We studied non-hospitalized 30-69 y-old Japanese subjects to ascertain the influences of a 677C-T methylene-tetrahydrofolate reductase (MTHFR) genotype, nutri-tional intake and lifestyle-related factors on plasma homocysteine (Hcys) and serum folate concentrations. Hcys was higher and serum folate was lower in males than in females (p<0, 01). The Hcys concentration was higher in the VV group than in the AA and AV groups for both males and females. However, a relatively low serum folate concentration of 18±7 nmol/L was found in the entire male group as compared with 22±10 nmol/L in all females. In the female subjects, serum folate concentrations differed among MTHFR geno-types, being lowest in the VV group. In all male subjects, log folate intake per 1, 000 kcal was a significant positive predictor of log serum folate concentration (p<0.01), while in females the log vitamin C intake per standard body weight was a significant positive variable (p<0.001) predicting the log serum folate concentration. Smokers had significantly lower serum folate concentrations, regardless of dietary folate intake. High folate and vitamin C consumptions, appears to be beneficial to normal and heterozygous MTHF genotype sub-jects for maintaining serum folate concentrations. Even a 400μg daily intake of folate might be less than what is needed, especially for homozygous MTHFR subjects and smokers, to maintain an adequate serum folate concentration.

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We examined the effect of dietary conjugated linoleic acid (CLA) on liver regeneration after a partial hepatectomy (PH) in Sprague-Dawley rats. PH Was performed on rats fed a 0 or 1 wt.% CLA diet for 3 wk. Average liver weight in the CLA fed rat popula-tion was heavier than the control rat population at the time of PH and 1-d after PH. Con-versely, CLA fed raduced liver weight was significantly lower than control rats at 7-d after PH. This suggests that dietary CLA reduced liver weight gain after PH. Dietary CLA did not affect serum aspartate aminotransferase (AST) or alanine aminotransferase (ALT) activities. How-ever, CLA significantly reduced serum albumin levels at 1-d but not at 7-d after PH. 5-Promo- and 5-iododeoxyuridine incorporation into hepatocytes 1-d post PH was lower in the CLA group. In conclusion, the data suggests that dietary CLA inhibits DNA synthesis after PH, which results in hepatocyte proliferation inhibition.

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The effects of protocatechuic acid on serum cholesterol and gene expression related to cholesterol metabolism in rats were investigated. Rats were fed a cholesterol-free diet with or without 5g protocatechuic acid/kg diet for 4 wk. There were no significant dif ferences in body weight and food intake among groups through the experimental period. The liver weight in the protocatechuic acid group was significantly lower than that in the control group. The serum total cholesterol, high-density lipoprotein (HDL)-cholesterol and very low-density lipoprotein (VLDL)+intermediate density lipoprotein (IDL)+low-density lipoprotein (LDL)-cholesterol concentrations in the protocatechuic acid group were signifi-cantly lower than those in the control group through the feeding period. The hepatic cholesterol concentration in the protocatechuic acid group was significantly higher than in the control group at the end of the 4-wk feeding period. The relative hepatic LDL receptor, apo B, apo E, lecithin-cholesterol acyltransferase (LCAT) and hepatic triglyceride lipase (HTGL) mRNA levels in the protocatechuic acid group were significantly higher than those in the control group. The results of this study suggest the possibility that the increase in the hepatic LDL receptor, apo E, LCAT and HTGL guessed by these mRNAs increase in the pro-tocatechuic acid group lowers the serum total cholesterol level.

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This study was conducted to estimate the requirements of calcium (Ca), mag-nesium (Mg) and phosphorus (P) in Japanese young adults. From 1986 to 2000, 109 vol-unteers (23 males, 86 females), ranging from 18 to 28 y old, took part in mineral balance studies after written informed consent was obtained. The duration of the study periods ranged from 5 to 12 d, with 2-4 d of adaptation. Foodstuffs used in each study were selected from those commercially available. The minerals present in diet, feces, urine and sweat were measured by atomic absorption spectrophotometer (Ca, Mg) or spectrophotometer (P). The dietary intakes of Ca, Mg and P ranged from 4.83-23.58, 2.44-7.83 and 13.46-45.69mg/kg BW/d, respectively, Dietary intake (Intake) of Ca was positively correlated to apparent absorption (AA.) (r²=0.425), which was also correlated with urine excretion (Urine) (r²=0.327) and balance (Bal) (r²=0, 382). Intake of Ca was slightly but significantly corre-lated with Bal (r²=0.036, p=0.048). The mean value and upper limit of the 95% confi-dence interval for the regression equation between Intake and Bal when balance is equal to zero (Mean and upper limit) for Ca were 11.752 and 12.555mg/kg BW/d, respectively, Intake of Mg was positively correlated to A.A. (r2=0.451), which was also correlated with Urine (r²=0.486) and Bal (r²=0.349). However, Intake of Mg was not correlated with Bal. Intake of P was positively correlated with A.A. (r²=0.959), which was also correlated with both Urine (r²=0.908) and Bal (r²=0.135). Intake of P was slightly but significantly corre-lated with Bal (r2=0.103, p=0.0013). Mean and upper limits for P were 22.584 and 24.059mg/kg BW/d, respectively. Intakes of Mg and P correlated negatively with their respective A.A, rates (%) (r²=0.120 for Mg, r²=0.109 for P). However, there was not much of a correlation for Ca. Balance of Ca was positively correlated with that of Mg (r²=0.541), but not with that of P.

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The changes in body fat ratio and serum lipids induced by the ingestion of β-conglycinin were examined in 41 healthy female university student volunteers. The trend of change in body fat ratio following the ingestion of β-conglycinin differed between students with a baseline body fat ratio over 25% and those less than 25%. In the former group, the ingestion of β-conglycinin suppressed the increase in body fat ratio. Moreover the six sub-jects who had a high total cholesterol level (5.72 mmol/L or higher) tended to have reduced levels of serum triglyceride, free fatty acid, total cholesterol and lipoprotein (a) after the ingestion of β-conglycinin, although those levels did not change significantly. The number of subjects was only six, therefore it was inferred that significant changes were not observed. Thus, ingestion of soybean β-conglycinin suppressed the increase in body fat ratio in indi-viduals with a high baseline body fat ratio and reduced relatively high serum levels of lipids. Those results suggest that if soybean β-conglycinin is ingested continuously (5g daily), it will be effective in keeping body fat ratio and serum lipid levels normal and eliminating excessive lipids from the body

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Six polyphenolic compounds were isolated from ethylacetate extract second-ary to 80% ethanol extraction of defatted safflower seeds. They were categorized into three types: lignans, flavones and serotonin derivatives. Female Sprague-Dawley rats weighing 163.4±6.3g were ovariectomized (Ovx) and fed either ethylacetate extract at a level of 1% (w/w) or three types of safflower polyphenolic compounds at a level of 200 mg/leg in a diet containing 0.5% (w/w) cholesterol for four wk. The sham and Ovx control groups were fed the same diet without safflower components. Plasma GOT and GPT levels did not differ among the six experimental groups. The plasma levels of total cholesterol were reduced in the four safflower groups by 20-30% as compared to the Ovx control. The plasma level of HDL-cholesterol was higher in the Ovx+ethylacetate extract group or appeared to be in the three Ovx+safflower polyphenolic groups than in the Ovx control. The level of plasma triglyceride was also significantly lower in the Ovx+lignan group than in the Ovx control. The liver level of cholesterol was significantly reduced in the Ovx+ethylacetate extract group. Fecal excretion of cholesterol increased by the safflower lignans and flavones, whereas that of bile acid was not significantly changed by the safflower polyphenols. Matairesinol and acacetin isolated from safflower seeds reduced the cholesterol content in cultured HepG2 cells at a concentration of 0.01-0.1μM and all three safflower polyphenolics decreased tri-glyceride content at the concentration of 0.1μM. These results suggest that safflower polyphenols have the effect of improving blood lipid status via increasing HDL-cholesterol formation and cholesterol excretion without significant uterotropic action in estrogen-deficient animals.

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We have already reported that instant coffee powder (ICP) and ICP-loade rat sera could suppress proliferation and invasion of rat ascites hepatoma cell line of AH109A in vitro. In this report, we examined the mechanisms for suppression of tumor cell prolifer-ation and invasion y ICP, and the effect of ICP on in vivo tumor growth, metastasis and abnormal lipoprotein profiles in hepatoma-bearing rats. ICP, when directly added to the cuh ture media, induced cell cycle arrest (elongation of S phase) at a lower concentration (0.3mg/mL) and apoptosis at a higher concentration (0.6-1.2mg/mL). ICP and ICP-loade rat sera showed reactive oxygen species (OS)-scavenging property and canceled the enhance-ment of invasive activity of hepatoma cells induced by OS in vitro. These results suggest that ICP suppresses the proliferation by inducing cell cycle arrest and apoptosis, and the invasion by scavenging ROS and that ICP could retain these properties after their gas-trointestinal absorption. The hepatoma-bearing rats were fed with a 20% casein diet (20C) or 20C supplemented with 0.1% ICP for 14 d. Dietary ICP significantly reduced solid tumor growth and tended to reduce hepatoma metastases to lung and lymphatic nodes, suggesting that ICP could suppress tumor cell proliferation and invasion in vivo. In addition, dietary ICP significantly increased serum high density lipoprotein (HDL)-cholesterol and tended to reduce very low-density and low-density lipoprotein (VLDL+LDL)-cholesterol, resulting in amelioration of abnormal lipoprotein profiles occurred in hepatoma-bearing rats. In conclu-sion, ICP has the ability to induce cell cycle arrest and apoptosis in hepatoma cells and to suppress tumor cell invasion by reducing oxidative stresses in vitro, and it could also exhibit these effects in vivo, leading to the inhibition of tumor growth and metastases.

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Changes in protein conformation and proteolysis in chick myotubes in response to the induction of oxidative stress by H₂O₂ treatment were studied. Myotubes were treated for 1 h with H₂O₂. After this treatment, the H₂O₂ was removed and the cells were cultured in serum-free medium for 6 and 24 h. Protein carbonyl content, as an index of protein modification, was increased at 6 and 24 h after H₂O₂ treatment. N^T-methylhistidine release, as an index of myofibrillar proteolysis, was also increased at 6 and 24 h after H₂O₂ treatment. Calpain and cathepsin (B+L and D) activities were increased at 24 but not 6 h after H₂O₂ treatment. Proteasome activity was increased at 6 and 24 h after H₂O₂ treatment. These results indicate that oxidative stress increased proteasome activity and caused an increase in myofibrillar proteolysis during short-term incubation, whereas it increased calpain, proteasome and cathepsin activities during long-term incubation, finally resulting in an increase of myofibrillar proteolysis in chick myotubes.

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We have already reported that indigestible fructooligosaccharides (FOS) increased calcium absorption in rat large intestines and that calbindin-D9k (CaBP), which is an intestine-specific calcium-binding protein, is involved in that increasing effect. In this study, not only the CaBP gene, every gene that changed expression profiles as the result of FOS feeding was identified by cDNA expression arrays. Sprague-Dawley male rats were fed an experimental diet containing 10% FOS for 10 d. To compare gene expression with rats fed a control diet, total mRNA was extracted from the colorectum and analyzed using a Rat cDNA Expression Arrays filter. This arrays filter contains probes of 588 genes, and 195 of them showed detectable changes in their expression by FOS feeding. There were six genes that increased their expression more than twice that of the control. Among them, genes related to the induction of cell growth such as Map kinase 1 and Max were included. Expres-sions that decreased to less than half were observed in 20 genes, such as somatostatin and prohibitin, which prohibit cell growth. These results are consistent with the other observa-tion that FOS increases cell growth in the colorectum. This approach has revealed that cDNA array technology is an effective tool for nutritional sciences that involve the regulation of a large number of genes, especially for molecular mechanisms of regulation, by nutri-tional constituents.

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The indispensable branched-chain amino acid leucine acts as a key regulator of mRNA translation by modulating the phosphorylation of proteins that represent impor-tant control points in translation initiation, including the translational repressor, eukaryotic initiation factor (eIF) 4E-binding protein 1 (4E-BP1) and ribosomal protein S6 kinase (S6K1). In the current study, we compared the effects of L- and D-enantiomers of leucine on the phosphorylation of 4E-BP1 and S6K1. We also assessed whether leucine itself or its metabolite, α-ketoisocaproate (α-KIC), mediates the effects of leucine. Food-deprived (18h) rats were orally administered 135mg/100g body weight L-leucine, D-leucine or α-KIC and were sacrificed after 1h. L-Leucine administration had an obvious stimulatory effect on the phosphorylation of 4E-BP 1 and S6K1 in both skeletal muscle and liver while D-leucine was much less effective, indicating that the effect of leucine is stereospecific. Oral administration of α-KIC mimicked the stimulatory effect of L-leucine in skeletal muscle. In contrast to skel-etal muscle, provision of α-KIC was significantly less effective than L-leucine in the liver. The results showing that the efficacy of L-leucine and α-KIC in stimulating phosphorylation of S6K1 and 4E-BP 1 is equivalent in skeletal muscle, may be explained by the conversion of a-KIC to L-leucine.

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Increased levels or overexpression of ornithine decarboxylase (ODC), a rate-limiting enzyme in polyamine biosynthesis pathway, is characteristic of tumor cells. Similarly, prostaglandins (PGs) appear to be important in the pathogenesis of cancer because these affect mitogenesis, cellular adhesion, immune surveillance and apoptosis. Cancers form much more PGs than the original tissue from which they have arisen. This study has revealed that pretreatment of mice with resveratrol at a dose of 2.5mg/kg body weight for two weeks blocked the N-nitrosodiethylamine(NDEA)-induced cytosolic ODC levels in the liver and lungs. The blockage was pronounced in hepatic tissue compared to pulmonary tissue. Resveratrol feeding caused a significant reduction in microsomal cyclooxygenase (COX) activities in the liver and lungs, while the dosage of NDEA (200mg/kg body weight) induced COX activity 24 h after its administration. In any case, resveratrol pretreatment turned out to effectively block the induction of COX activity in the lungs by NDEA.

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Antioxidant activity of L-ascorbic acid (AsA) against oxidative modification of apolipoprotein (apo) E in human very-low-density lipoprotein (VLDL) Was investigated. The VLDL oxidation induced by peroxyl radicals and peroxynitrite led to lipid peroxidation and oxidative modification of apoE. The binding activity of apoE to heparin was decreased by the oxidative modification. AsA (200, 500, and 1, 000μM) inhibited both the lipid peroxidation and the oxidative modification of apoE. These results suggest that AsA prevents the biologi-cal function of apoE in VLDL from the oxidation by free radicals.

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