Urinary biomarkers for monitoring disease progression in the Han:SPRD-cy rat model of autosomal-dominant polycystic kidney disease

The Han:SRPD-cy rat is a well-recognized model of human autosomal-dominant polycystic kidney disease. The disease is characterized by the development of progressive renal cysts, leading to declining renal function. Disease progression typically is monitored by measurement of plasma urea concentration. Although plasma urea may be an adequate measure of overall renal function, urinary biomarkers capable of accurately monitoring disease progression may be equally useful. The goal of this study was to assess several urinary biomarkers as potential markers of disease progression in male and female Han:SPRD-cy rats. These biomarkers were compared with changes in plasma urea concentration and morphometric changes as the disease progressed. Urinary activity of N-acetyl-β-D-glucosaminidase and concentration of α-glutathione S-transferase were measured as markers of proximal tubular dysfunction, glutathione S-transferase Yb1 as a distal tubular marker, and collagen IV as a biomarker for glomerular lesions. Urinary albumin was used as biomarker of glomerular or proximal tubular lesions. Albuminuria increased in male rats as the disease progressed, correlating with increasing plasma urea and morphologic changes. Urine concentrations of α-glutathione S-transferase decreased significantly in the male heterozygotic compared with wildtype rats in the later stages of the disease. Urinary concentrations of glutathione S-transferase Yb1 and collagen IV and activity of N-acetyl-β-D-glucosaminidase did not change during disease progression. Measurement of urinary albumin and concentrations of α-glutathione S-transferase may be useful for monitoring disease progression in the male Han:SPRD-cy rat model in future experiments.     

A dietary conjugated linoleic acid treatment that slows renal disease progression alters renal cyclooxygenase-2-derived prostanoids in the Han: SPRD-cy rat

A mixture of dietary conjugated linoleic acid (CLA) isomers reduces inflammation and mitigates disease progression in the Han:SPRD-cy rat model of chronic kidney disease. Since cyclooxygenase (COX) activities and prostanoid levels are higher in diseased kidneys in this rat, and dietary CLA can inhibit COX2 and prostanoid production in other tissues, the effects of dietary CLA were investigated. Kidney homogenates from normal and diseased Han:SPRD-cy rats were analyzed for prostanoid levels under various conditions: endogenous levels, steady-state levels (60-min incubations) and produced by COX isoforms. Thromboxane B(2) (TXB(2); TXA(2) metabolite), 6-keto-prostaglandin F(1α) (6-keto-PGF(1α); PGI(2) metabolite) and PGE(2) levels under these conditions were two- to ninefold higher in diseased kidneys. Dietary CLA resulted in ～32%-53% lower levels of prostanoids produced by total COX and COX2 activities in normal and diseased kidneys and partially mitigated alterations in COX2 protein levels associated with disease. The COX1 protein and activity were higher in renal disease, resulting in increased production of TXB(2) and 6-ketoPGF(1α), but not PGE(2). Dietary CLA had no effect on COX1, however. Disease resulted in up to twofold higher ratios of TXB(2)/6-ketoPGF(1α), TXB(2)/PGE(2) and 6-ketoPGF(1α) /PGE(2), and dietary CLA partially mitigated these increases under several conditions. Elevated levels of renal membrane associated cytosolic phospholipase A(2) in diseased kidneys also were reduced by 50% with CLA feeding. The effects of CLA feeding on COX2 protein levels and activity indicate that the beneficial effect of dietary CLA in this renal disorder is mediated in part via effects on COX2-derived prostanoids.     

Betaine administration corrects ethanol-induced defective VLDL secretion

Our previous studies, demonstrating ethanol-induced alterations in phosphatidylcholine (PC) synthesis via the phosphatidylethanolamine methyltransferase (PEMT) pathway, implicated a defect in very low-density lipoprotein (VLDL) secretion in the pathogenesis of hepatic steatosis. The objective of this study was to determine whether VLDL secretion was reduced by chronic ethanol consumption and whether betaine supplementation, that restores PEMT activity and prevents the development of alcoholic steatosis, could normalize VLDL secretion. The VLDL secretion in rats fed with control, ethanol and the betaine supplemented diets was determined using Triton WR-1339 to inhibit plasma VLDL metabolism. We observed reduced VLDL production rates in chronic alcohol-fed rats compared to control animals. Supplementation of betaine in the ethanol diet increased VLDL production rate to values significantly higher than those observed in the control diet-fed rats. To conclude, chronic ethanol consumption impairs PC generation via the PEMT pathway resulting in diminished VLDL secretion which contributes to the development of hepatic steatosis. By increasing PEMT-mediated PC generation, betaine results in increased fat export from the liver and attenuates the development of alcoholic fatty liver.     

Betaine Down Regulates Apelin Gene Expression in Cardiac and Adipose Tissues of Insulin Resistant Diabetic Rats Fed by High-Calorie Diet

Apelin is a newly discovered peptide that its serum level increases in diabetic patients with cardiovascular dysfunction. Recent studies indicate the beneficial actions of betaine in reducing the cardiovascular and metabolic complications, however data related to its effect on adipocytokine expression is limited. The aim of this study was to evaluate the effect of betaine supplementation on Apelin gene expression in cardiac muscle and adipose tissue of insulin resistance, diabetic rats fed by a high calorie diet. To induce insulin resistance rats were fed with high fat/high carbohydrate diet for five weeks and then 30 mg/kg STZ was injected intraperitoneally. After confirming of diabetes incidence (serum glucose above 7.5 mmol/l) the animals were treated with 1 % betaine in drinking water for 28 days. At days 14 and 28 after treatment, animals were euthanized and Apelin gene expression was evaluated by real time PCR and western blot in heart and adipose tissues. Serum levels of insulin, Apelin and glucose and HOMA–IR were also measured. Our results showed that feeding of rats by a high calorie diets caused insulin resistance, which was manifested by elevated plasma insulin, glucose and Apelin levels and also HOMA–IR. Apelin gene expression in heart and adipose tissues were significantly increased simultaneously with the progression of diabetes. Betaine supplementation decreased serum Apelin and down regulated Apelin expression in adipose tissue and cardiac muscle, particularly at day 28 of treatment. We concluded that betaine might improve metabolic and cardiovascular complications in diabetic patients by regulation of Apelin expression and secretion.     

Involvement of AMP-activated protein kinase in beneficial effects of betaine on high-sucrose diet-induced hepatic steatosis

Although simple steatosis was originally thought to be a pathologically inert histological change, fat accumulation in the liver may play a critical role not only in disease initiation, but also in the progression to nonalcoholic steatohepatitis and cirrhosis. Therefore, prevention of fat accumulation in the liver may be an effective therapy for multiple stages of nonalcoholic fatty liver disease (NAFLD). Promising beneficial effects of betaine supplementation on human NAFLD have been reported in some pilot clinical studies; however, data related to betaine therapy in NAFLD are limited. In this study, we examined the effects of betaine on fat accumulation in the liver induced by high-sucrose diet and evaluated mechanisms by which betaine could attenuate or prevent hepatic steatosis in this model. Male C57BL/6 mice weighing 20 +/- 0.5 g (means +/- SE) were divided into four groups (8 mice per group) and started on one of four treatments: standard diet (SD), SD+betaine, high-sucrose diet (HS), and HS + betaine. Betaine was supplemented in the drinking water at a concentration of 1% (wt/vol) (anhydrous). Long-term feeding of high-sucrose diet to mice caused significant hepatic steatosis accompanied by markedly increased lipogenic activity. Betaine significantly attenuated hepatic steatosis in this animal model, and this change was associated with increased activation of hepatic AMP-activated protein kinase (AMPK) and attenuated lipogenic capability (enzyme activities and gene expression) in the liver. Our findings are the first to suggest that betaine might serve as a therapeutic tool to attenuate hepatic steatosis by targeting the hepatic AMPK system.     

Amelioration of D-galactosamine-induced acute liver injury in rats by dietary supplementation with betaine derived from sugar beet molasses

The effects of betaine supplementation on D-galactosamine-induced liver injury were examined in terms of hepatic and serum enzyme activities and of the levels of glutathione and betaine-derived intermediates. The rats induced with liver injury showed marked increases in serum enzyme activity, but those receiving dietary supplementation of 1% betaine showed enzyme activity levels similar to a control group without liver injury. Administration of betaine also increased both hepatic and serum glutathione levels, even following D-galactosamine injection. The activity of glutathione-related enzymes was markedly decreased following injection of D-galactosamine, but remained comparable to that of the control group in rats receiving 1% betaine. The concentrations of hepatic S-adenosyl methionine and cysteine showed similar trends to that observed for hepatic glutathione levels. These results indicate that 1% betaine has a hepatoprotective effect by increasing hepatic and serum glutathione levels along with glutathione-related enzyme activities in rats.     

Overexpression of kidney phosphatidylinositol 4-kinasebeta and phospholipase C(gamma1) proteins in two rodent models of polycystic kidney disease

Our studies of renal phosphoinositide levels and metabolism in the pcy mouse with polycystic kidney disease (PKD) suggest that phosphatidylinositol kinase (PtdInsK) and phospholipase C (PLC) are elevated in this renal disorder. Therefore, the steady-state levels of select isoforms of these enzymes were examined in renal cytosolic and particulate (detergent-soluble) fractions in male and female normal and CD1-pcy/pcy (pcy) mice at 60, 120 and 180 days of age, and in male and female normal and diseased (Han:SPRD-cy) rats at 28 and 70 days of age. Disease-related increases in phosphatidylinositol 4-kinasebeta (PtdIns4Kbeta) and PLC(gamma1) levels were present in both models. PtdIns4Kbeta levels were higher by as much as 233% in pcy mice and by 95% in diseased Han:SPRD-cy rats compared to normals of the same age and gender. Steady-state levels of PLC(gamma1) were as much as 74% and 35% higher in pcy mice and diseased Han:SPRD-cy rats, respectively, compared to their controls. The consistency of these alterations in two accepted models of PKD indicates the importance of the phosphoinositide signalling pathway in the evolution of this disorder, and represents a potential site for therapeutic intervention.     

Betaine supplementation prevents fatty liver induced by a high-fat diet: effects on one-carbon metabolism

The purpose of this study was to examine the effects of betaine supplementation on the regulation of one-carbon metabolism and liver lipid accumulation induced by a high-fat diet in rats. Rats were fed one of three different liquid diets: control diet, high-fat diet and high-fat diet supplemented with betaine. The control and high-fat liquid diets contained, respectively, 35 and 71 % of energy derived from fat. Betaine supplementation involved the addition of 1 % (g/L) to the diet. After three weeks on the high-fat diet the rats had increased total liver fat concentration, liver triglycerides, liver TBARS and plasma TNF-α. The high-fat diet decreased the hepatic S-adenosylmethionine concentration and the S-adenosylmethionine/S-adenosylhomocysteine ratio compared to the control as well as altering the expression of genes involved in one-carbon metabolism. Betaine supplementation substantially increased the hepatic S-adenosylmethionine concentration (~fourfold) and prevented fatty liver and hepatic injury induced by the high-fat diet. It was accompanied by the normalization of the gene expression of BHMT, GNMT and MGAT, which code for key enzymes of one-carbon metabolism related to liver fat accumulation. In conclusion, the regulation of the expression of MGAT by betaine supplementation provides an additional and novel mechanism by which betaine supplementation regulates lipid metabolism and prevents accumulation of fat in the liver.     

Feeding flaxseed oil but not secoisolariciresinol diglucoside results in higher bone mass in healthy rats and rats with kidney disease

Flaxseed's oil and lignan, secoisolariciresinol diglucoside (SDG), are implicated in attainment of health and treatment of renal injury and osteoporosis. To test for these benefits, weanling Han:SPRD-cy rats (n=171) with or without kidney disease were randomized to diets made with either corn oil or flaxseed oil and with or without SDG for 12 weeks. In females, weight was lower with the SDG diet. In males fed flaxseed oil, lean mass was higher and fat % was lower. In both sexes, fat % was lower in diseased rats. Bone mineral content (BMC) and density were higher in rats fed flaxseed oil and lower in diseased rats, additionally; BMC was lower in SDG-supplemented females. The benefit of flaxseed oil on body composition is sex specific but the effect on bone mass is not. Lastly, reduced weight due to early rat kidney disease is not due to loss of lean body mass.     

Cysteine supplementation decreases plasma homocysteine concentration in rats fed on a low-casein diet in rats

The effect of dietary supplementation with cysteine on the plasma homocysteine concentration was investigated in rats fed on 10% casein (10C) and 30% casein (30C) diets. The 10C diet significantly increased the plasma homocysteine concentration as compared with the 30C diet. The hyperhomocysteinemia induced by the 10C diet was significantly suppressed by cysteine supplementation even at a 0.3% level, whereas cysteine did not decrease the plasma homocysteine concentration when added to the 30C diet. In contrast, 0.3% methionine supplementation of the 10C diet tended to increase the plasma homocysteine concentration. Cysteine supplementation to rats fed on the 10C diet did not alter the plasma cysteine concentration and the hepatic activities of cystathionine beta-synthase and betaine:homocysteine S-methyltransferase, whereas it significantly decreased the hepatic concentrations of S-adenosylmethionine and betaine. These results suggest that cysteine supplementation might be effective for suppressing the hyperhomocysteinemia induced by a low-protein diet.     

Betaine attenuates hepatic steatosis by reducing methylation of the MTTP promoter and elevating genomic methylation in mice fed a high-fat diet

Aberrant DNA methylation contributes to the abnormality of hepatic gene expression, one of the main factors in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). Betaine is a methyl donor and has been considered to be a lipotropic agent. However, whether betaine supplementation improves NAFLD via its effect on the DNA methylation of specific genes and the genome has not been explored. Male C57BL/6 mice were fed either a control diet or high-fat diet (HFD) supplemented with 0%, 1% and 2% betaine in water (wt/vol) for 12 weeks. Betaine supplementation ameliorated HFD-induced hepatic steatosis in a dose-dependent manner. HFD up-regulated FAS and ACOX messenger RNA (mRNA) expression and down-regulated PPARα, ApoB and MTTP mRNA expression; however, these alterations were reversed by betaine supplementation, except ApoB. MTTP mRNA expression was negatively correlated with the DNA methylation of its CpG sites at −184, −156, −63 and −60. Methylation of these CpG sites was lower in both the 1% and 2% betaine-supplemented groups than in the HFD group (averages; 25.55% and 14.33% vs. 30.13%). In addition, both 1% and 2% betaine supplementation significantly restored the methylation capacity [S-adenosylmethionine (SAM) concentration and SAM/S-adenosylhomocysteine ratios] and genomic methylation level, which had been decreased by HFD (0.37% and 0.47% vs. 0.25%). These results suggest that the regulation of aberrant DNA methylation by betaine might be a possible mechanism of the improvements in NAFLD upon betaine supplementation.     

Betaine supplementation protects against high-fructose-induced renal injury in rats

High fructose intake causes metabolic syndrome, being an increased risk of chronic kidney disease development in humans and animals. In this study, we examined the influence of betaine on high-fructose-induced renal damage involving renal inflammation, insulin resistance and lipid accumulation in rats and explored its possible mechanisms. Betaine was found to improve high-fructose-induced metabolic syndrome including hyperuricemia, dyslipidemia and insulin resistance in rats with systemic inflammation. Betaine also showed a protection against renal dysfunction and tubular injury with its restoration of the increased glucose transporter 9 and renal-specific transporter in renal brush bolder membrane and the decreased organic anion transporter 1 and adenosine-triphosphate-binding cassette transporter 2 in the renal cortex in this model. These protective effects were relevant to the anti-inflammatory action by inhibiting the production of inflammatory cytokines including interleukin (IL)-1β, IL-18, IL-6 and tumor necrosis factor-α in renal tissue of high-fructose-fed rat, being more likely to suppress renal NOD-like receptor superfamily, pyrin domain containing 3 inflammasome activation than nuclear factor κB activation. Subsequently, betaine with anti-inflammation ameliorated insulin signaling impairment by reducing the up-regulation of suppressor of cytokine signaling 3 and lipid accumulation partly by regulating peroxisome proliferator-activated receptor α/palmityltransferase 1/carnitine/organic cation transporter 2 pathway in kidney of high-fructose-fed rats. These results indicate that the inflammatory inhibition plays a pivotal role in betaine’s improvement of high-fructose-induced renal injury with insulin resistance and lipid accumulation in rats.     

Dietary betaine accumulates in the liver and intestinal tissue and stabilizes the intestinal epithelial structure in healthy and coccidia-infected broiler chicks

The aim of this experiment was to study the patterns of betaine accumulation into intestinal tissue, liver and plasma of broiler chicks with or without coccidial infection. The chicks were raised on a corn-based, low-betaine diet with or without 1000 ppm betaine supplementation and with or without intestinal microparasite (Eimeria maxima) challenge to the age of 21 days. Plasma, liver, intestinal tissue and digesta of non-challenged (NC) birds and plasma and intestinal tissue of coccidiosis challenged (CC) birds were analysed for betaine content. NC birds were also analyzed for homocysteine in plasma and S-adenosylmethionine (S-AM) in liver. The jejunal epithelium was histologically examined for the presence of coccidia and the crypt-villus ratio was measured. Dietary betaine supplementation decreased the plasma homocysteine concentration but had no effect on liver S-AM of NC birds. The data suggest that chicks on a low-betaine diet accumulate betaine into the intestinal tissue. When the diet was supplemented with betaine, betaine accumulated heavily into liver and to a lesser degree into intestinal tissue. The concentration of betaine in jejunal and ileal digesta was low suggesting that dietary betaine was mainly absorbed from the proximal small intestine. The coccidial challenge decreased the concentration of betaine in the liver, but greatly increased that in the intestinal tissue. The crypt-villus ratio was decreased by the dietary betaine supplementation in healthy and challenged chicks, suggesting that dietary betaine both protects the jejunal villi against coccidial infection and also stabilizes the mucosal structure in healthy broiler chicks. These results support our earlier findings suggesting that betaine is likely to act as an important intestinal osmolyte in broiler chicks.     

Maternal betaine supplementation attenuates glucocorticoid-induced hepatic lipid accumulation through epigenetic modification in adult offspring rats

There are lots of reports about alleviation of NAFLD by dietary supplements of betaine. However, it remains unclear whether maternal betaine supplementation can also ameliorate NAFLD in offspring. Hence, twenty pregnant rats were fed with a basal diet with or without betaine (1%), and then the female offspring rats were raised at 3 months of age followed by 3 weeks of physiological saline or dexamethasone in a dose of 0.1 mg/kg body mass every day via intraperitoneal injection. In this study, maternal betaine supplementation significantly (P<.05) reduced the increase of hepatic triglycerides concentration in dexamethasone-induced rats, which is associated with the expression of hepatic lipogenic genes (ACC1, FASN and SCD1). Moreover, the hypomethylation of lipogenic genes in dexamethasone-induced rats were reserved by prenatal betaine exposure. Furthermore, the increase of hepatic GR or SP1 content in dexamethasone-injected rats were significantly decreased (P<.05), which were in line with the binding of GR or SP1 to lipogenic genes, in betaine -exposed rats. Together, these results suggest that maternal betaine supplementation attenuates dexamethason-induced fatty liver in the female adult offspring rats, which may be attributed to DNA methylation and GR or SP1-mediated the regulation of lipogenic genes.     

The effects of chronic betaine supplementation on exercise performance, skeletal muscle oxygen saturation and associated biochemical parameters in resistance trained men

Trepanowski, JF, Farney, TM, McCarthy, CG, Schilling, BK, Craig, SA, and Bloomer, RJ. The effects of chronic betaine supplementation on exercise performance, skeletal muscle oxygen saturation, and associated biochemical parameters in resistance trained men. J Strength Cond Res 25(12): 3461-3471, 2011-We examined the effects of chronic betaine supplementation on exercise performance and associated parameters in resistance trained men. Men were randomly assigned in a double-blind manner using a crossover design to consume betaine (2.5 g of betaine mixed in 500 ml of Gatorade?) or a placebo (500 ml of Gatorade?) for 14 days, with a 21-day washout period. Before and after each treatment period, tests of lower- and upper-body muscular power and isometric force were conducted, including a test of upper-body muscular endurance (10 sets of bench press exercise to failure). Muscle tissue oxygen saturation (StO2) during the bench press protocol was measured via near infrared spectroscopy. Blood samples were collected before and after the exercise test protocol for analysis of lactate, nitrate/nitrite (NOx), and malondialdehyde (MDA). When analyzed using a repeated measures analysis of variance, no significant differences were noted between conditions for exercise performance variables (p > 0.05). However, an increase in total repetitions (p = 0.01) and total volume load (p = 0.02) in the 10-set bench press protocol was noted with betaine supplementation (paired t-tests), with values increasing approximately 6.5% from preintervention to postintervention. Although not of statistical significance (p = 0.14), postexercise blood lactate increased to a lesser extent with betaine supplementation (210%) compared with placebo administration (270%). NOx was lower postintervention as compared with preintervention (p = 0.06), and MDA was relatively unchanged. The decrease in StO2 during the bench press protocol was greater with betaine vs. placebo (p = 0.01), possibly suggesting enhanced muscle oxygen consumption. These findings indicate that betaine supplementation results in a moderate increase in total repetitions and volume load in the bench press exercise, without favorably impacting other performance measures.     

Supplementation with chromium picolinate recovers renal Cr concentration and improves carbohydrate metabolism and renal function in type 2 diabetic mice

To study the preventive effect of supplemented chromium picolinate (CrPic) on the development of diabetic nephropathy in mice, we analyzed the effects of CrPic supplementation on renal function and concentrations of serum glucose and tissue chromium (Cr). In experiment 1, male KK-Ay obese diabetic mice were fed either a control diet (control) or a diet supplemented with 2 mg/kg diet (Cr2) or 10 mg/kg diet (Cr10) of Cr for 12 wk. Cr10 significantly ameliorated hyperglycemia after a glucose load, creatinine clearance rates, and urinary microalbumin levels (p<0.05). In experiment 2, the Cr10 diet was fed to male KK-Ay obese diabetic mice and C57BL nondiabetic mice for 4 wk. The CrPic diet reduced urinary albumin excretion in the diabetic mice (p<0.05). Inductively coupled plasma-mass spectrometry analysis revealed that the renal Cr content and the recovery of renal Cr concentration after Cr supplementation were significantly lower in the diabetic mice than in the nondiabetic mice (p<0.01). These observations suggest that Cr supplementation of type 2 diabetic mice reduces the symptoms of hyperglycemia and improves the renal function by recovering renal Cr concentration.     

Betaine analogues alter homocysteine metabolism in rats

Glycine betaine supplementation lowers homocysteine levels in homocystinuria and in chronic renal failure patients through methylation catalysed by betaine-homocysteine methyltransferase (BHMT). The aim of this study was to determine the effect of glycine betaine analogues on homocysteine metabolism in Lewis rats. Glycine betaine, proline betaine, trigonelline, dimethylsulfoniopropionate (DMSP) or dimethylthetin (1.5 mmoles) was subcutaneously administered to rats fed a low betaine diet. The effect of each betaine on total plasma homocysteine and urinary and plasma betaine concentrations was monitored for 24h following administration. Baseline plasma homocysteine was 8.5 +/- micromol/l (S.E.M., n=44) and compared to controls concentrations decreased following glycine betaine (0.8+/-0.4 micromol/l, P = 0.064), DMSP (1.0+/-0.5 micromol/l, P = 0.041) and dimethylthetin (1.5 +/- 0.7micromol/l, P = 0.033) treatment, while concentrations increased following proline betaine (2.24 +/-0.7micromol/l, P = 0.002) and trigonelline (1.6 +/-0.3 micromol/l, P < 0.001) treatment. The effect of glycine betaine, DMSP and dimethylthetin on circulating homocysteine concentrations was thought to be mediated by BHMT in vivo. This hypothesis was supported by the finding that circulating glycine betaine concentrations increased following DMSP and dimethylthetin treatment. Proline betaine and trigonelline appeared to be poor BHMT substrates, being largely excreted in the urine unchanged, yet increased circulating homocysteine levels. This suggests they are inhibitors of BHMT. Urinary excretion of glycine betaine increased following treatment with all betaines, suggesting that the resorption of glycine betaine in the kidney was inhibited. The study shows that glycine betaine analogues have multiple effects on homocysteine metabolism (250).     

Alleviation of dimethylnitrosamine-induced liver injury and fibrosis by betaine supplementation in rats

Previous studies suggested that betaine intake might antagonize the induction of oxidative stress-mediated acute liver injury through regulation of the sulfur-amino acid metabolism. In this study we examined the protective effects of betaine on chronic liver injury and fibrosis induced by dimethylnitrosamine (DMN). Male rats were supplemented with betaine (1%, w/v) in drinking water from 2 weeks prior to the initiation of DMN treatment (10 mg/(kg day), i.p., 3 days/week, for 1, 2, or 4 weeks) until sacrifice. Induction of liver injury was determined by quantifying serum alanine aminotransferase, aspartate aminotransferase activities, bilirubin levels, hepatic xenobiotic-metabolizing capacity, histopathological changes and 4-hydroxyproline levels. Development of oxidative injury was estimated by malondialdehyde (MDA) levels and total oxyradical scavenging capacity (TOSC) of liver and serum toward hydroxyl, peroxyl radicals, and peroxynitrite. Progressive changes in the parameters of liver injury and fibrosis were evident in the rats challenged with DMN. Elevation of MDA levels in liver was significant before the onset of a change in any parameters determined in this study. Betaine supplementation markedly attenuated the induction of hepatotoxicity and fibrosis by DMN. Elevation of MDA and the reduction of TOSC were also depressed significantly. Development of liver injury corresponded well with the induction of oxidative stress in rats treated with DMN, both of which are inhibited effectively by betaine supplementation. It is suggested that betaine may protect liver from fibrogenesis by maintaining the cellular antioxidant capacity.     

Grape pomace reduces the severity of non-alcoholic hepatic steatosis and the development of steatohepatitis by improving insulin sensitivity and reducing ectopic fat deposition in mice

While non-alcoholic fatty liver disease (NAFLD) represents the common cause of chronic liver disease, specific therapies are currently unavailable. The wine industry produces millions of tons of residue (pomace), which contains high levels of bioactive phytochemicals. The aim of this study was to clarify the potential benefits of grape pomace for the treatment of NAFLD at different levels of severity, and to clarify the mechanism of action. C57Bl/6 mice were given high fat diet (HFD) or western diet (WD) as models of obesity and hepatic steatosis or steatohepatitis, respectively, with or without pomace supplementation (50–250 mg/day). Pomace inhibited food intake, and reduced serum leptin and body weight gain. Ectopic fat deposition was reduced, while white adipose tissue mass was preserved. In addition, pomace improved glucose tolerance and insulin sensitivity, prevented the development of adipose tissue inflammation, and reduced hepatic steatosis. Higher expression of genes involved in fatty acids transport and oxidation was observed in adipose tissue, while lipogenic genes were attenuated in the liver of pomace-treated mice. In WD-fed mice, pomace reduced the severity of hepatic steatosis and inflammation and improved blood lipid profile, but was ineffective in reversing hepatic damage of advanced NASH. In conclusion, pomace improved insulin sensitivity and reduced ectopic fat deposition, leading to a healthier metabolic profile. Pomace may hold the potential as a supplement with beneficial health outcomes for the prevention and treatment of hepatic steatosis and other obesity-related pathologies.