Ascorbic acid suppresses endotoxemia and NF-κB signaling cascade in alcoholic liver fibrosis in guinea pigs: a mechanistic approach.

Alcohol consumption increases the small intestinal bacterial overgrowth (SIBO) and intestinal permeability of endotoxin. The endotoxin mediated inflammatory signaling plays a major role in alcoholic liver fibrosis. We evaluated the effect of ascorbic acid (AA), silymarin and alcohol abstention on the alcohol induced endotoxemia and NF-κB activation cascade pathway in guinea pigs (Cavia porcellus). Guinea pigs were administered ethanol at a daily dose of 4g/kg b.wt for 90days. After 90days, ethanol administration was stopped. The ethanol treated animals were divided into abstention, silymarin (250mg/kg b.wt) and AA (250mg/kg b.wt) supplemented groups and maintained for 30days. The SIBO, intestinal permeability and endotoxin were significantly increased in the ethanol group. The mRNA expressions of intestinal proteins claudin, occludin and zona occludens-1 were significantly decreased in ethanol group. The mRNA levels of inflammatory receptors, activity of IKKß and the protein expressions of phospho-IκBα, NF-κB, TNF-α, TGF-ß1 and IL-6 were also altered in ethanol group. The expressions of fibrosis markers α-SMA, α1 (I) collagen and sirius red staining in the liver revealed the induction of fibrosis. But the supplementation of AA could induce greater reduction of ethanol induced SIBO, intestinal barrier defects, NF-κB activation and liver fibrosis than silymarin. The possible mechanism may be the inhibitory effect of AA on SIBO, intestinal barrier defect and IKKß, which decreased the activation of NF-κB and synthesis of cytokines. This might have led to suppression of HSCs activation and liver fibrosis.

Ascorbic acid is superior to silymarin in the recovery of ethanol-induced inflammatory reactions in hepatocytes of guinea pigs.

Both oxidative stress and inflammatory reactions play a major role in alcoholic liver fibrosis. We evaluated the efficacy of ascorbic acid (AA) and silymarin in the regression of alcohol-induced inflammation in hepatocytes of guinea pigs (Cavia porcellus). Animals were administered with ethanol at a daily dose of 4 g/kg body weight (b.wt) for 90 days. On the ninety-first day, ethanol administration was stopped and animals were divided into alcohol abstention group and silymarin- (25 mg/100 g b.wt) and AA- (25 mg/100 g b.wt) supplemented groups and maintained for 30 days. There was a significant increase in the activities of alanine aminotransferase, aspartate aminotransferase, and γ-glutamyl transpeptidase in the serum of the ethanol group. The intracellular reactive oxygen species (ROS) and expressions of cytochrome P4502E1 and nuclear factor κB1, tumor necrosis factor-α, and transforming growth factor-ß(1) in hepatocytes were significantly increased in ethanol group. The fibrotic markers α-smooth muscle actin and α(1)(I) collagen and activity of cytotoxicity marker caspase-3 were significantly increased and AA content was significantly reduced in hepatocytes of alcohol-treated guinea pigs. But the AA and silymarin supplementation significantly reduced these changes in comparison with alcohol abstention group. AA could induce greater reduction of inflammatory and fibrotic markers in hepatocytes than silymarin. This indicates that AA is superior to silymarin in inhibiting intracellular ROS generation and thereby reducing the ethanol-induced inflammation in hepatocytes.

Boerhaavia diffusa L. attenuates angiotensin II-induced hypertrophy in H9c2 cardiac myoblast cells via modulating oxidative stress and down-regulating NF-κß and transforming growth factor ß1.

The present study evaluated the antihypertrophic potential of the ethanolic extract of Boerhaavia diffusa (BDE), a well-known edible cardiotonic plant reported in Ayurveda against angiotensin II-induced hypertrophy in H9c2 cardiac myoblast cells. Markers of hypertrophy such as cell size, protein content and the concentrations of atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP) were analysed for the confirmation of hypertrophy induction. Angiotensin II (100 nM) caused an increase in cell volume (69·26 (SD 1·21)%),protein content (48·48 (SD 1·64)%), ANP (81·90 (SD 1·22)%) and BNP (108·57 (SD 1·47)%). BDE treatment significantly reduced cell volume, protein content and the concentrations of ANP and BNP (P#0·05) in H9c2 cells. The activity of various antioxidant enzymes and the concentration of reduced glutathione, which was lowered due to hypertrophy, were increased in BDE-treated cells. The BDE treatment also reduced intracellular reactive oxygen species generation, lipid peroxidation and protein carbonyls in cells. In addition,the expression patterns of NF-kb and transforming growth factor b1 were found to be increased during hypertrophy, and their expressions were reduced on BDE treatment. In vitro chemical assays showed that BDE inhibits angiotensin-converting enzyme and xanthine oxidase in a dose-dependent manner with an estimated 50% effective concentration (EC50) value of 166·12 (SD 2·42) and 60·05 (SD 1·54) mg/ml,respectively. The overall results clearly indicate the therapeutic potential of B. diffusa against cardiac hypertrophy, in addition to its nutritional qualities.

Additive effect of alpha-tocopherol and ascorbic acid in combating ethanol-induced hepatic fibrosis.

OBJECTIVE: To investigate the efficacy of combined administration of alpha-tocopherol (AT) and ascorbic acid (AA) in reducing ethanol-induced hepatotoxicity. METHODS: Rats were maintained for 90 days and grouped as follows: I-control rats, II-ethanol, III-alpha-tocopherol, IV-ethanol+alpha-tocopherol, V-AA, VI-ethanol+ascorbic acid, VII-alpha-tocopherol+ascorbic acid, VIII-ethanol+alpha-tocopherol+ascorbic acid. At the end of the experimental period, markers of hepatic function, oxidative stress, and the expression of markers of inflammation and fibrosis were assayed. RESULTS: The markers of hepatic function, lipid peroxidation products, protein carbonyls, and the expression of nuclear factor kappa B, tumor necrosis factor alpha, transforming growth factor beta 1, cytochrome P4502E1, and collagen Type I were elevated after ethanol administration. All these parameters were reduced in the ethanol group administered AT and AA in combination. The activities of antioxidant enzymes which were reduced by ethanol administration were enhanced on combined administration of AT and AA. The reduction in hepatic fibrosis was almost 20% more in AT and AA co-administered group compared with AT and AA alone treated groups. DISCUSSION: Combined administration of fat soluble AT and water soluble AA was beneficial against ethanol-induced hepatotoxicity. This may be due to their different subcellular localizations.

Protective effect of ascorbic acid against ethanol-induced reproductive toxicity in male guinea pigs.

The present study was undertaken to elucidate the effect of ascorbic acid on alcohol-induced reproductive toxicity and also to compare it with that of abstention. A total of thirty-six male guinea pigs were divided into two groups and were maintained for 90 d as control and ethanol-treated groups (4 g/kg body weight (b.wt.)). After 90 d, ethanol administration was stopped and animals in the control group were divided into two groups and then maintained for 30 d as the control and control+ascorbic acid groups and those in the ethanol-treated group as ethanol abstention and ethanol+ascorbic acid (25 mg/100 g b. wt.) groups. Animals treated with ethanol showed a significant decline in sperm quality (P<0·001), decreased activity of steroidogenic enzymes (P<0·05) and reduced serum testosterone (P<0·05), luteinising hormone and follicle-stimulating hormone levels, decrease in the activity of testicular succinate dehydrogenase, adenosine triphosphatase, sorbitol dehydrogenase and reduction in fructose content (P<0·05). It also caused an increase in testicular malondialdehyde levels (P<0·05) and decrease in the levels of glutathione content (P<0·001) of testes. Ascorbic acid levels in testes and plasma were also reduced (P<0·001) in ethanol-fed animals. Ascorbic acid supplementation altered all these parameters and produced a better and faster recovery from alcohol-induced reproductive toxicity than abstention. The mechanism of action of ascorbic acid may be by reducing the oxidative stress and improving antioxidant status, which eventually changed the microenvironment of testes and enhanced the energy needed for motility of sperms, improved the sperm morphology and elevated the testosterone and gonadotropin levels.

Ascorbic acid supplementation causes faster restoration of reduced glutathione content in the regression of alcohol-induced hepatotoxicity in male guinea pigs.

Alcoholic liver disease is caused mainly by free radicals. Ascorbic acid (AA) and glutathione (GSH) are the major water-soluble antioxidants in the liver. The impact of AA supplementation on GSH, AA and activities of GSH-dependent enzymes in alcoholic guinea pigs was studied and was compared with alcohol abstention. Guinea pigs were administered ethanol at a dose of 4 g/kg body weight (b.wt)/day for 90 days. After 90 days, alcohol administration was stopped and one-half of the ethanol-treated animals were supplemented with AA (25 mg/100 g b.wt) for 30 days and the other half was maintained as the abstention group. There was a significant increase in the activities of alanine aminotransferase, aspartate aminotransferase, and gamma-glutamyl transpeptidase in the serum of the ethanol group. In addition, a significant decrease in the GSH content, activities of GSH peroxidase, GSH reductase, and increased activity of GSH-S-transferase were observed in the liver of the ethanol group. Histopathological analysis and triglycerides content in the liver of the ethanol group showed induction of steatosis. But AA supplementation and abstention altered the changes caused by ethanol. However, maximum protective effect was observed in the AA-supplemented group indicating the ameliorative effect of AA in the liver.

Ascorbic acid supplementation down-regulates the alcohol induced oxidative stress, hepatic stellate cell activation, cytotoxicity and mRNA levels of selected fibrotic genes in guinea pigs.

Both oxidative stress and endotoxins mediated immunological reactions play a major role in the progression of alcoholic hepatic fibrosis. Ascorbic acid has been reported to reduce alcohol-induced toxicity and ascorbic acid levels are reduced in alcoholics. Hence, we investigated the hepatoprotective action of ascorbic acid in the reversal of alcohol-induced hepatic fibrosis in male guinea pigs (n = 36), and it was compared with the animals abstenting from alcohol treatment. In comparison with the alcohol abstention group, there was a reduction in the activities of toxicity markers and levels of lipid and protein peroxidation products, expression of α-SMA, caspase-3 activity and mRNA levels of CYP2E1, TGF-ß(1), TNF-α and α(1)(I) collagen in liver of the ascorbic acid-supplemented group. The ascorbic acid content in liver was significantly reduced in the alcohol-treated guinea pigs. But it was reversed to normal level in the ascorbic acid-supplemented group. The anti-fibrotic action of ascorbic acid in the rapid regression of alcoholic liver fibrosis may be attributed to decrease in the oxidative stress, hepatic stellate cells activation, cytotoxicity and mRNA expression of fibrotic genes CYP2E1, TGF-ß(1), TNF-α and α(1) (I) collagen in hepatic tissues.

In vitro antioxidant activities of the methanol extract and its different solvent fractions obtained from the fruit pericarp of Terminalia bellerica.

Terminalia bellerica has been used as a traditional medicine in a variety of ailments including anaemia, asthma, cancer, inflammation, rheumatism and hypertension. In this study, the free radical scavenging and antioxidant activities of methanol extract (ME) and its different solvent fractions (namely hexane (HE), ethyl acetate (EA), butanol (BL) and water (WA)) of the T. bellerica fruit pericarp were evaluated and compared with standard antioxidant compounds like gallic acid (GA), catechin and ascorbic acid. Among the different fractions tested, the EA fraction exhibited higher antioxidant and radical scavenging activity against 1,1-diphenyl-2-picrylhydrazyl (DPPH), superoxide and hydroxyl radicals than the other fractions, which may be attributed to its higher phenolic and flavonoid content, since a linear relation was observed between the phenolic content and the antioxidant parameters. The HPTLC analysis of the EA fraction revealed that it mainly contains GA and ferulic acid (FA) as major phenolics, and the higher antioxidant activities of EA fraction may be due to the presence of these compounds.

Cardioprotective effects of aqueous extract of Oxalis corniculata in experimental myocardial infarction.

The present study evaluated the protective potential of aqueous extract of Oxalis corniculata (OCE) against isoproterenol (ISO) induced myocardial infarction in rats. Myocardial infarction in rats was induced by isoproterenol (200 mg/kg) at an interval of 24 h for 2 days. OCE was given to rats as pretreatment for 30 days orally using an intragastric tube. Isoproterenol caused a significant increase in the activity of cardiac injury marker enzymes like creatine phosphokinase (CPK) and lactate dehydrogenase (LDH) and increased the concentration of serum lipids. OCE pretreatment significantly reduced the concentration of CPK, LDH, serum total cholesterol, LDL cholesterol and triglycerides. OCE also reduced the activity of lipogenic enzyme, glucose-6-phosphate dehydrogenase in ISO administered rats. Oxidative stress produced by isoproterenol was significantly lowered by the administration of OCE which was evident from increased activities of antioxidant enzymes (catalase and superoxide dismutase) and reduced concentration of lipid peroxidation products (TBARS and conjugated dienes). Concentration of vitamin C, protein sulfhydryl groups and reduced glutathione (GSH) was also high in OCE pretreated rats. Histopathology of heart of ISO administered rat pretreated with OCE showed normal myocardium with very little evidence of inflammatory infiltration. Results of our in vitro findings also confirmed that OCE exhibits significant antioxidant and radical scavenging activity against DPPH, superoxide and nitric oxide radicals. These findings provided evidence that O. corniculata was found to be protecting the myocardium against ischemic insult and the protective effect could attribute to its antioxidative and antihyperlipidemic activities.