Inhibitory effects of extracts from Eucalyptus gunnii on α-synuclein amyloid fibrils.

Amyloid fibril formation is associated with various amyloidoses, including neurodegenerative diseases such as Alzheimer's and Parkinson's diseases. Despite the numerous studies on the inhibition of amyloid formation, the prevention and treatment of a majority of amyloid-related disorders are still challenging. In this study, we investigated the effects of various plant extracts on amyloid formation of α-synuclein. We found that the extracts from Eucalyptus gunnii are able to inhibit amyloid formation, and to disaggregate preformed fibrils, in vitro. The extract itself did not lead cell damage. In the extract, miquelianin, which is a glycosylated form of quercetin and has been detected in the plasma and the brain, was identified and assessed to have a moderate inhibitory activity, compared to the effects of ellagic acid and quercetin, which are strong inhibitors for amyloid formation. The properties of miquelianin provide insights into the mechanisms controlling the assembly of α-synuclein in the brain.

Perilla Pomace, a By-product of Oil Extraction, Is Rich in Nutrients and Can Favorably Modulate Lipid Metabolism in Sprague-Dawley Rats.

Perilla pomace, a by-product of oil extraction, is rich in nutrients, such as proteins, but it has not been used for purposes other than livestock feeding. The aim of this study was to determine how perilla pomace modulates glucose and lipid metabolism in Sprague-Dawley rats. Dried perilla pomace was added to diet at a concentration of 16%. One experimental group was administered perilla oil equivalent to that in the perilla pomace. After four weeks, the animals were euthanized, and biochemical parameters were measured. Two experiments were conducted using a low-fat (7% by weight) and a high-fat (21% by weight) diet. Regardless of the level of fat in the diets, no differences in food intake were found among the groups. In the low-fat diet-fed rats (Experiment 1), epididymal adipose tissue weight was slightly, but not significantly, lower in perilla pomace-fed rats than in those fed the control diet. Hepatic triglyceride and cholesterol levels were significantly reduced by perilla pomace compared to those in the control group. Serum lipid profiles (triglycerides and cholesterol) were similar to those in the liver, without statistically significant differences. Perilla pomace significantly diminished hepatic fatty acid synthase (FAS) activity. In high-fat diet-fed rats (Experiment 2), pomace did not significantly lower epididymal adipose tissue weight. Hepatic cholesterol levels were lower in rats on the perilla oil than in control rats. The activity of hepatic enzymes involved in fat oxidation was significantly higher in rats fed the perilla pomace than in those fed the control diet. Collectively, these results show that perilla pomace favorably modulates fat metabolism, and the specific effects depend on the fat content in the diet.

Perilla pomace obtained from four different varieties have different levels and types of polyphenols and anti-allergic activity.

Perilla frutescens (L.) Britton var. frutescens (egoma in Japan) is a traditional oilseed that has several varieties with different photoperiod responses. Although egoma pomace, industrial waste produced during oil extraction, is a rich source of macro- and micro-nutrients such as protein, fiber, minerals, and polyphenols, it has not yet been used for purposes other than livestock feeding. To find out a better use of perilla pomace and its function, we selected four varieties of egoma originating from different regions with different photoperiod responses: two varieties were from Japan, which are broadly cultivated for oilseed and are highly sensitive to light and temperature. The other two varieties from Nepal, which are tolerant to low light and low temperature. Rosmarinic acid-3-O-glucoside, rosmarinic acid, and apigenin-7-O-glucoside were detected as the main polyphenolic constituents in every variety, while apigenin and luteolin were present only in perilla pomace from Japan. In IgE-sensitized RBL-2H3 cells, polyphenols derived from two varieties of Japan suppressed degranulation of mast cells, but those derived from the two varieties of Nepal did not, indicating that apigenin and luteolin may be in part responsible for the anti-allergic response. In addition, it was found that proteins involved in the degranulation signaling pathway, such as PLCγ2, Syk, and Akt, were less phosphorylated in cells treated with the egoma pomace extracts of Japanese origin. Taken together, pomace from egoma varieties derived from different regions may differently modulate allergic response in part due to the difference in polyphenol composition and may be applied to develop nutraceuticals and functional foods fortified with anti-allergic properties.

The Contents of Polyphenols in Perilla frutescens (L.) Britton var. frutescens (Egoma) Leaves are Determined by Vegetative Stage, Spatial Leaf Position, and Timing of Harvesting during the Day.

The leaf of Perilla frutescens (L.) Britton var. frutescens (egoma) is a rich source of polyphenolic compounds, including rosmarinic acid. However, there is still a lack of detailed information concerning the content of phenolic compounds in these leaves. Since some flavonoids were found as a conjugated form, leaves were used untreated or hydrolyzed using ß-glucuronidase for analysis. Enzymatic hydrolysis method successfully identified some polyphenols, which have not been reported before. Scutellarin, a flavone glucuronide with a molecular mass similar to that of luteolin 7-O-glucuronide, was present in egoma leaves. Scutellarin was the second most abundant polyphenolic compound, after rosmarinic acid. Egoma leaves at the top of the plant contained a higher amount of rosmarinic acid and scutellarin compared to that in the leaves below. The difference in plant growth stage also influenced the rosmarinic acid and scutellarin contents, while the time of harvesting during the day did rosmarinic acid contents only. This is the first time that scutellarin, a traditional Chinese medicine, widely used for the treatment of cerebrovascular disease, was quantitatively determined in egoma leaves. The present study may help adding value to egoma leaves, developing dietary supplements, functional foods, and cosmetics.

Inhibitory Effects of Green Asparagus Extract, Especially Phospholipids, on Allergic Responses in Vitro and in Vivo.

Asparagus (Asparagus officinalis L.) is one of the widely consumed vegetables. To investigate the mechanism underlying the anti-allergic responses of asparagus, we extracted different fractions from asparagus and measured their inhibitory effects on ß-hexosaminidase release in RBL-2H3 cells in vitro and an atopic dermatitis NC/Nga mouse model in vivo. The lipid fractions from asparagus were extracted with 50% ethanol, separated using chloroform by liquid-liquid phase separation, and fractionated by solid-phase extraction. Among them, acetone fraction (rich in glycolipid) and MeOH fraction (rich in phospholipid) markedly inhibited ß-hexosaminidase release from RBL-2H3 cells. In NC/Nga mice treated with picryl chloride, atopic dermatitis was alleviated following exposure to the 50% EtOH extract, acetone fraction, and methanol fraction. The inhibitory effects of asparagus fractions in vivo were supported by the significant decrease in serum immunoglobulin E (IgE) levels. The phospholipid fractions showed significantly better inhibitory effects, and phosphatidic acid from this fraction showed the best inhibitory effect on ß-hexosaminidase release. In mice challenged with ovalbumin (OVA), oral administration of asparagus extract and its fractions decreased the OVA-specific IgE level and total IgE, indicating that these effects may be partly mediated through the downregulation of antigen-specific IgE production. Taken together, the present study shows for the first time that asparagus extract and its lipid fractions could potentially mitigate allergic reactions by decreasing degranulation in granulocytes. Our study provides useful information to develop nutraceuticals and functional foods fortified with asparagus.

Control Mechanisms of the Tumor Suppressor PDCD4: Expression and Functions.

PDCD4 is a novel tumor suppressor to show multi-functions inhibiting cell growth, tumor invasion, metastasis, and inducing apoptosis. PDCD4 protein binds to the translation initiation factor eIF4A, some transcription factors, and many other factors and modulates the function of the binding partners. PDCD4 downregulation stimulates and PDCD4 upregulation inhibits the TPA-induced transformation of cells. However, PDCD4 gene mutations have not been found in tumor cells but gene expression was post transcriptionally downregulated by micro environmental factors such as growth factors and interleukins. In this review, we focus on the suppression mechanisms of PDCD4 protein that is induced by the tumor promotors EGF and TPA, and in the inflammatory conditions. PDCD4-protein is phosphorylated at 2 serines in the SCFßTRCP ubiquitin ligase binding sequences via EGF and/or TPA induced signaling pathway, ubiquitinated, by the ubiquitin ligase and degraded in the proteasome system. The PDCD4 protein synthesis is inhibited by microRNAs including miR21.

Koji glycosylceramide commonly contained in Japanese traditional fermented foods alters cholesterol metabolism in obese mice.

Koji, which is manufactured by proliferating non-pathogenic fungus Aspergillus oryzae on steamed rice, is the base for Japanese traditional fermented foods. We have revealed that koji and related Japanese fermented foods and drinks such as amazake, shio-koji, unfiltered sake and miso contain abundant glycosylceramide. Here, we report that feeding of koji glycosylceramide to obese mice alters the cholesterol metabolism . Liver cholesterol was significantly decreased in obese mice fed with koji glycosylceramide. We hypothesized that their liver cholesterol was decreased because it was converted to bile acids. Consistent with the hypothesis, many bile acids were increased in the cecum and feces of obese mice fed with koji glycosylceramide. Expressions of CYP7A1 and ABCG8 involved in the metabolism of cholesterol were significantly increased in the liver of mice fed with koji glycosylceramide. Therefore, it was considered that koji glycosylceramide affects the cholesterol metabolism in obese mice.

Erratum to: Japanese traditional dietary fungus koji Aspergillus oryzae functions as a prebiotic for Blautia coccoides through glycosylceramide: Japanese dietary fungus koji is a new prebiotic.

[This corrects the article DOI: 10.1186/s40064-016-2950-6.].

Japanese traditional dietary fungus koji Aspergillus oryzae functions as a prebiotic for Blautia coccoides through glycosylceramide: Japanese dietary fungus koji is a new prebiotic.

BACKGROUND: The Japanese traditional cuisine, Washoku, considered to be responsible for increased longevity among the Japanese, comprises various foods fermented with the non-pathogenic fungus Aspergillus oryzae (koji). We have recently revealed that koji contains an abundant amount of glycosylceramide. Intestinal microbes have significant effect on health. However, the effects of koji glycosylceramide on intestinal microbes have not been studied. MATERIALS AND METHODS: Glycosylceramide was extracted and purified from koji. C57BL/6N mice were fed a diet containing 1 % purified koji glycosylceramide for 1 week. Nutritional parameters and faecal lipid constituents were analyzed. The intestinal microbial flora of mice on this diet was investigated. RESULTS: Ingested koji glycosylceramide was neither digested by intestinal enzymes nor was it detected in the faeces, suggesting that koji glycosylceramide was digested by the intestinal microbial flora. Intestinal microbial flora that digested koji glycosylceramide had an increased ratio of Blautia coccoides. Stimulation of B. coccoides growth by pure koji glycosylceramide was confirmed in vitro. CONCLUSIONS: Koji functions as a prebiotic for B. coccoides through glycosylceramide. Since there are many reports of the effects of B. coccoides on health, an increase in intestinal B. coccoides by koji glycosylceramide might be the connection between Japanese cuisine, intestinal microbial flora, and longevity.

Highly efficient NIR to NIR upconversion of ZnMoO4:Tm3+,Yb3+ phosphors and their application in biological imaging of deep tumors.

ZnMoO4:Tm3+,Yb3+,K+ nano-phosphors with intense NIR to NIR (excitation by 980 nm, emission at ∼800 nm) upconversion were synthesized by a facile hydrothermal method. The nanoparticles were of the order of 200-400 nm. The XRD patterns confirmed a single phase triclinic structure despite doping small amounts of RE3+ and alkali ions. The optimum concentration of Tm3+, Yb3+ and alkali ions were determined to be 0.1 mol%, 10 mol% and 10 mol%, respectively. Besides charge neutrality, the doped K+ ions affected the crystal field symmetry around the Tm3+ ions which increased the f-f transition probabilities of the RE3+ ions, and hence increased the UC intensities. Compared with ZnMoO4:Tm3+,Yb3+, the NIR to NIR upconversion emission intensity of 10 mol% K+ substituted ZnMoO4:Tm3+,Yb3+ nanocrystals increased by 21-fold and can be pumped by less than 1 mW laser power. The brightest ZnMoO4:Tm3+,Yb3+,K+ nano-phosphor was applied for non-invasively visualizing the tumors in nude mice and successfully detected deep tumors in the thigh muscles. So far, this is the first report of oxide based UCNPs used for in vivo NIR-to-NIR biological imaging and opens the door to the possibility of achieving improved features using non-fluoride based UCNPs.

Glucosylceramide Contained in Koji Mold-Cultured Cereal Confers Membrane and Flavor Modification and Stress Tolerance to Saccharomyces cerevisiae during Coculture Fermentation.

In nature, different microorganisms create communities through their physiochemical and metabolic interactions. Many fermenting microbes, such as yeasts, lactic acid bacteria, and acetic acid bacteria, secrete acidic substances and grow faster at acidic pH values. However, on the surface of cereals, the pH is neutral to alkaline. Therefore, in order to grow on cereals, microbes must adapt to the alkaline environment at the initial stage of colonization; such adaptations are also crucial for industrial fermentation. Here, we show that the yeast Saccharomyces cerevisiae, which is incapable of synthesizing glucosylceramide (GlcCer), adapted to alkaline conditions after exposure to GlcCer from koji cereal cultured with Aspergillus kawachii. We also show that various species of GlcCer derived from different plants and fungi similarly conferred alkali tolerance to yeast. Although exogenous ceramide also enhanced the alkali tolerance of yeast, no discernible degradation of GlcCer to ceramide was observed in the yeast culture, suggesting that exogenous GlcCer itself exerted the activity. Exogenous GlcCer also increased ethanol tolerance and modified the flavor profile of the yeast cells by altering the membrane properties. These results indicate that GlcCer from A. kawachii modifies the physiology of the yeast S. cerevisiae and demonstrate a new mechanism for cooperation between microbes in food fermentation.

Control of a tumor suppressor PDCD4: Degradation mechanisms of the protein in hepatocellular carcinoma cells.

In this study, we demonstrate that EGF inhibits the TGF-ß1-induced apoptosis of Huh7 cells. TGF-ß1 up-regulates the expression of PDCD4 causing apoptosis, by stimulating the synthesis of PDCD4 mRNA via the Smad signaling pathway. TGF-ß1 also inhibits the activation of S6 kinase 1 which phosphorylates the serine 67 residue of PDCD4 and leads to the phosphorylation of serine 71 and serine 76 in the ß-TRCP binding sequence. This phosphorylation sequence causes the protein to be degraded in the ubiquitin-proteasome system. EGF activates S6 kinase 1 via the PI3K-Akt-mTOR signaling pathway and stimulates the degradation of PDCD4. EGF also suppresses PDCD4 mRNA levels. As the mTOR inhibitor rapamycin up-regulated PDCD4 mRNA levels, the PI3K-Akt-mTOR signaling pathway may control the transcription of the PDCD4 gene as well as the degradation of the protein. TPA also inhibited the TGF-ß1-induced apoptosis of Huh7 cells, stimulating the degradation of the PDCD4-protein. Analyses using PDCD4 mutants with changes of serines 67, 71 and 76 to alanine revealed that the phosphorylation of serine 67 is not essential for the TPA-induced suppression of the protein. The mitogens could not suppress the PDCD4-mutant proteins with changes of serine 71 and/or serine 76 to alanine, however, indicating that phosphorylations at these residues are necessary for the proteasome-mediated degradation of PDCD4. The phosphor-mimic S71/D and S76/D mutants were able to be degraded in the ubiquitin-proteasome system unlike the mutants with changes of serine to alanine. The expression of S71/D mutant was suppressed with EGF but that of S76/D mutant was not indicating that at least partly the phosphorylation of both sites was mediated by different enzymes.

The role of PKC isoforms in the inhibition of NF-κB activation by vitamin K2 in human hepatocellular carcinoma cells.

Vitamin K (VK) has diverse protective effects against osteoporosis, atherosclerosis and carcinogenesis. We recently reported that menatetrenone, a VK2 analogue, suppressed nuclear factor (NF)-κB activation in human hepatoma cells. Although NF-κB is regulated by isoforms of protein kinase C (PKC), the involvement of PKCs in VK2-mediated NF-κB inhibition remains unknown. Therefore, the effects of VK2 on the activation and the kinase activity of each PKC isoform were investigated. The human hepatoma Huh7 cells were treated with PKC isoform-specific inhibitors and/or siRNAs against each PKC isoform with or without 12-O-tetradecanoylphorbol-13-acetate (TPA). VK2 inhibited the TPA-induced NF-κB activation in Huh7 cells. NF-κB activity was inhibited by the pan-PKC inhibitor Ro-31-8425, but not by the PKCα-specific inhibitor Gö6976. The knockdown of individual PKC isoforms including PKCα, δ and ɛ showed only marginal effects on the NF-κB activity. However, the knockdown of both PKCδ and PKCɛ, together with treatment with a PKCα-specific inhibitor, depressed the NF-κB activity. VK2 suppressed the PKCα kinase activity and the phosphorylation of PKCɛ after TPA treatment, but neither the activation nor the enzyme activity of PKCδ was affected. The knockdown of PKCɛ abolished the TPA-induced phosphorylation of PKD1, and the effects of PKD1 knockdown on NF-κB activation were similar to those of PKCɛ knockdown. Collectively, all of the PKCs, including α, δ and ɛ, and PKD1 are involved in the TPA-mediated activation of NF-κB. VK2 inhibited the NF-κB activation through the inhibition of PKCα and ɛ kinase activities, as well as subsequent inhibition of PKD1 activation.

Regulation of TGF-ß1-Induced Pro-Apoptotic Signaling by Growth Factor Receptors and Extracellular Matrix Receptor Integrins in the Liver.

Hepatocellular carcinoma (HCC) often arises from chronically diseased livers. Persistent liver inflammation causes the accumulation of excessive extracellular matrix (ECM) proteins and impairs the liver function, finally leading to the development of HCC. A pleiotropic cytokine, transforming growth factor (TGF)-ß1, plays critical roles throughout the process of fibrogenesis and hepatocarcinogenesis. In the liver, TGF-ß1 inhibits the proliferation of hepatocytes and stimulates the production of ECM from hepatic stellate cells (HSCs) to maintain tissue homeostasis. During disease progression, both growth factors/cytokines and the ECM alter the TGF-ß1 signals by modifying the phosphorylation of Smad proteins at their C-terminal and linker regions. TGF-ß1 stimulates the expression of integrins, cellular receptors for ECM, along with an increase in ECM accumulation. The activation of integrins by the ECM modulates the response to TGF-ß1 in hepatic cells, resulting in their resistance to TGF-ß1-induced growth suppression in hepatocytes and the sustained production of ECM proteins in activated HSCs/myofibroblasts. Both growth factor receptors and integrins modify the expression and/or functions of the downstream effectors of TGF-ß1, resulting in the escape of hepatocytes from TGF-ß1-induced apoptosis. Recent studies have revealed that the alterations of Smad phosphorylation that occur as the results of the crosstalk between TGF-ß1, growth factors and integrins could change the nature of TGF-ß1 signals from tumor suppression to promotion. Therefore, the modification of Smad phosphorylation could be an attractive target for the prevention and/or treatment of HCC.

Expression patterns of the tumor suppressor PDCD4 and correlation with ß-catenin expression in gastric cancers.

The expression patterns of PDCD4, a tumor suppressor, and ß-catenin were immunohistologically investigated in gastric carcinoma tissues. In normal gastric tissues, PDCD4 was strongly expressed in the cell nuclei, but weakly expressed in the cytoplasm. In gastric adenocarcinoma tissues, nuclear PDCD4 expression was decreased, while cytoplasmic PDCD4 expression was unchanged or somewhat increased. In gastric signet ring cell carcinoma tissues, PDCD4 expression patterns were different from the expression patterns of the adenocarcinoma tissues, and PDCD4 was localized in the nuclei of the carcinoma cells as a belt in the middle of the epithelial layer. The nuclear localization of PDCD4 in the adenocarcinoma tissues was correlated with the membrane localization of ß-catenin, the activation of which stimulates invasion of colon cancer cells. PDCD4 expression was correlated with ß-catenin expression in gastric carcinoma cell lines, but not with E-cadherin, as the binding partner in the cell membrane.

Vitamin K2 augments 5-fluorouracil-induced growth inhibition of human hepatocellular carcinoma cells by inhibiting NF-κB activation.

Although 5-fluorouracil (5-FU) is one of the most commonly used chemotherapeutic agents in various cancer including hepatocellular carcinoma (HCC), chemoresistance has precluded single use of 5-FU in clinical settings. Since menatetrenone, an analogue of vitamin K2 (VK2), inhibits growth of cancer cells including HCC cells in vitro and in vivo, we examined VK2 modulation of HCC cell response to 5-FU. VK2 pretreatment dose-dependently enhanced growth-inhibition by 5-FU through a G1 cell cycle arrest. VK2 inhibited 5-FU-induced NF-κB activation and cyclin D1 expression. Therefore, combination of VK2 and 5-FU might represent a new therapeutic strategy for patients with HCC.

Regulation of tumor suppressor PDCD4 by novel protein kinase C isoforms.

Transforming growth factor-beta1 (TGF-beta1) induces apoptosis in normal hepatocytes and hepatoma cells. PDCD4 is involved in TGF-beta1-induced apoptosis via the Smad pathway. The tumor promoter 12-O-tetradecanoylphorbor-13-acetate (TPA), a protein kinase C stimulator, inhibits TGF-beta1-induced apoptosis. However, the mechanisms of TPA action on PDCD4 expression remain to be elucidated. Therefore. the regulatory mechanism of PDCD4 expression by PKC was investigated. The treatment of the human hepatoma cell line, Huh7 with TPA suppressed PDCD4 protein expression and TGF-beta1 failed to increase the PDCD4 protein expression. PKC inhibitors Ro-31-8425 or bisindolylmaleimide-1-hydrocholoride (pan-PKC inhibitors) and rottlerin (PKCdelta inhibitor), but not Go6976 (PKCalpha inhibitor), enhanced the induction of PDCD4 protein by TGF-beta1. Furthermore, siRNA-mediated knockdown of PKCdelta and epsilon, but not PKCalpha, augmented the TGF-beta1-stimulated PDCD4 protein expression. However, TPA or pan-PKC inhibitor did not alter the PDCD4 mRNA expression either under basal- and TGF-beta1-treated conditions. The down-regulation of PDCD4 by TPA was restored by treatment with the proteasome inhibitor MG132. These data suggest that two isoforms of PKCs are involved in the regulation of the PDCD4 protein expression related to the proteasomal degradation pathway.

Modulation of the transforming growth factor-beta1-induced Smad phosphorylation by the extracellular matrix receptor beta1-integrin.

Integrins, heterodimeric receptors for the extracellular matrix, are known to modulate transforming growth factor-beta1 (TGF-beta1)-mediated cell behavior. However, the interplay between beta1-integrin and Smad signaling, regulated by TGF-beta1, is not clearly understood. This study focuses on the alterations of the regulatory Smads (R-Smads) by TGF-beta1 in beta1-integrin-transfected HepG2 cells. The phosphorylation at the C-terminal site of R-Smads by TGF-beta1 was impaired in the beta1-integrin-transfected cells. However, the R-Smads were constitutively phosphorylated at the linker region in a MAP kinase-dependent manner. Furthermore, the expression of a mutant Smad3, that lacks the phosphorylation sites in the linker region, restored the TGF-beta1-induced Smad transcriptional activity. These results suggest that beta1-integrin impairs the TGF-beta1-mediated signals through the altered phosphorylation of the R-Smads.

Inhibition of matrix metalloproteinase expression by menatetrenone, a vitamin K2 analogue.

Vitamin K2 (VK2) has been shown to have a potent anti-tumor effect against several cancer types including hepatocellular carcinoma (HCC), but the mechanisms remain to be elucidated. Matrix metalloproteinase (MMP) plays an important role in the invasion and metastasis of cancer cells, but it is not known whether VK2 regulates the expression of MMPs. Human HCC cell lines were treated with VK2 combined with 12-O-tetradecanoyl phorbol-13 acetate (TPA) and the expression of MMPs was examined by reporter gene assay, RT-PCR and Western blotting. VK2 inhibited the basal and TPA-induced expression of MMP-1, -3 and -7 at the transcriptional, mRNA and protein levels in a dose-dependent manner. VK2 also inhibited the TPA-induced activation of NF-kappaB and AP-1 activity. The inhibitors against NF-kappaB and mitogen-activated protein kinases (MAP kinase) including ERK and JNK pathways suppressed TPA-induced luciferase activity of MMP-1, -3 and -7 promoters. These data suggest that VK2 inhibits MMP expression by suppressing NF-kappaB and MAP kinase activity and might be potentially useful in the treatment of HCC.

A gene-gene interaction between ALDH2 Glu487Lys and ADH2 His47Arg polymorphisms regarding the risk of colorectal cancer in Japan.

Alcohol consumption is recognized as a potential risk factor for colorectal cancer (CRC). Genetic polymorphisms, aldehyde dehydrogenase (ALDH2) Glu487Lys and alcohol dehydrogenase 2 (ADH2) His47Arg, which have a strong impact on alcohol metabolism, are common in Japanese population but their significance for CRC carcinogenesis remains to be clarified in detail. We, therefore, conducted a matched case-control study with 257 incident CRC cases and 771 non-cancer controls at Aichi Cancer Center, including analysis of interactions between polymorphisms, drinking and folate consumption. The ADH2 Arg allele was found to be associated with increased risk, the odds ratios (ORs) being 1.35 (95% confidence interval: 1.00-1.84) and 1.93 (1.06-3.53) for the His/Arg and Arg/Arg genotypes, respectively. In contrast, no apparent links were observed with the ALDH2 genotypes. Individuals having ALDH2 Glu/Glu with ADH2 Arg+, ALDH2 Lys+ with ADH2 His/His and ALDH2 Lys+ with ADH2 Arg+ showed ORs of 0.10(0.04-0.21), 0.10 (0.06-0.19) and 1.36 (0.94-1.97), respectively, compared with ALDH2 Glu/Glu with ADH2 His/His. Statistical gene-gene interaction was significant between the two polymorphisms for the risk of CRC (P< 0.001). The impact of ALDH2 Lys+ with ADH2 Arg+ was more evident in low folate consumer (OR = 2.32, 1.19-4.55) than high folate consumer (OR 1.38, 0.80-2.38). In conclusion, while we failed to find any significant association with the ALDH2 polymorphism itself, significant interaction between ALDH2 and ADH2 polymorphism was observed. Replication in the future study is warranted.