Improvement of mitochondrial function in muscle of genetically obese rats after chronic supplementation with proanthocyanidins

The aim of this study was to determine the effect of chronic dietary supplementation of a grape seed proanthocyanidin extract (GSPE) at a dose of 35 mg/kg body weight on energy metabolism and mitochondrial function in the skeletal muscle of Zucker obese rats. Three groups of 10 animals each were used: lean Fa/fa lean group (LG) rats, a control fa/fa obese group (OG) of rats, and an obese supplemented fa/fa proanthocyanidins obese group (POG) of rats, which were supplemented with a dose of 35 mg GSPE/kg of body weight/day during the 68 days of experimentation. Skeletal muscle energy metabolism was evaluated by determining enzyme activities, key metabolic gene expression, and immunoblotting of oxidative phosphorylation complexes. Mitochondrial function was analyzed by high-resolution respirometry using both a glycosidic and a lipid substrate. In muscle, chronic GSPE administration decreased citrate synthase activity, the amount of oxidative phosphorylation complexes I and II, and Nrf1 gene expression, without any effects on the mitochondrial oxidative capacity. This situation was associated with lower reactive oxygen species (ROS) generation. Additionally, GSPE administration enhanced the ability to oxidize pyruvate, and it also increased the activity of enzymes involved in oxidative phosphorylation including cytochrome c oxidase. There is strong evidence to suggest that GSPE administration stimulates mitochondrial function in skeletal muscle specifically by increasing the capacity to oxidize pyruvate and contributes to reduced muscle ROS generation in obese Zucker rats.     

Effects of proanthocyanidin preparations on hyperlipidemia and other biomarkers in mouse model of type 2 diabetes

The protective effect of proanthocyanidins from persimmon peel, using both oligomers and polymers, was investigated in a db/db type 2 diabetes model. Male db/db mice were divided into three groups: control (vehicle), polymer-, or oligomer- (10 mg/(kg body weight x day x p.o.)) administered mice. Age-matched nondiabetic m/m mice were used as a normal group. The administration of proanthocyanidins reduced hyperglycemia in db/db mice through a decline in the serum level of glucose and glycosylated protein. In addition, it had a strong effect on hyperlipidemia through lowering levels of triglyceride, total cholesterol, and nonesterified fatty acids. The protective effect against hyperglycemia and hyperlipidemia was greater in the groups administered the oligomeric rather than polymeric form. The increased oxidative stress in db/db mice was attenuated by the administration of oligomers through inhibiting the generation of reactive oxygen species and lipid peroxidation and elevating the reduced glutathione/oxidized glutathione ratio. On the other hand, polymers did not show such an effect. Moreover, expressions in the liver of sterol regulatory element binding protein (SREBP)-1 and SREBP-2 were downregulated by the administration of proanthocyanidins, especially the oligomeric form. Oligomers caused a slight elevation in the expression of peroxisome proliferator-activated receptors alpha. Furthermore, oligomeric proanthocyanidin regulated the expression of nuclear factor kappaB in db/db type 2 diabetes via the activation of inhibitor protein kappaB-alpha. It also attenuated the protein expressions of cyclooxygenase-2 and inducible nitric oxide synthase. This suggests that oligomers would act as a regulator in inflammatory reactions associated with oxidative stress in type 2 diabetes. The present study results suggest that proanthocyanidin administration, especially the oligomeric form, may improve oxidative stress via the regulation of hyperlipidemia than hyperglycemia in type 2 diabetes.     

Increase of antioxidative potential of rat plasma by oral administration of proanthocyanidin-rich extract from grape seeds.

The effect of a single oral administration of proanthocyanidins, oligomeric and polymeric polyhydroxyflavan-3-ol units, on the antioxidative potential of blood plasma was studied in rats. Proanthocyanidin-rich extract from grape seeds was administered by intragastric intubation to fasted rats at 250 mg/kg of body weight. The plasma obtained from water- or proanthocyanidin-administered rats was oxidized by incubation with copper sulfate or 2, 2'-azobis(2-amidinopropane) dihydrochloride (AAPH) at 37 degrees C, and the formation of cholesteryl ester hydroperoxides (CE-OOH) was followed. The plasma obtained from proanthocyanidin-administered rats was significantly more resistant against both copper ion-induced and AAPH-induced formation of CE-OOH than that from control rats. The lag phase in the copper ion-induced oxidation of rat plasma was remarkably increased at 15 min after administration of proanthocyanidins and reached a maximum level at 30 min. When the plasma from proanthocyanidin-administered rat was hydrolyzed by sulfatase and beta-glucuronidase following analysis by high-performance liquid chromatography with electrochemical detection, metabolites of proanthocyanidins occurred in rat plasma at 15 min after administration, three peaks of which were identified as gallic acid, (+)-catechin, and (-)-epicatechin. These results suggest that the intake of proanthocyanidins, the major polyphenols in red wine, increases the resistance of blood plasma against oxidative stress and may contribute to physiological functions of plant food including wine through their in vivo antioxidative ability.     

Glutathione peroxidase activity, TBARS, and alpha-tocopherol in meat from chickens fed different diets.

This study investigated the effect of feeding broilers with diets differing in dietary fat source (lard, sunflower oil, olive oil) and vitamin E (basal vs supplemented with 200 mg of alpha-tocopheryl acetate/kg) on meat lipid oxidative stability. The diets differed by their degree of unsaturation and included the natural antioxidant alpha-tocopherol (vitamin E). Glutathione peroxidase (GSHPx) activity was measured in raw meat and ranged from 3.62 to 8.06 nmol NADPH/min/mg protein. The enzyme activity was influenced by the degree of unsaturation of the diet. Capillary gas chromatography analyses showed that dietary alpha-tocopherol accumulated in the muscle tissue and contributed to a better oxidative stability of the raw and cooked meat. Thigh meat alpha-tocopherol levels ranged from 2.73 to 3.62 microg/g in unsupplemented chickens whereas levels from 8.69 to 13.37 microg/g were observed in the thigh meat from alpha-tocopherol supplemented animals. The inclusion of olive oil and alpha-tocopherol in the animal diet gave lower thiobarbituric acid reactive substance (TBARS) values and lower GSHPx activity. High correlations were found between the parameters studied. The results suggest that the glutathione peroxidase activity could be used as an indicator of the meat oxidative stability. A negative relationship was observed between GSHPx activity and tissue alpha-tocopherol levels, and a positive relationship was evidenced between TBARS and antioxidant enzyme activity.     

Grape Seed-Derived Procyanidins Decrease Dipeptidyl-peptidase 4 Activity and Expression

Dipeptidyl-peptidase 4 (DPP4) inhibitors are among the newest treatments against type 2 diabetes. Since some flavonoids modulate DPP4 activity, we evaluated whether grape seed-derived procyanidins (GSPEs), which are antihyperglycemic, modulate DPP4 activity and/or expression. In vitro inhibition assays showed that GSPEs inhibit pure DPP4. Chronic GSPE treatments in intestinal human cells (Caco-2) showed a decrease of DPP4 activity and gene expression. GSPE was also assayed in vivo. Intestinal but not plasmatic DPP4 activity and gene expression were decreased by GSPE in healthy and diet-induced obese animals. Healthy rats also showed glycemia improvement after oral glucose consumption but not after an intraperitoneal glucose challenge. In genetically obese rats, only DPP4 gene expression was down-regulated. Thus, procyanidin inhibition of intestinal DPP4 activity, either directly and/or via gene expression down-regulation, could be responsible for some of their effects in glucose homeostasis.     

Hydroxytyrosol-rich olive mill wastewater extract protects brain cells in vitro and ex vivo

Elevated oxidative and nitrosative stress both impair the integrity and functioning of brain tissue, especially in aging. As long-term intake of plant foods rich in antioxidant phenolics, such as extra virgin olive oil, positively modulates surrogate markers of many human pathological alterations, the interest in cheap and abundant sources of such phenolics is rapidly growing. Olive mill wastewater is particularly rich in hydroxytyrosol, an o-diphenol with powerful antioxidant, anti-inflammatory, and antithrombotic activities. Due to the deleterious effect of oxidative stress on brain cell survival, the efficacy of a hydroxytyrosol-rich extract to attenuate Fe2+- and nitric oxide (NO)-induced cytotoxicity in murine-dissociated brain cells was investigated. The addition of either Fe2+ or SNP (an NO donor) caused both a severe loss of cellular ATP and a markedly depolarized mitochondrial membrane potential. Preincubation with hydroxytyrosol significantly attenuated the cytotoxic effect of both stressors, although with different efficiencies. Mice feeding studies were performed to assess the brain bioactivity of hydroxytyrosol ex vivo. Subchronic, but not acute, administration of 100 mg of hydroxytyrosol per kilogram body weight for 12 days enhanced resistance of dissociated brain cells to oxidative stress, as shown by reduced basal and stress-induced lipid peroxidation. Also, basal mitochondrial membrane potential was moderately hyperpolarized (P < 0.05), an effect suggestive of cytoprotection. In synthesis, the ex vivo data provide the first evidence of neuroprotective effects of oral hydroxytyrosol intake. Keywords: Hydroxytyrosol; olive mill wastewater; dissociated brain cells; oxidative stress; brain; Mediterranean diet.     

Postmortem oxygen consumption by mitochondria and its effects on myoglobin form and stability

The objective of this study was to assess the morphological integrity and functional potential of mitochondria from postmortem bovine cardiac muscle and evaluate mitochondrial interactions with myoglobin (Mb) in vitro. Electron microscopy revealed that mitochondria maintained structural integrity at 2 h postmortem; prolonged storage resulted in swelling and breakage. At 2 h, 96 h, and 60 days postmortem, the mitochondrial state III oxygen consumption rate (OCR) and respiratory control ratio decreased with time at pH 7.2 and 5.6 (p < 0.05). Mitochondria isolated at 60 days did not exhibit ADP-induced transitions from state IV to state III oxygen consumption. Tissue oxygen consumption also decreased with time postmortem (p < 0.05). Mitochondrial oxygen consumption was inhibited by decreased pH in vitro (p < 0.05). In a closed system, mitochondrial respiration resulted in decreased oxygen partial pressure (pO(2)) and enhanced conversion of oxymyoglobin (OxyMb) to deoxymyoglobin (DeoMb) or metmyoglobin (MetMb). Greater mitochondrial densities caused rapid decreases in pO(2) and favored DeoMb formation at pH 7.2 in closed systems (p < 0.05); there was no effect on MetMb formation (p > 0.05). MetMb formation was inversely proportional to mitochondrial density at pH 5.6 in closed systems. Mitochondrial respiration in open systems resulted in greater MetMb and DeoMb formation at pH 5.6 and pH 7.2, respectively, vs controls (p < 0.05). The greatest MetMb formation was observed with a mitochondrial density of 0.5 mg/mL at both pH values in open systems. Mitochondrial respiration facilitated a shift in Mb form from OxyMb to DeoMb or MetMb, and this was dependent on pH, oxygen availability, and mitochondrial density.     

Metabolic fate of glucose on 3T3-L1 adipocytes treated with grape seed-derived procyanidin extract (GSPE). Comparison with the effects of insulin

In this paper we investigate the effects of a grape seed procyanidin extract (GSPE) on the metabolic fate of glucose in adipocytes. Differentiated 3T3-L1 cells were treated with 140 mg/L GSPE or 100 nM insulin for a short period (1 h, acute treatment) or for a long period (15 h, chronic treatment). 2-Deoxy-[1-(3)H]glucose uptake and [1-(14)C]glucose incorporation into cells, glycogen, and lipid were measured. We found that GSPE mimicked the anabolic effects of insulin but there were several important differences. GSPE stimulated glycogen synthesis less than insulin. After chronic exposure, GSPE induced a higher incorporation of glucose into lipid, mainly due to the increase in glucose directed to glycerol synthesis. Our main conclusions, therefore, are that GSPE has insulinomimetic properties and activates glycogen and lipid synthesis. However, the differences between the effects of GSPE and the effects of insulin indicate that GSPE uses mechanisms complementary to those of insulin signaling pathways to bring about these effects.     

Effect of soybean phospholipids on canthaxanthin lipoproteins transport, digestibility, and deposition in rainbow trout (Oncorhynchus mykiss) muscle

This study was designed to assess the effect of dietary soybean phospholipids on canthaxanthin transport by serum lipoproteins and canthaxanthin muscle deposition in trout. Three groups of 12 immature trout in triplicate with a mean body weight of 130 g were fed with three experimental diets containing (1) canthaxanthin plus lecithin plus fish oil, (2) canthaxanthin plus lecithin, and (3) canthaxanthin alone, for 12 days. The two major lipoprotein classes in rainbow trout are high-density lipoproteins, which transport principally carotenoids present in the serum, and low-density lipoproteins, which are responsible for the transport of cholesterol, both independently of the administered diet. In addition, very low density lipoproteins are responsible for triglyceride transport in serum. Nevertheless, the amount of canthaxanthin in the serum increased when carotenoid was associated with phospholipids plus fish oil. When canthaxanthin is transported by lecithin plus fish oil, the amount of phospholipids, cantaxanthin, and cholesterol deposited in muscle increased but not significantly. The highest apparent canthaxanthin digestibility coefficient was obtained when canthaxanthin was carried by lecithin plus fish oil. The administration of canthaxanthin carried by phospholipids improved its accumulation in the muscle of rainbow trout. This accumulation could be enhanced if the time of administration of canthaxanthin is increased.     

Interactions between mitochondrial lipid oxidation and oxymyoglobin oxidation and the effects of vitamin E

Off-flavor and discoloration of meat products result from lipid oxidation and myoglobin (Mb) oxidation, respectively, and these two processes appear to be interrelated. The objective of this study was to investigate their potential interaction in mitochondria and the effects of mitochondrial alpha-tocopherol concentrations on lipid oxidation and metmyoglobin (MetMb) formation in vitro. The addition of ascorbic acid and ferric chloride (AA-Fe(3+)) increased ovine and bovine mitochondrial lipid oxidation when compared with their controls (p < 0.05); MetMb formation also increased with increased lipid oxidation relative to controls (p < 0.05). Reactions containing Mb and mitochondria with greater alpha-tocopherol concentrations demonstrated less lipid oxidation and MetMb formation than mitochondria with lower alpha-tocopherol concentrations. Greater mitochondrial alpha-tocopherol concentration was also correlated with increased mitochondrial oxygen consumption in vitro and with a more pronounced effect at pH 7.2 than at pH 5.6. Relative to controls, succinate addition to bovine mitochondria resulted in increased concentrations of ubiquinol 10 and alpha-tocopherol and decreased lipid and Mb oxidation (p < 0.05). Mitochondrial lipid oxidation was closely related to MetMb formation; both processes were inhibited by alpha-tocopherol in a concentration-dependent manner.     

Protective effect of supplementation of fish oil with high n-3 polyunsaturated fatty acids against oxidative stress-induced DNA damage of rat liver in vivo

The present study was undertaken to know the effect of supplementation of fish oil with high n-3 polyunsaturated fatty acids (PUFA) on oxidative stress-induced DNA damage of rat liver in vivo. Male Wistar rats were fed a diet containing fish oil or safflower oil with high n-6 PUFA at 50 g/kg of diet and an equal amount of vitamin E at 59 mg/kg of diet for 6 weeks. Livers of rats fed fish oil were rich in n-3 PUFA, whereas those of rats fed safflower oil were rich in n-6 PUFA. Ferric nitrilotriacetate was intraperitoneally injected to induce oxidative stress. The degree of lipid peroxidation of the liver was assessed by the levels of phospholipid hydroperoxides and thiobarbituric acid-reactive substances (TBARS), and the degree of oxidative DNA damage was assessed by comet type characterization in alkaline single-cell gel electrophoresis and 8-hydroxy-2'-deoxyguanosine levels. The levels of TBARS of the livers of the fish oil diet group increased to a greater extent than those of the safflower oil diet group, whereas the levels of the hydroperoxides of the livers of both diet groups increased to a similar extent. The vitamin E level of livers of the fish oil diet group was remarkably decreased. The degree of DNA damage of both diet groups was increased, but the increased level of the fish oil diet group was remarkably lower than that of the safflower oil diet group. The above results indicate that fish oil supplementation does not enhance but appears to protect against oxidative stress-induced DNA damage and suggest that lipid peroxidation does not enhance but lowers the DNA damage.     

Effect of beta-carotene and vitamin E on oxidative stability in leg meat of broilers fed different supplemental fats

The objectives of this study were to investigate the effects of dietary fat (6% lard and sunflower and olive oil) and supplementation of alpha-tocopheryl acetate or beta-carotene on vitamin E content and lipid oxidation in raw, cooked, and chilled-stored broiler leg meat. Vitamin E increased its tissue level, reducing lipid oxidation. The oxidative stability of leg meat tended to decrease with dietary sunflower oil. Effects of beta-carotene on vitamin E levels and oxidation depended on dietary fat and its concentration in feed, decreasing vitamin E, mainly at 50 ppm. beta-Carotene at 15 ppm acted as antioxidant in fresh and cooked meat in the sunflower and olive oil diets. However, in stored meat, beta-carotene at 50 ppm increased TBARS, probably due to a decrease in vitamin E content and direct prooxidant effects per se. It is suggested that the antioxidant effect of beta-carotene requires the presence of vitamin E in tissues.     

Erythritol attenuates the diabetic oxidative stress through modulating glucose metabolism and lipid peroxidation in streptozotocin-induced diabetic rats

We investigated the effects of erythritol on rats with streptozotocin- (STZ-) induced diabetes mellitus. Oral administration of erythritol [100, 200, or 400 mg (kg body weight)(-1) day(-1) for 10 days] to rats with STZ-induced diabetes resulted in significant decreases in the glucose levels of serum, liver, and kidney. Erythritol also reduced the elevated serum 5-hydroxymethylfurfural level that is glycosylated with protein as an indicator of oxidative stress. In addition, thiobarbituric acid-reactive substance levels of serum and liver and kidney mitochondria were dose-dependently lower in the erythritol-treated groups than in the control diabetic group. Furthermore, the serum creatinine level was reduced by oral administration of erythritol in a dose-dependent manner. These results suggest that erythritol affects glucose metabolism and reduces lipid peroxidation, thereby improving the damage caused by oxidative stress involved in the pathogenesis of diabetes.     

Amla (Emblica officinalis Gaertn.) attenuates age-related renal dysfunction by oxidative stress

To investigate the effects of amla on renal dysfunction involved in oxidative stress during the aging process, we employed young (2 months old) and aged (13 months old) male rats and administered SunAmla (Taiyo Kagaku Co., Ltd., Japan) or an ethyl acetate (EtOAc) extract of amla, a polyphenol-rich fraction, at a dose of 40 or 10 mg/kg body weight/day for 100 days. The administration of SunAmla or EtOAc extract of amla reduced the elevated levels of serum creatinine and urea nitrogen in the aged rats. In addition, the tail arterial blood pressure was markedly elevated in aged control rats as compared with young rats, while the systolic blood pressure was significantly decreased by the administration of SunAmla or EtOAc extract of amla. Furthermore, the oral administration of SunAmla or EtOAc extract of amla significantly reduced thiobarbituric acid-reactive substance levels of serum, renal homogenate, and mitochondria in aged rats, suggesting that amla would ameliorate oxidative stress under aging. The increases of inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2 expression in the aorta of aging rats were also significantly suppressed by SunAmla extract or EtOAc extract of amla, respectively. Moreover, the elevated expression level of bax, a proapoptotic protein, was significantly decreased after oral administration of SunAmla or EtOAc extract of amla. However, the level of bcl-2, an antiapoptotic protein, did not show any difference among the groups. The expressions of renal nuclear factor-kappaB (NF-kappaB), inhibitory kappaB in cytoplasm, iNOS, and COX-2 protein levels were also increased with aging. However, SunAmla or EtOAc extract of amla reduced the iNOS and COX-2 expression levels by inhibiting NF-kappaB activation in the aged rats. These results indicate that amla would be a very useful antioxidant for the prevention of age-related renal disease.     

Influence of dietary fat and vitamin E supplementation on free radical production and on lipid and protein oxidation in turkey muscle extracts

The objectives of this study were to investigate the effects of dietary fat (6% soy oil or rapeseed oil or tallow) and alpha-tocopheryl acetate supplementation at two levels (30 or 200 ppm) on radical production, measured by ESR spectroscopy, and on lipid and protein oxidation in turkey muscle extracts oxidized by an enzymic system (NADPH, ADP, FeSO(4)/cytochrome P450 reductase). Two muscles were tested: pectoralis major (glycolytic) and sartorius (oxidative) muscles. Radical production measured by ESR was higher in pectoralis major muscle than in sartorius muscle, whereas lipid and protein oxidation was more important in sartorius muscle, showing the importance of the pro-/antioxidant ratio in oxidative processes in muscular cells and of the measurement methodology to appreciate the free radical production. Dietary fat had no effect on the level of ESR signals, whereas feeding of animals with soy oil induced higher oxidation of lipids. Protein oxidation was less sensitive to the nature of the dietary fat than lipid oxidation. Vitamin E supplementation significantly decreased radical production, as measured by ESR spectroscopy. Vitamin E also decreased lipid and protein oxidation, but the effect of vitamin E on protein oxidation was less pronounced than on lipid oxidation.     

Model system for testing the efficacy of antioxidants in muscle foods

The objective of this research was to study the effect of the antioxidants, delta-tocopherol, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), tertiary butylhydroquinone (TBHQ), and propyl gallate in a model system of lean muscle and canola oil and to compare the effects with those in minced herring. Two carrier solvents with different dielectric constants (epsilon), ethanol (epsilon = 24) and oil (epsilon= 2), were used. Oxidation was measured using thiobarbituric acid reactive substances (TBARS) and sensory analysis. In both the lean muscle-canola oil model system and in herring muscle, the hydrophilic antioxidants, propyl gallate and TBHQ, were more effective in providing oxidative stability than the lipophilic antioxidants, delta-tocopherol and BHT. The oxidative stability of a cod muscle-canola oil system in the presence of propyl gallate, and delta-tocopherol was not affected by the dielectric constant of the carrier solvent, while BHA was more effective as an antioxidant when added in the polar solvent ethanol.     

IGF2基因在通城猪和长白猪胚胎骨骼肌中的表达研究

IGF2基因对骨骼肌的生长发育起着非常重要的调控作用。为了研究IGF2对猪胚胎骨骼肌生长发育的调控,本文建立猪IGF2基因的SYBR-Green荧光定量PCR方法,并利用此方法分析该基因在通城猪（脂肪型）和长白猪（瘦肉型）妊娠33天、65天和90天胚胎骨骼肌中的表达规律。结果表明,建立的SYBR-Green荧光定量方法可以有效地用于IGF2的表达分析。IGF2基因在两品种中都以妊娠65天时表达水平最高,呈波浪式表达模式。有趣的是,在所研究的三个胚胎时期,IGF2的表达在通城猪中均高于长白猪,其原因值得深入研究。     

Beneficial effects of dietary fish-oil-derived monounsaturated fatty acids on metabolic syndrome risk factors and insulin resistance in mice

The aim of this study was to elucidate the effect of fish-oil-derived monounsaturated fatty acids (MUFAs) containing large amounts of C20:1 and C22:1 isomers on metabolic disorders in mice. Male C57BL/6J mice were fed a 32% lard diet (control) or a 27% lard plus 5% saury-oil-derived MUFA diet for 6 weeks. Dietary MUFA improved insulin resistance and alleviated metabolic syndrome risk factors by reducing blood glucose and lipids. These favorable changes may be attributed to an improved adipocytokine profile. MUFA ingestion resulted in favorable changes in mRNA expression of genes involved in glucose/lipid metabolism (SCD-1, CPT1a, UCPs, and CS) as well as inflammation (MAC1, MMP3, and SAA3) and alterations in fatty acid composition. Our data suggest that marine MUFA improved glucose/lipid homeostasis and hindered the development of metabolic syndrome in obese mice.