AMPK activation by AICAR reduces diet induced fatty liver in C57BL/6 mice.

Dysregulation of 5'-adenosine monophosphate-activated protein kinase (AMPK) occurs in metabolic disorders including non-alcoholic fatty liver disease (NAFLD) which makes it a molecular target for treatment. An AMPK activator, 5-aminoimidazole-4-carboxamide-1-ß-D-ribofuranoside (AICAR) alleviates NAFLD in experimental rats, however the specific mechanism remains to be explored. We aimed to study the effect of AICAR on lipid levels, oxidant-antioxidant balance, AMPK and mTOR activation and FOXO3 gene expression in liver of mice model. Fatty liver was induced in two groups of C57BL/6 mice (groups 2 and 3) by providing a high fat high fructose diet (HFFD) for 10 weeks while groups 1 and 4 animals were fed normal pellet. For the last two weeks, groups 3 and 4 were administered AICAR (150 mg/kg bw/day, i.p.) while groups 1 and 2 were administered saline. AICAR decreased fatty liver, decreased glucose and insulin in circulation, prevented the accumulation of triglycerides and collagen and ameliorated oxidative stress in HFFD fed mice. At the molecular level, AICAR upregulated FOXO3 and p-AMPK expression and reduced p-mTOR expression. AMPK activation may involve FOXO3 in protection against NAFLD. The role of AMPK, mTOR and FOXO3 crosstalk in NAFLD needs to be characterised in future.

Effect of protocatechuic acid on lipid profile and DNA damage in D-galactosamine-induced hepatotoxic rats.

BACKGROUND: Our aim in this study is to investigate the effect of protocatechuic acid (PCA) on lipid profile and DNA damage in D-galactosamine (D-GalN)-induced hepatotoxic rats. METHODS: Hepatotoxicity was induced by a single intraperitoneal dose of D-GalN in male Wistar rats. The activities of hepatic markers and levels of kidney function markers were determined. The plasma and tissue lipid levels were estimated. DNA damage was determined by COMET assay. Histopathological examination was also performed using portions of the liver and kidney tissues. RESULTS: D-GalN-induced hepatotoxic rats showed increased in the activities of hepatic marker enzymes such as aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), and γ-glutamyl transpeptidase (GGT) in serum. The levels of kidney function markers such as urea, uric acid, and creatinine increased in serum. Levels of lipid profile such as total cholesterol (TC), triglycerides (TG), free fatty acid (FFA), and phospholipids (PLs) in the plasma and tissues (liver and kidney) were significantly increased in D-GalN-induced rats. In plasma, levels of very low density lipoprotein cholesterol (VLDL-C) and low-density lipoprotein cholesterol (LDL-C) significantly increased, whereas high-density lipoprotein cholesterol (HDL-C) level decreased in D-GalN-induced rats. Furthermore, D-GalN-induced rats showed increased percentage of tail DNA and tail length and decreased percentage of head DNA. Oral administration of PCA (100 mg/ kg BW) for 20 days improved these levels when compared to D-GalN-induced rats. These biochemical changes were reflected on the attenuation and the structural alteration of the liver and kidney integrity. CONCLUSIONS: The results of the study suggest that PCA has a potent hepatoprotective activity that may be linked to its antihyperlipidemic effect.

Rosmarinic acid inhibits DMH-induced cell proliferation in experimental rats.

BACKGROUND: Colon cancer is one of the most common cancers in both men and women. The present study is an effort to unravel the anticarcinogenic effects of rosmarinic acid (RA) in 1,2-dimethylhydrazine (DMH)-induced rat colon carcinogenesis. Administration of DMH induces multiple tumors in the rat colon, which mimics human colon cancer. METHODS: Male Wistar rats were divided into six groups and fed a high-fat diet. Group 1 served as control, group 2 rats were given RA [5 mg/kg body weight (b.w.)] orally every day for a total period of 30 weeks, and groups 3-6 were given weekly injections of DMH (20 mg/kg b.w. subcutaneous) once a week in the groin for the first 15 weeks. In addition to DMH, groups 4-6 received RA at a dose of 5 mg/kg b.w. during the initiation and postinitiation stages, and also throughout the entire study period. Colon tissues were examined histologically; further, the extent of oxidative stress was assessed by measuring lipid peroxidation and antioxidant levels in the colonic mucosa of rats. RESULTS: Macroscopic and microscopic tumors were identified in all the groups that received DMH. The results revealed that supplementation with RA significantly inhibited the tumor formation and tumor multiplicity in DMH-treated rats. RA supplementation to DMH-administered rats significantly reduced the cell proliferation markers, namely, argyrophilic nucleolar organizing regions as well as proliferative cell nuclear antigen labeling index. In addition, RA supplementation reduces the expressions of tumor necrosis factor-α, interlukin-6, and cyclooxygenase-2, and modulates the expression of p65. CONCLUSIONS: The above findings clearly underline the chemopreventive efficacy of RA against DMH-induced colon carcinogenesis.

Renoprotective and blood pressure-lowering effect of dietary soy protein via protein kinase C beta II inhibition in a rat model of metabolic syndrome.

We studied whether substitution of soy protein for casein can improve insulin sensitivity, lower blood pressure (BP), and inhibit protein kinase C betaII (PKCbetaII) activation in kidney in an acquired model of metabolic syndrome. Adult male rats were fed 4 different diets: (i) starch (60%) and casein (20%) (CCD), (ii) fructose (60%) and casein (20%) (FCD), (iii) fructose (60%) and soy protein (20%) (FSD), and (iv) starch (60%) and soy protein (20%) (CSD). Renal function parameters, BP, pressor mechanisms, PKCbetaII expression, oxidative stress, and renal histology were evaluated after 60 days. FCD rats displayed insulin resistance and significant changes in body weight, kidney weight, urine volume, plasma and urine electrolytes accompanied by significant changes in renal function parameters compared with CCD rats. Elevated BP, plasma angiotensin-converting enzyme (ACE) activity, renal oxidative stress, and reduced nitrite (NO) and kallikrein activity were observed. Western blot analysis revealed enhanced renal expression of membrane-associated PKCbetaII in the FCD group. Histology showed fatty infiltration and thickening of glomeruli while urinary protein profile revealed a 5-fold increase in albumin. Substitution of soy protein for casein improved insulin sensitivity, lowered BP and PKCbetaII activation and restored renal function. Antioxidant action, inhibitory effect on ACE and PKCbetaII activation, and increased availability of kinins and NO could be contributing mechanisms for the benefits of dietary soy protein.

Investigation of antioxidant, anti-inflammatory and DNA-protective properties of eugenol in thioacetamide-induced liver injury in rats.

The present study investigated the preventive effect of eugenol, a naturally occurring food flavouring agent on thioacetamide (TA)-induced hepatic injury in rats. Adult male Wistar rats of body weight 150-180 g were used for the study. Eugenol (10.7 mg/kg b.w./day) was administered to rats by oral intubation for 15 days. TA was administered (300 mg/kg b.w., i.p.) for the last 2 days at 24h interval and the rats were sacrificed on the 16th day. Markers of liver injury (aspartate transaminase, alanine transaminase, alkaline phosphatase, gamma-glutamyl transferase and bilirubin), inflammation (myeloperoxidase, tumor necrosis factor-alpha and interleukin-6), oxidative stress (lipid peroxidation indices, protein carbonyl and antioxidant status) and cytochrome P4502E1 activity were assessed. Expression of cyclooxygenase-2 (COX-2) and the extent of DNA damage were analyzed using immunoblotting and comet assay, respectively. Liver injury and collagen accumulation were assessed using histological studies by hematoxylin and eosin and Masson trichrome staining. Rats exposed to TA alone showed increased activities of hepatocellular enzymes in plasma, lipid peroxidation indices, inflammatory markers and pro-inflammatory cytokines and decreased antioxidant status in circulation and liver. Hepatic injury and necrosis were also evidenced by histology. Eugenol pretreatment prevented liver injury by decreasing CYP2E1 activity, lipid peroxidation indices, protein oxidation and inflammatory markers and by improving the antioxidant status. Single-cell gel electrophoresis revealed that eugenol pretreatment prevented DNA strand break induced by TA. Increased expression of COX-2 gene induced by TA was also abolished by eugenol. These findings suggest that eugenol curtails the toxic effects of TA in liver.

Effect of genistein, a soy isoflavone, on whole body insulin sensitivity and renal damage induced by a high-fructose diet.

The study evaluates the effect of genistein, a soy isoflavone, on insulin sensitivity and renal functional and structural injury in rats rendered insulin-resistant by feeding on a high-fructose diet for 60 days. Fructose-fed animals (60 g /100 g) displayed insulin resistance as indicated by the measures of insulin sensitivity [insulin sensitivity index (ISI(0,120)), quantitative insulin check index (QUICKI), and homeostatic model assessment (HOMA)]. Alterations in body weight, kidney weight, urine volume, plasma, and urine electrolytes accompanied by significant increases in plasma and urinary levels of urea, uric acid, creatinine, total protein, and albumin were observed in fructose-fed rats. Oxidative stress in kidney was noted by an elevation in lipid peroxides and a decline in glutathione (GSH). Insulin sensitivity and renal function were improved in fructose-fed rats administered genistein. Histological changes such as fatty infiltration and thickening of glomeruli observed in fructose-fed rats were also ameliorated when genistein was co-administered. The study shows that genistein improves insulin sensitivity and kidney function in a dietary model of insulin resistance. We suggest that genistein may have benefits for patients suffering from kidney disease associated with insulin resistance.

Protective effect of Umbelliferone on membranous fatty acid composition in streptozotocin-induced diabetic rats.

Umbelliferone (UMB), a natural antioxidant, is benzopyrone in nature, and it is present in the fruits of golden apple and bitter orange. Earlier we evaluated and reported the effect of Umbelliferone on antidiabetic, antioxidant and antihyperlipidemic properties, and this study was designed to evaluate the effect of Umbelliferone on membrane fatty acid composition and histopathology of liver and kidney of control and streptozotocin (STZ) diabetic rats. Male albino Wistar rats (180-200 g) were made diabetic by an intraperitonial administration of STZ (40 mg/kg). The control and diabetic rats were treated with Umbelliferone and glibenclamide dissolved in 10% dimethyl sulfoxide for 45 days. Diabetic rats had decreased insulin and increased glucose, and increased levels of thiobarbituric acid reactive substances, lipid hydroperoxides and conjugated dienes. The levels of palmitic, stearic and oleic acids increased and the levels of linolenic and arachidonic acids decreased in diabetic rats as compared with control rats. Thus, the saturated fatty acids and monounsaturated fatty acids increased and the polyunsaturated fatty acids decreased in diabetic rats. Diabetic rats had decreased liver weight and increased activities of alanine transaminase and aspartate transaminase; increased kidney weight and urine albumin, and decreased levels of urea, uric acid and creatinine in the urine. Histopathological studies of liver and kidney in diabetic rats showed fatty changes surrounding portal triad; enlargement of lining cells of tubules, fatty infiltration, large area of hemorrhage and lymphocyte infiltration. Treatment with Umbelliferone and glibenclamide reversed these changes to near normalcy. Our results showed that Umbelliferone has a protective effect on membrane fatty acid composition of liver and kidney as supported by antioxidant and antihyperlipidemic effects of Umbelliferone reported earlier as evidenced by improved histopathological changes, hepatic and nephritic markers, indicating recovery from the risk of diabetic complications.

Taurine prevents ethanol-induced alterations in lipids and ATPases in rat tissues.

The study investigates the cytoprotective effect of taurine on ethanol-induced alterations in lipids and enzymes involved in ion-transport in rat tissues. Male albino rats were divided into 4 groups (n = 8) and maintained for 28 days as follows: control group, ethanol (6 g/kg/day) group, ethanol plus taurine (10 g/kg diet/day) group and control plus taurine group. Ethanol administration caused significant increases in the lipids in plasma and tissues, such as liver, kidney and brain, together with reductions in the activities of ATPases in tissues as compared to control animals. Histological studies revealed lipid accumulation and inflammatory cell infiltrates in the liver and kidney while edematous changes were observed in the brain. Simultaneous administration of taurine along with alcohol attenuated the rise in lipid levels and normalized ATPase activities. Histological changes were alleviated on treatment with taurine. The study suggests a bioprotective effect of taurine in ethanol intoxication.

Comparative effects of curcumin and an analogue of curcumin in carbon tetrachloride-induced hepatotoxicity in rats.

We have evaluated the comparative effect of curcumin (diferuloyl methane) and its analogue [bis-1,7-(2-hydroxyphenyl)-hepta-1,6-diene-3,5-dione] (BDMC-A) on carbon tetrachloride-induced hepatotoxicity in rats. Administration of carbon tetrachloride (3 ml/kg/week) for three months significantly (P<0.05) increased the levels of marker enzymes such as aspartate transaminase (AST), alkaline phosphatase (ALP) and gamma-glutamyl transferase (GGT). The levels of plasma thiobarbituric acid reactive substances (TBARS) and lipid hydroperoxides were also significantly (P<0.05) increased. We have observed a significant (P<0.05) decrease in the levels of plasma reduced glutathione (GSH), vitamin C and vitamin E. There was a significant (P<0.05) increase in the levels of TBARS and hydroperoxides in liver and kidney and a significant (P<0.05) decrease in the activities of enzymic antioxidants- superoxide dismutase (SOD), catalase and GSH peroxidase along with GSH in CCl(4)-treated rats. Oral administration of curcumin and BDMC-A to CCl(4)-induced rats for a period of three months significantly (P<0.05) decreased the levels of marker enzymes, plasma TBARS and hydroperoxides and increased the levels of plasma and tissue antioxidants. Histopathological studies of liver also showed protective effect of curcumin and BDMC-A. We have observed thickening of blood vessels and microvesicular fatty changes around the portal triad in CCl(4)-treated rat liver. Treatment with curcumin showed only mild sinusoidal dilatation while with BDMC-A there was only mild portal inflammation. The effect exerted by BDMC-A was found to be more promising than curcumin.

Glycine modulates hepatic lipid accumulation in alcohol-induced liver injury.

We studied the effect of administering glycine, a non-essential amino acid, on serum and tissue lipids in experimental hepatotoxic Wistar rats. All the rats were fed standard pellet diet. Hepatotoxicity was induced by administering ethanol (7.9 g kg(-1)) for 30 days by intragastric intubation. Control rats were given isocaloric glucose solution. Glycine was subsequently administered at a dose of 0.6 g kg(-1) every day by intragastric intubation for the next 30 days. Average body weight gain at the end of the total experimental period of 60 days was significantly lower in rats supplemented with alcohol, but improved on glycine treatment. Feeding alcohol significantly elevated the levels of cholesterol, phospholipids, free fatty acids and triglycerides in the serum, liver and brain as compared with those of the control rats. Subsequent glycine supplementation to alcohol-fed rats significantly lowered the serum and tissue lipid levels to near those of the control rats. Microscopic examination of alcohol-treated rat liver showed inflammatory cell infiltrates and fatty changes, which were alleviated on treatment with glycine. Alcohol-treated rat brain demonstrated edema, which was significantly lowered on treatment with glycine. In conclusion, this study shows that oral administration of glycine to alcohol-supplemented rats markedly reduced the accumulation of cholesterol, phospholipids, free fatty acids and triglycerides in the circulation, liver and brain, which was associated with a reversal of steatosis in the liver and edema in the brain.

Effect of Cassia auriculata leaf extract on lipids in rats with alcoholic liver injury.

We studied the effect of administering Cassia auriculata leaf extract to rats with experimentally induced liver damage. Hepatotoxicity was induced by administering 9.875 g/kg bodyweight ethanol for 30 days by intragastric intubation. C. auriculata leaf extract was administered at a dose of 250 mg/kg bodyweight daily in one group and 500 mg/kg bodyweight daily in another group of alcohol-treated rats. All rats were fed with standard pellets. The control rats were also given isocaloric glucose solution. The average bodyweight gain was significantly lower in alcohol-treated rats, but improved on supplementation with C. auriculata leaf extract. Alcohol supplementation significantly elevated the cholesterol, phospholipid and triglyceride concentration in the liver, brain, kidney and intestine, as compared with those of the normal control rats. Treatment with C. auriculata leaf extract and alcohol significantly lowered the tissue lipid levels to almost normal levels. Microscopic examination of alcohol-treated rat liver showed inflammatory cell infiltrates and fatty changes, which were reversed on treatment with C. auriculata leaf extract. Similarly, alcohol-treated rat brain demonstrated spongiosis, which was markedly reduced on treatment with C. auriculata. In conclusion, this study shows that treatment with C. auriculata leaf extract has a lipid-lowering effect in rats with experimentally induced, alcohol-related liver damage. This is associated with a reversal of steatosis in the liver and of spongiosis in the brain. The mechanism of C. auriculata leaf extract lipid-lowering potential is unclear.