioxidant properties which is comparable to the reference antioxidant, vitamin E, used in this study.

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.

Deproteinization is Necessary for the Accurate Determination of Ammonia Levels by Glutamate Dehydrogenase Assay in Blood Plasma From Subjects With Liver Injury.

To determine the effect of presence of high concentrations of nicotinamide adenine dinucleotide (NADH)- and nicotinamide adenine dinucleotide phosphate (NADPH)-consuming enzymes on the accuracy of glutamate dehydrogenase (GLDH) assay for ammonia. We measured ammonia concentrations using GLDH and NADH or NADPH in blood-plasma specimens and specimens deproteinized by sulfosalicylic acid from CCl4-treated or control rats. The nonspecific oxidation of NADH and NADPH was measured in mixtures without GLDH. We observed a gradual decrease (~0.5%) in absorbance in the plasma of controls after the addition of NADH but not after adding NADPH. The decrease in absorbance in plasma of CCl4-treated animals was 13.2% and 5.2% after the addition of NADH and NADPH, respectively. The decrease in absorbance was not detected in deproteinized specimens. The values of ammonia concentration were higher in the plasma specimens compared with the deproteinized ones. Deproteinization is necessary for accurate measurement of ammonia using GLDH assay in the blood plasma of subjects with liver injury.

Antioxidant Potential of Stem Bark Extract of Shorea roxburghii against CCl 4 induced Liver Damage in Wistar Rats

The aim of the present study was to evaluate the antioxidant potential of the stem bark extract of Shorea roxburghii (S. roxburghii) on CCl4 induced liver toxicity. Stem bark extract of Shorea roxburghii (SRE) was prepared using ethyl acetate. Administration of S. roxburghii inhibited the elevation marker enzymes such as aminotransferase (AST), alkaline phosphatase (ALP) and alanine aminotransferase (ALT) and liver lipid peroxides in CCl4-treated Wistar rats. The natural antioxidant enzymes such catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPx) and lipid peroxidation were significantly improved after the treatment of S.roxburghii. In this study, the results indicate that S.roxburghii can inhibit lipid peroxidation and improve the activities of antioxidant enzymes. The quantitative analysis of biochemical parameters, marker enzymes, antioxidant activities and histological study suggest the administration of the stem bark extract of S.roxburghii protecting the liver from CCl4 toxicity.

A novel three-dimensional culture system maintaining the physiological extracellular matrix of fibrotic model livers accelerates progression of hepatocellular carcinoma cells

Liver fibrosis is characterized by the progressive accumulation of extracellular matrix (ECM) and is a strong predictor of hepatocellular carcinoma (HCC) development and progression. However, the effect of ECM in fibrotic livers on HCC cells is poorly understood. The aims of this study were to create a new culture system that retained the natural ECM of fibrotic model livers and to establish whether natural ECM regulated the characteristics of HCC cells. Using an organ decellularization technique, we created a new culture system that preserved the tissue-specific ECM of fibrotic model livers from CCl4-treated rats. The content of ECM in fibrotic model liver scaffolds was increased and the ECM microstructure was distorted. Quantitative polymerase chain reaction and immunofluorescence assays of HCC cells cultured in fibrotic model liver scaffolds for 7 days showed an epithelial-mesenchymal transition phenotype. Moreover, the ECM of fibrotic model livers promoted proliferation and chemoresistance of HCC cells. These results showed a novel effect of natural ECM in fibrotic model livers on the malignant behaviour of HCC cells. This new culture system will be useful for both understanding the cell biology of fibrotic livers and developing novel anti-cancer drugs.

MicroRNA-146a-5p attenuates liver fibrosis by suppressing profibrogenic effects of TGFβ1 and lipopolysaccharide

Liver fibrosis is characterized by proliferation and activation of hepatic stellate cells (HSCs). Transforming growth factor-β1 (TGFβ1) is crucial for liver fibrogenesis, and gut-derived endotoxin (LPS) also plays an important role in liver fibrogenesis. In the present study, we found that microRNA-146a-5p (miR-146a-5p) could regulate TGFβ1/Smad and LPS/NF-κB/Bambi pathways to attenuate liver fibrosis. Downregulated miR-146a-5p and upregulated level of LPS were found in liver of CCl4-treated rats. On cellular level, expression of miR-146a-5p is reduced during primary rat HSCs naturally activation and changed in response to TGFβ1 and/or LPS stimulation in primary rat HSCs and human HSC line LX-2. Further overexpression of miR-146a-5p suppresses proliferation and activation of HSCs. The underlying mechanism involved that miR-146a-5p directly suppresses profibrogenic effects of TGFβ1 by down-regulating the expression of Smad4 and phosphorylation of Smad2. Moreover, miR-146a-5p indirectly suppresses TGFβ1/Smad pathway by targeting IL-1 receptor-associated kinase 1 (IRAK1) and TNF receptor associated factor-6 (TRAF6), two major components of LPS/NF-κB/Bambi pathway, to reduce inhibition of TGFβ pseudoreceptor Bambi. These results indicate that miR-146a-5p abrogate hepatic fibrosis by suppressing both TGFβ/Smad and LPS/NF-κB/Bambi signaling pathway in HSCs and suggest that miR-146a-5p is a potential therapeutic target for liver fibrosis.

Hepatoprotective effects of Vaccinium arctostaphylos against CCl4-induced acute liver injury in rats.

This study was carried out to evaluate the antioxidant and hepatoprotective effects of Vaccinium arctostaphylos (V.a) methanolic extract on carbon tetrachloride (CCl4)-induced acute liver injury in Wistar rats. Total phenolic and total flavonoid contents as well as antioxidant activity of V.a were determined. Extracts of V.a at doses of 200 and 400 mg/kg were administered by oral gavage to rats once per day for 7 days and then were given an intraperitoneal injection of 1 mL/kg CCl4 (1:1 in olive oil) for 3 consecutive days. Serum biochemical markers of liver injury, oxidative markers, as well as hydroxyproline (HP) content and histopathology of liver were evaluated. The obtained results showed that V.a had strong antioxidant activity. Treatment of rats with V.a blocked the CCl4-induced elevation of serum markers of liver function and enhanced albumin and total protein levels. The level of hepatic HP content was also reduced by the administration of V.a treatment. Histological examination of the liver section revealed that V.a prevented the occurrence of pathological changes in CCl4-treated rats. These findings suggested that V.a may be useful in the treatment and prevention of hepatic injury induced by CCl4.

In vivo Pharmacological Effectiveness of Heat-treated Cucumber (Cucumis sativus L.) Juice against CCI4- induced Detoxification in a Rat Model

Abstract: Background: Detoxification of heavy metals such as carbon tetrachloride (CCl4) poisoning has become a major focus of researchers. Objectives: In this study, heat-treated cucumber juice was assessed for its pharmacological effect on protection of carbon tetrachloride (CCl4)-induced acute liver and kidney damages. Methods: Initially, during detoxification of CCl4, the rats of all groups except the control group (without treatment) were injected with a single intraperitoneal dose of 2.5 mL CCl4/kg with 50% concentration (1:1 ratio; v/v in olive oil). Additionally, CCl4-treated rats were also dosed with saline, 10 mg/kg, 100 mg/kg, and 500 mg/kg cucumber juice for 1, 3 and 5 days once in a day after 6 h from CCl4 administration. Results: As a result, the best concentration of cucumber juice was noted to be 10 mg/kg, which showed remarkable protective effect on body weight (212-217 g), enzymatic activities of aspartate aminotransferase (AST; 53 IU/L), alanine aminotransferase (ALT; 125 IU/L), blood urea nitrogen (BUN; 18.8 mg/dL), and creatinine content (0.65 mg/dL) as compared to control (220-232 g; 477 IU/L; 987 IU/L; 16.8 mg/dL; and 0.95 mg/dL), respectively. Moreover, histology and histomorphometry analysis of liver and kidney led to significant reduction in percentage of hepatopathy and nephropathy parameters such as hepatic (46.90±2.63%) and kidney (39.86±2.01) degenerative regions, number of hepatocytes (638.00±47.25), tubules (327.60±26.67) and glomerulus (29.20±1.36). Conclusion: These findings suggest that heat-treated cucumber juice has a significant pharmacological effect on CCl4-induced acute liver and kidney damages in experimental rats. Key words: Cucumis sativus, Cucumber juice, CCl4 detoxification, AST, ALT, BUN.