Effects of Rice Proteins from Two Cultivars,Koshihikari and Shunyo,on Cholesterol and Triglyceride Metabolism in Growing and Adult Rats

The effect and mechanism of two types of rice protein, one from regular japonica rice Koshihikari and another from rice cultivar Shunyo, with low glutelin and high prolamin content, on cholesterol and triglyceride metabolism were compared by feeding casein and soy protein to male Wistar strain rats 7 and 20 weeks old ad libitum for 2 weeks. The results in adult rats clearly indicated that both rice proteins had cholesterol-lowering effects in the plasma and the liver, comparable to soy protein, and the effects were accompanied with TG-lowering effects in the liver. Similar effects were also observed in growing rats when the diets were supplemented with cholesterol. The mechanism of the cholesterol-lowering effects by these rice proteins cannot be explained solely by fecal steroid excretion, but the results indicate that not only regular rice protein but also Shunyo rice protein possesses improving effects on lipid metabolism, especially in the adult period.     

Hypocholesterolemic effect of rice protein is due to regulating hepatic cholesterol metabolism in adult rats

Aging is one of major risk factors for developing hypercholesterolemia. To elucidate the cholesterol-lowering mechanism exerted by rice protein (RP), the effects on hepatic cholesterol outputs and cholesterol metabolism related enzymes were investigated in adult rats, which were fed by casein (CAS) and RP without cholesterol in diets. After 2 weeks of feeding, the significant cholesterol-lowering effect was observed in adult rats fed by RP compared to CAS. The hepatic total- and VLDL-cholesterol secretions into circulation were significantly depressed in RP group, whereas biliary outputs of bile acids and cholesterol were effectively stimulated by RP-feeding, causing an increase in fecal sterol excretion compared to CAS. As a result, the apparent cholesterol absorption was significantly inhibited by RP. RP-feeding significantly increased the activity and gene expression of cholesterol 7α-hydroxylase, whereas acyl-CoA:cholesterol acyltransferase-2 activity and gene expression were significantly decreased by RP as compared with CAS. Neither activity nor gene expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase of RP did differ from CAS in the liver. The present study demonstrates that rice protein can prevent hypercholesterolemia through modifying hepatic cholesterol metabolism under cholesterol-free dietary condition. The findings suggest that hypocholesterolemic action induced by rice protein is attributed in part to the inhibition of cholesterol absorption during the adult period.     

Effect of phytate in soy protein on the serum and liver cholesterol levels and liver fatty acid profile in rats

Dietary soy protein, in comparison with casein, generally lowers the serum cholesterol concentration in rats fed on a cholesterol-enriched diet, while mixed results were observed in rats fed on a diet free of cholesterol. Soy protein also suppresses the conversion of linoleic acid to arachidonic acid in the rat liver. The present study examines whether phytate, a minor component of a soy protein isolate, is responsible for these beneficial effects of soy protein. Weanling male rats were fed for 4 weeks on a purified diet containing a 20% level of protein (either casein (CAS), soy protein (SOY), phytate-depleted SOY (PDSOY) or phytate-replenished PDSOY (PRSOY)) and cholesterol (0 or 0.5%). The dietary protein source and phytate level only affected the serum and liver cholesterol concentrations when the animals were fed on the cholesterol-enriched diet, being significantly lower in those rats fed on the SOY and PRSOY diets than in those fed on the CAS diet, while the concentrations in the rats fed on the PDSOY diet were intermediate. When the animals were fed on the cholesterol-free diet, the ratio of (20:3n-6 + 20:4n-6)/18:2n-6 in liver phosphatidylcholine, a delta6 desaturation index, was significantly lower in the SOY diet group than in the CAS, PDSOY and PRSOY diet groups. Dietary cholesterol significantly depressed the ratio, but neither depletion nor replenishment of phytate affected the ratio. These results suggest that phytate in soy protein played a limited role in the cholesterol-lowering effect of soy protein and was not involved in the metabolism of linoleic acid.     

Antioxidant capacity responsible for a hypocholesterolemia is independent of dietary cholesterol in adult rats fed rice protein

Dietary cholesterol and aging are major risk factors to accelerate oxidation process for developing hypercholesterolemia. The major aim of this study is to elucidate the effects of rice protein on cholesterol level and oxidative stress in adult rats fed with and without cholesterol. After 2 weeks of feeding, hepatic and plasma contents of cholesterol, reduced glutathione (GSH), oxidized glutathione (GSSG), malondialdehyde (MDA) and protein carbonyl (PCO) were measured. In liver, total antioxidative capacity (T-AOC), activities of antioxidant enzymes (total superoxide dismutase, T-SOD; catalase, CAT), glutathione metabolizing enzyme activities and gene expression levels (γ-glutamylcysteine synthetase, γ-GCS; glutathione reductase, GR; glutathione peroxidase, GPx) were determined. Under cholesterol-free/enriched dietary condition, T-AOC, activities of T-SOD and CAT, glutathione metabolism related enzymes' activities and mRNA levels (γ-GCS, GR and GPx) were effectively stimulated by rice proteins as compared to caseins. Compared with caseins, rice proteins significantly increased hepatic and plasma GSH contents, whereas hepatic and plasma accumulations of MDA, PCO and GSSG were significantly reduced by rice protein-feedings. As a result, the marked reductions of cholesterol in the plasma and in the liver were observed in adult rats fed rice proteins with and without cholesterol. The present study demonstrates that the hypocholesterolemic effect of rice protein is attributable to inducing antioxidative response and depressing oxidative damage in adult rats fed cholesterol-free/enriched diets. Results suggest that the antioxidant capability involved in the hypocholesterolemic action exerted by rice protein is independent of dietary cholesterol during adult period.     

Isoflavone-poor soy protein alters the lipid metabolism of rats by SREBP-mediated down-regulation of hepatic genes

Soy intake acts hypolipidemically. Besides isoflavones, soy protein itself is suggested to influence plasma lipid concentrations. We investigated the effects of an alcohol-washed isoflavone-poor soy protein isolate on plasma and liver lipids and the hepatic expression of genes encoding proteins involved in cholesterol and fatty acid metabolism. Therefore, rats were fed diets containing 200 g/kg of either ethanol-extracted soy protein isolate or casein over 22 days. Rats fed soy protein isolate had markedly lower concentrations of liver cholesterol and lower concentrations of triglycerides in the liver and in plasma than rats fed casein (P<.05). Rats fed soy protein isolate had lower relative mRNA concentrations of sterol-regulatory element-binding protein (SREBP)-2, 3-hydroxy-3-methylglutaryl coenzyme A reductase, low-density lipoprotein receptor, cholesterol 7alpha-hydroxylase, apolipoprotein B, Delta9-desaturase and glucose-6-phosphate dehydrogenase in the liver than rats fed casein (P<.05). Hepatic mRNA concentration of SREBP-1c tended to be lower in rats fed soy protein isolate (P<.10). Hepatic mRNA concentrations of insulin-induced gene (Insig) 1 and Insig-2 and of microsomal triglyceride transfer protein, as well as plasma concentrations of free fatty acids, insulin and glucagon, were not different between the two groups. In conclusion, this study suggests that isoflavone-poor soy protein isolate affects cellular lipid homeostasis by the down-regulation of SREBPs and its target genes in the liver, which are involved in the synthesis of cholesterol and triglycerides.     

Effects of dietary protein type on oxidized cholesterol-induced alteration in age-related modulation of lipid metabolism and indices of immune function in rats

Exogenous oxidized cholesterol disturbs both lipid metabolism and immune functions. Therefore, it may perturb these modulations with ageing. Effects of the dietary protein type on oxidized cholesterol-induced modulations of age-related changes in lipid metabolism and immune function was examined using differently aged (4 weeks versus 8 months) male Sprague-Dawley rats when casein, soybean protein or milk whey protein isolate (WPI) was the dietary protein source, respectively. The rats were given one of the three proteins in diet containing 0.2% oxidized cholesterols mixture. Soybean protein, as compared with the other two proteins, significantly lowered both the serum thiobarbituric acid reactive substances value and cholesterol, whereas it elevated the ratio of high density lipoprotein-cholesterol/cholesterol in young rats, but not in adult. Moreover, soybean protein, but not casein and WPI, suppressed the elevation of Delta6 desaturation indices of phospholipids in both liver and spleen, particularly in young. On the other hand, WPI, compared to the other two proteins, inhibited the leukotriene B4 production of spleen, irrespective of age. Soybean protein reduced the ratio of CD4(+)/CD8(+) T-cells in splenic lymphocytes. Therefore, the levels of immunoglobulin (Ig)A, IgE and IgG in serum were lowered in rats given soybean protein in both age groups except for IgA in adult, although these observations were not shown in rats given other proteins. Thus, various perturbations of lipid metabolism and immune function caused by oxidized cholesterol were modified depending on the type of dietary protein. The moderation by soybean protein on the change of lipid metabolism seems to be susceptible in young rats whose homeostatic ability is immature. These observations may be exerted through both the promotion of oxidized cholesterol excretion to feces and the change of hormonal release, while WPI may suppress the disturbance of immune function by oxidized cholesterol in both ages. This alleviation may be associated with a large amount of lactoglobulin in WPI. These results thus showed a possibility that oxidized cholesterol-induced perturbations of age-related changes of lipid metabolism and immune function can be moderated by both the selection and combination of dietary protein.     

Effects of soy protein diet on the expression of adipose genes and plasma adiponectin.

Many studies have reported the cholesterol-lowering, anti-lipogenic, anti-obesity and anti-hypertensive effects of soy protein. Adipose tissue-specific plasma protein, adiponectin, has anti-atherogenic and anti-insulin-resistance properties. Here, we investigated the effects of soy protein diet on body fat composition, plasma glucose, lipid and adiponectin levels and expression of genes involved in glucose and fatty acid metabolism in obese KK-A y mice. Body weights and adipose tissue weights of mesenteric, epididymal, and brown fat were lower in mice on calorie-restricted diet containing soy protein isolate. Plasma cholesterol, triglyceride, free fatty acid, and glucose levels were also decreased by this diet. Body fat content and plasma glucose levels in mice on a soy protein isolate diet were still lower than those treated with an isocaloric casein-protein-diet. Among the genes related to glucose and fatty acid metabolism, adiponectin mRNA levels in adipose tissue and adiponectin plasma concentrations were elevated in mice on a calorie-restricted diet, although there were no significant differences between soy protein and casein protein groups. Our results indicate that that soy protein diet decreased body fat content and plasma glucose levels more effectively than isocaloric casein-protein diet in obese mice.     

Effect of the Supplementation of Sulfur Amino Acids to a Diet Containing Soy Protein Isolate on Lipid Metabolism in Rats

The effect of the supplementation of sulfur amino acids to a low casein or soy protein isolate diet on tissue lipid metabolism was investigated. Supplementation of methionine to a 8% casein diet produced a fatty liver in rats, however, supplementation of methionine to a 8.8% soy protein diet (corresponding to a 8% casein diet as to sulfur amino acids content) did not produce a fatty liver. At the level of 8% or less of soy protein in the diet, the accumulation of liver lipids and serum triglyceride was observed. An amino acid mixture simulating the composition of soy protein isolate caused significant accumulation of liver lipids, but serum triglyceride was not changed. Serum cholesterol in rats fed the soy protein diet was lower than that in rats fed the casein diet, but on feeding the amino acid mixtures simulating these protein diets, there was no difference between the two groups. The small differences between soy protein isolate and casein as to lipid metabolism might be due to the small differences in the contents of sulfur amino acids or the specific nature of the soy protein or casein. The supplemental effect of methionine and cystine was also studied. About 60% of total sulfur amino acids could be substituted by cystine for maximum growth.     

Comparison of regulative functions between dietary soy isoflavones aglycone and glucoside on lipid metabolism in rats fed cholesterol

The effects of dietary soy isoflavones aglycone and glucoside on lipid metabolism were compared in male Sprague-Dawley rats (4 weeks old) given purified diets containing 0.3% cholesterol. The rats were fed a diet supplemented with either isoflavone aglycone-rich powder (IF-A group) or isoflavone glucoside-rich powder (IF-G group) or isoflavone-free diet (control group) for 40 days. The additional level of isoflavone aglycone moiety in the diet was prepared to the same level (approximately 0.096 g/100 g: approximately 0.1% in diet). The activity of hepatic cholesterol 7alpha-hydroxylase tended to be slightly higher in the rats fed isoflavones than in those fed the isoflavone-free diet. On the other hand, the activity of hepatic Delta6 desaturase in the IF-A group was lower than that of the control group. Reflecting this effect, the Delta6 desaturation indices [(20:3n-6+20:4n-6)/18:2n-6] in liver phospholipids of the IF-A group were lower than those in the control group. Liver and serum total cholesterol levels and liver TG level were also reduced by consumption of isoflavone aglycone. Moreover, serum TG level was lowered by consumption of both isoflavones aglycone and glucoside. The level of serum total isoflavones in the IF-A group was significantly higher than that in the IF-G group. Therefore, we speculate that the absorption speed of isoflavone aglycones might be faster than that of isoflavone glucosides in rats. This study suggests that dietary soy isoflavones, particularly their aglycone form, may exert a beneficial effect on lipid metabolism in rats fed cholesterol.     

大米蛋白调控成熟期大鼠LDLR基因及蛋白表达

为了探讨大米蛋白对成熟期大鼠胆固醇代谢调控因子一低密度脂蛋白受体（low—densitv lipoprotein receptor．LDLR）的调控作用,以18周龄雄性Wistar成熟期大鼠为研究对象,应用大米蛋白及酪蛋白为食物蛋白源,饲喂无胆固醇及富含胆固醇饲料,经18日自由摄食后,测定实验鼠血浆总胆固醇、血浆高密度胆固醇水平及肝脏LDLR基因及蛋白表达水平。对照酪蛋白,大米蛋白均能显著降低大鼠血浆总胆固醇、血浆非高密度胆固醇水平及动脉粥样硬化指数,并且,显著刺激肝脏LDL基因及蛋白表达水平。实验结果表明,大米蛋白降低成熟期大鼠血浆胆固醇水平的作用功效与膳食胆固醇添加与否无关,大米蛋白降胆固醇的作用机制之一是能够有效刺激LDLR的表达,从而抑制LDL—C的转运入血。     

Mechanism for the cholesterol-lowering action of egg white protein in rats

Eggs are a popular source of dietary cholesterol, but their consumption does not necessarily result in an increased serum cholesterol concentration. We investigated the cholesterol-lowering activity of egg white protein (EWP) and its potential mechanism in rats. The consumption of EWP resulted in a decreased concentration of cholesterol in the serum, liver and intestinal mucosa. The excretion of fecal neutral sterols and bile acids was greater by rats fed with EWP than by those fed with casein. The ratio of cholesterol and bile acids in the micellar phase to those in the solid phase was lower in the intestinal contents from rats fed with EWP than from those fed with casein. These results suggest that the cholesterol-lowering activity of EWP can be attributed to lowering the cholesterol absorption by intervening in the micellar formation in the intestines.     

Effects of soybean isoflavones, probiotics, and their interactions on lipid metabolism and endocrine system in an animal model of obesity and diabetes

The effects of soybean isoflavones with or without probiotics on tissue fat deposition, plasma cholesterol, and steroid and thyroid hormones were studied in SHR/N-cp rats, an animal model of obesity, and were compared to lean phenotype. We tested the hypothesis that probiotics by promoting the conversion of isoflavone glycosides to their metabolically active aglycone form will have a synergistic effect on body fat, cholesterol metabolism, and the endocrine system. Obese and lean SHR/N-cp rats were fed AIN-93 diets containing 0.1% soy isoflavone mixture, 0.1% probiotic mixture, or both together. Different fat tissues were teased and weighed. Plasma was analyzed for cholesterol and steroid and thyroid hormones. In both phenotypes, isoflavones lowered fat deposition in several fat depots. Probiotics alone had no significant effect on fat depots. Isoflavones lowered total, LDL, and HDL cholesterol in lean rats, but in obese rats isoflavones lowered only total and LDL cholesterol. Isoflavones also lowered many of the steroid hormones involved in lipid metabolism but had no significant effect on thyroid hormones. Probiotics had no significant effect on cholesterol or hormones. Thus, our data show that soy isoflavones also lower plasma cholesterol and that this hypocholesterolemic effect appears to be due in part to the modulation of steroid hormones. Probiotics do not seem to enhance the effect of isoflavones.     

Soy protein influences insulin sensitivity and cardiovascular risk in male lean SHHF rats.

Previous investigations have demonstrated a marked effect of soy protein on multiple physiological parameters associated with the metabolic syndrome (MS). This preliminary study investigated the physiological effects of soy-based diets on cardiovascular risk in a unique rodent model that reflects early stages of MS. Briefly, lean male SHHF (+/cp) rats were randomly assigned to the following treatment groups: casein (control, C); low-isoflavone (LIS) soy protein isolate; high-isoflavone (HIS) soy protein isolate; or C+ 0.01 % rosiglitazone (CR). Rats were fed for thirty-six weeks. Liver weight, heart weight, total plasma cholesterol, fasting blood glucose were lower in soy-fed animals compared to control (p < 0.01). Body weight, kidney weight, alanine aminotransferase (ALT), fasting plasma insulin, and homeostasis model assessment (HOMA) score were also lower in LIS-fed rodents (p < 0.05) compared to casein treatment. All diet groups exhibited lower urine protein (p < 0.01) and small arteriole content (p < 0.05) compared to controls. LIS feed had a slightly more profound influence on body weight, liver metabolism, and insulin sensitivity. However, both soy diets exhibited marked improvements over a casein-based diet.     

Effects of Lactobacillus Fermented Soymilk and Soy Yogurt on Hepatic Lipid Accumulation in Rats Fed a Cholesterol-Free Diet

We examined the effects of lactic acid fermented soymilk, in which part of the soymilk was replaced with okara (soy yogurt), on plasma and hepatic lipid profiles in rats fed a cholesterol-free diet. Additionally, we investigated the effects of soy yogurt on hepatic gene expression in rats using DNA microarray analysis. Male Sprague-Dawley rats aged 5 weeks (n=5/group) were fed a control diet (AIN-93) or a test diet in which 20% of the diet was replaced by soy yogurt for 7 weeks. Soy yogurt consumption did not affect body weight or adipose tissue weight as compared with control diet. In the soy yogurt group, the liver weight and hepatic triglyceride content were significantly lower than the control group, and the level of plasma cholesterol was also lower. Furthermore, DNA microarray analysis indicated that soy yogurt ingestion down-regulated the expression of the SREBP-1 gene and enzymes related to lipogenesis in the rat liver, while expression of β-oxidation-related genes was up-regulated. These results suggest that soy yogurt is beneficial in preventing hepatic lipid accumulation in rats.     

Alimentary proteins, amino acids and cholesterolemia

Numerous data from both epidemiological and experimental origins indicate that some alimentary proteins and amino acids in supplements can modify the blood LDL cholesterol, HDL cholesterol and total cholesterol. After an initial approval of the health claim for soy protein consumption for the prevention of coronary heart disease, more recently it has been concluded from an overall analysis of literature that isolated soy protein with isoflavones only slightly decrease LDL and total cholesterol. Other plant extracts and also some proteins from animal origin have been reported to exert a lowering effect on blood cholesterol when compared with a reference protein (often casein). The underlying mechanisms are still little understood. Individual amino acids and mixture of amino acids have also been tested (mostly in animal studies) for their effects on cholesterol parameters and on cholesterol metabolism. Methionine, lysine, cystine, leucine, aspartate and glutamate have been tested individually and in combination in different models of either normo or hypercholesterolemic animals and found to be able to modify blood cholesterol and/or LDL cholesterol and/or HDL cholesterol. It is however not known if these results are relevant to human nutrition.     

Lupin protein attenuates the development of hypertension and normalises the vascular function of NaCl-loaded Goto-Kakizaki rats.

The beneficial cardiovascular effects of soy protein have been studied intensively in recent years. Another protein-rich legume is lupin, which has been shown to have similar effects to those of soy in lowering serum cholesterol levels. In this study we compared the effects of lupin and soy protein on hypertension and vascular functions in spontaneously diabetic Goto-Kakizaki (GK) rat, which develop hypertension when fed a high-salt diet. The rats were fed with a 6% NaCl diet containing either lupin or soy protein isolate (20% weight/weight) for two weeks. In the end of the study the SBP was 18.6 mmHg lower (p<0.001) in the lupin group, and 12.0 mmHg lower (p<0.01) in the soy group than in the control group. Lupin and soy treatments normalised the decreased vasocontraction observed in the NaCl-fed control group, but only lupin treatment improved the impaired endothelium-dependent vasorelaxation. The attenuation of hypertension is likely to be mediated by the corrected vascular dysfunction, whose precise mechanism and the possible clinical relevance remains to be studied further.     

Prenatal exposure to valproate induces sex-, age-, and tissue-dependent alterations of cholesterol metabolism: Potential implications on autism

Here, we investigated the protein network regulating cholesterol metabolism in the liver and brain of adolescent and adult male and female rats prenatally exposed to valproate (VPA), a well validated experimental model of autism spectrum disorders (ASD). We were aimed at studying whether prenatal VPA exposure affected the proteins involved in cholesterol homeostasis in a sex-dependent manner. To this aim the protein network of cholesterol metabolism, in term of synthesis and plasma membrane trafficking, was analyzed by western blot in the liver and different brain areas (amygdala, cerebellum, cortex, hippocampus, nucleus accumbens, and dorsal striatum) of adolescent and adult male and female rats prenatally exposed to VPA. Our results show that physiological sex-dependent differences are present both in the liver and in brain of rats. Interestingly, VPA affects specifically the brain in an age- and region-specific manner; indeed, cerebellum, cortex, hippocampus and nucleus accumbens are affected in a sex-dependent way, while this does not occur in amygdala and dorsal striatum. Overall, we demonstrate that each brain area responds differently to the same external stimulus and males and females respond in a different way, suggesting that this could be related to the diverse incidences, between the sexes, of some neurodevelopmental pathologies such as autism, which displays a 3:1 male to female ratio.     

Piperidine alkaloids from Piper retrofractum Vahl. protect against high-fat diet-induced obesity by regulating lipid metabolism and activating AMP-activated protein kinase

The fruits of Piperretrofractum Vahl. have been used for their anti-flatulent, expectorant, antitussive, antifungal, and appetizing properties in traditional medicine, and they are reported to possess gastroprotective and cholesterol-lowering properties. However, their anti-obesity activity remains unexplored. The present study was conducted to isolate the anti-obesity constituents from P. retrofractum Vahl. and evaluate their effects in high-fat diet (HFD)-induced obese mice. Piperidine alkaloids from P. retrofractum Vahl. (PRPAs), including piperine, pipernonaline, and dehydropipernonaline, were isolated as the anti-obesity constituents through a peroxisome proliferator-activated receptor δ (PPARδ) transactivation assay. The molecular mechanism was investigated in 3T3-L1 adipocytes and L6 myocytes. PRPA treatment activated AMP-activated protein kinase (AMPK) signaling and PPARδ protein and also regulated the expression of lipid metabolism-related proteins. In the animal model, oral PRPA administration (50, 100, or 300 mg/kg/day for 8 weeks) significantly reduced HFD-induced body weight gain without altering the amount of food intake. Fat pad mass was reduced in the PRPA treatment groups, as evidenced by reduced adipocyte size. In addition, elevated serum levels of total cholesterol, low-density lipoprotein cholesterol, total lipid, leptin, and lipase were suppressed by PRPA treatment. PRPA also protected against the development of nonalcoholic fatty liver by decreasing hepatic triglyceride accumulation. Consistent with the in vitro results, PRPA activated AMPK signaling and altered the expression of lipid metabolism-related proteins in liver and skeletal muscle. Taken together, these findings demonstrate that PRPAs attenuate HFD-induced obesity by activating AMPK and PPARδ, and regulate lipid metabolism, suggesting their potential anti-obesity effects.