Yukio Furuichi

Tryptophans 286 and 288 in the C-terminal region of protein B23.1 are important for its nucleolar localization.

Nucleolar protein B23 can shuttle between the nucleolus and cytoplasm. However, the mechanism involved in the protein moving and staying in the nucleolus is not fully understood. To identify the nucleolar localization signal sequence of protein B23, we examined the subnuclear location of B23.1 mutant proteins fused with green fluorescent protein in HeLa cells. The results suggested that the two C-terminal tryptophan residues (Trp-286 and Trp-288) of protein B23.1 were important in this phenomenon.

Suppressive Effect of a Hot Water Extract of Adzuki Beans (Vigna angularis) on Hyperglycemia after Sucrose Loading in Mice and Diabetic Rats

A hot water extract obtained by boiling adzuki beans (Vigna angularis) to produce bean paste for Japanese cake showed inhibitory activity against alpha-glucosidase, alpha-amylase, maltase, sucrase, and isomaltase after HP-20 column chromatography. The IC50 values for each hydrolylase were 0.78 mg/ml (α-amylase), 2.45 mg/ml (maltase), 5.37 mg/ml (sucrase), and 1.75 mg/ml (isomaltase). The active fraction showed potential hypoglycemic activity in both normal mice and streptozotocin (STZ)-induced diabetic rats after an oral administration of sucrose, but did not show any effect on the blood glucose concentration after glucose administration, suggesting that the active fraction suppressed the postprandial blood glucose level by inhibiting α-glucosidase and α-amylase, irrespective of the endogenous blood insulin level.

Potential Ability of Hot Water Adzuki (Vigna angularis) Extracts to Inhibit the Adhesion, Invasion, and Metastasis of Murine B16 Melanoma Cells

The 40% ethanol eluent of the fraction of hot-water extract from adzuki beans (EtEx.40) adsorbed onto DIAION HP-20 resin has many biological activities, for example, antioxidant, antitumorigenesis, and intestinal α-glucosidase suppressing activities. This study examined the inhibitory effect of EtEx.40 on experimental lung metastasis and the invasion of B16-BL6 melanoma cells. EtEx.40 was found significantly to reduce the number of tumor colonies. It also inhibited the adhesion and migration of B16-BL6 melanoma cells into extracellular matrix components and their invasion into reconstituted basement membrane (matrigel) without affecting cell proliferation in vitro. These in vivo data suggest that EtEx.40 possesses a strong antimetastatic ability, which might be a lead compound in functional food development.

Lowering serum cholesterol level by feeding a 40% ethanol-eluted fraction from HP-20 resin treated with hot water extract of adzuki beans (Vigna angularis) to rats fed a high-fat cholesterol diet.

OBJECTIVE Hot water extract of adzuki beans (Vigna angularis) was subjected to HP-20 resin chromatography. The fraction eluted from the column using 40% ethanol (EtEx.40) was investigated by its effect on serum lipids in rats fed a high-fat cholesterol and/or cholesterol-free high-fat diet. METHODS The rats were divided into 4 groups. Groups 1 and 2 were fed a high-fat cholesterol diet with or without 3.5% EtEx.40 for 2 wk. Group 3s and 4 were fed a high-fat cholesterol-free diet with or without 3.5% EtEx.40 for 2 wk. RESULTS In the high-fat-cholesterol diet groups, there was no significant difference in food intake in the experimental diet group when compared with the control group. Serum total cholesterol level was significantly decreased in the rats fed the EtEx.40 diet, but there was no difference in fecal excretion of cholesterol and bile acid between the two dietary groups. Conversely, in the high-fat cholesterol-free diet groups, ingestion of EtEx.40 reduced serum triacylglycerol concentration. CONCLUSION Ingestion of EtEx.40 suppressed serum cholesterol level in rats fed the high-fat cholesterol and serum triacylglycerol level in rats fed the high-fat cholesterol-free diet. These mechanisms did not become clear in this experiment.

Effects of dietary alpha- or gamma-linolenic acid on levels and fatty acid compositions of serum and hepatic lipids, and activity and mRNA abundance of 3-hydroxy-3-methylglutaryl CoA reductase in rats

The effects of diets containing equal amounts of alpha (alpha)- or gamma (gamma)-linolenic acid on lipid metabolism were compared in rats. Four groups of male Wistar rats were given the diets containing 20% perilla/corn mixed oil or borage oil in the absence (PO- and BO-diets, respectively) or presence (CPO- and CBO-diets) of cholesterol for 20 days. The PO-diet yielded lower serum cholesterol than the BO-diet, although the difference was not observed between the CPO and CBO groups. The PO and CPO groups showed lower high-density lipoprotein cholesterol than the BO and CBO groups, respectively. A similar tendency was observed in serum phospholipids. The CPO-diet gave markedly lower hepatic triglycerides than the CBO-diet. The activity of hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase was much lower on the PO-diet than on the BO-diet. mRNA abundance of HMG-CoA reductase was lower in rats on the PO-diet than on the BO-diet, though there was no significant difference between the CPO and CBO groups. The present results indicate that alpha-linolenic acid exhibits a larger hypocholesterolemic effect than gamma-linolenic acid, and it may be displayed mainly through the repression of the activity and mRNA expression of HMG-CoA reductase.

Hot-water extracts from adzuki beans (Vigna angularis) stimulate not only melanogenesis in cultured mouse B16 melanoma cells but also pigmentation of hair color in C3H mice.

A hot-water extract of adzuki was obtained by boiling beans of adzuki (Vigna angularis). This hot-water extract was fractionated using HP-20 column chromatography. Its distilled water fraction (WEx) was found to stimulate tyrosinase activity in cultured mouse B16 melanoma cells and hair color pigmentation in C3H mice. At concentrations of 1–3 mg/ml, WEx stimulated melanogenesis without inhibiting cell growth. During this effect, WEx activated tyrosinase-inducing activity in the cells, but did not activate tyrosinase, which exists at an intracellular level. In this study, WEx increased cyclic adenosine-3′,5′-monophospate (cAMP) content in the cells and protein kinase A (PKA) activity, and stimulated translocation of cytosolic protein kinase C (PKC) to the membrane-bound PKC. These results suggest that the addition of WEx activates the adenylcyclase and protein kinase pathways and, as a result, stimulates melanogenesis. WEx was found to have pigmentation activity on hair color in C3H mice. It might be useful in anti-graying, protecting human skin from irradiation.

Comparative effects of short- and long-term feeding of safflower oil and perilla oil on lipid metabolism in rats

Diets high in linoleic acid (20% safflower oil contained 77.3% linoleic acid, SO-diet) and alpha-linolenic acid (20% perilla oil contained 58.4% alpha-linolenic acid, PO-diet) were fed to rats for 3, 7, 20, and 50 days, and effects of the diets on lipid metabolism were compared. Levels of serum total cholesterol and phospholipids in the rats fed the PO-diet were markedly lower than those fed the SO-diet after the seventh day. In serum and hepatic phosphatidylcholine and phosphatidylethanolamine, the proportion of n-3 fatty acids showed a greater increase in the PO group than in the SO group in the respective feeding-term. At the third and seventh days after the commencement of feeding the experimental diets, expressions of hepatic 3-hydroxy-3-methylglutaryl coenzyme A reductase mRNA were significantly higher in the SO group than those in the PO group, although the difference was not observed in the longer term. There were no significant differences in the LDL receptor mRNA levels between the two groups through the experimental term, except 3-days feeding. These results indicate that alpha-linolenic acid has a more potent serum cholesterol-lowering ability than linoleic acid both in short and long feeding-terms.

Comparative effects of safflower oil and perilla oil on serum and hepatic lipid levels, fatty acid compositions of serum and hepatic phospholipids, and hepatic mRNA expressions of 3-hydroxy-3-methylglutaryl CoA reductase, LDL receptor, and cholesterol 7alpha-hydroxylase in young and adult rats

Male Wistar rats, 4 and 33 weeks of age, were fed the diets containing safflower oil (SO-diet, 77.3% linoleic acid) or perilla oil (PO-diet, 58.4% α-linolenic acid) for 7 days. Serum total cholesterol was lower on the PO-diet in both ages. On the other hand, hepatic cholesterol and phospholipids were significantly higher in the PO group than in the SO group of the adult rats. The PO group showed significantly lower 20:4 n-6 but higher 18:2 n-6 in hepatic phosphatidylcholine compared with the SO group in both ages. Hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase mRNA was significantly lower on the PO-diet than on the SO-diet irrespective of age. The present results show that α-linolenic acid has a higher hypocholesterolemic ability than linoleic acid in rats irrespective of age and these fatty acids behaved differently in affecting hepatic mRNA expressions of HMG-CoA reductase, LDL receptor, and cholesterol 7α-hydroxylase.

Hot Water Extract of Adzuki (Vigna angularis) Suppresses Antigen-Stimulated Degranulation in Rat Basophilic Leukemia RBL-2H3 Cells and Passive Cutaneous Anaphylaxis Reaction in Mice

The hot water extract of adzuki (HWEA), which is produced as a byproduct in the adzuki bean boiling process, has anti‐tumor, antioxidative, and anti‐diabetic activities. In this study, we fractionated HWEA to 4 fractions using stepwise gradient column chromatography with water and ethanol, and demonstrated the effects of each fraction on antigen (Ag)‐stimulated degranulation in rat basophilic leukemia RBL‐2H3 cells. The 40% ethanol eluate extract (EtEx.40) showed the strongest inhibition level of these fractions. To reveal the inhibitory mechanisms underlying degranulation by EtEx.40, we investigated intracellular reactive oxygen species (ROS) production, intracellular free Ca2+ concentration ([Ca2+]i), and early intracellular signaling pathways. Treatment with EtEx.40 markedly inactivated Lyn following Ag stimulation, resulting in the suppressions of intracellular elevation of [Ca2+]i and production of ROS. To identify the active compound in EtEx.40, we isolated 7 flavonoids from EtEx.40 and calculated their inhibition levels on Ag‐stimulated degranulation. These flavonoids inhibited degranulation by about 25–60%. We further examined the in vivo effects of HWEA or EtEx.40 using a passive cutaneous anaphylaxis (PCA) reaction. Both extracts strongly suppressed the PCA reaction. These findings suggest that HWEA and/or EtEx.40 are beneficial for alleviating type I allergic symptoms. Copyright

Regulation of the differentiation of osteoblasts and osteoclasts by a hot-water extract of adzuki beans (Vigna angularis).

Osteoporosis is a global public health problem thought to be caused by an imbalance in bone metabolism. We examined in this study the 40% ethanol fraction of HP-20 resin in combination with a hot-water adzuki extract (EtEx.40) for its effect on osteoblast and osteoclast differentiation. EtEx.40-treated murine preosteoblast MC3T3-E1 cells exhibited significantly elevated alkaline phosphatase activity and mineralization. EtEx.40 facilitated osteoblast differentiation by up-regulating such osteoblast differentiation-related molecules as runt-related transcription factor 2, distal-less homeobox 5, and osterix via p38 mitogen-activated protein kinase. EtEx.40 also suppressed the formation of large tartrate-resistant acid phosphatase-positive multinucleated cells in RAW264.7 cells that had been stimulated with the receptor activator of the nuclear factor κB ligand/macrophage colony-stimulating factor. EtEx.40 significantly inhibited NF-κB activation, thus reducing the expression of such downstream molecules as c-Fos and NFATc1. Our findings suggest that EtEx.40 could be used to maintain bone mass. Graphical Abstract We prepared a 40% ethanol fraction of HP-20 resin treated with a hot water extract of adzuki (EtEx.40). EtEx.40 regulates osteoblast and osteoclast differentiation.