Symbiobacterium thermophilum is a syntrophic bacterium whose growth depends on coculture with a cognate Bacillus sp. We have been studying the unique features of S. thermophilum in terms of taxonomy, ecology, genome biology, and physiology. Here we overview current knowledge of this bacterium. Although S. thermophilum shows several physiological properties of Gram-negative bacteria, 16S rRNA gene-based phylogeny indicated that it represents a distinct lineage of Gram-positive bacteria with deep branching between the clades for the high-G+C (Actinobacteria) and the low-G+C (Firmicutes) groups. Ecological study has revealed that S. thermophilum and its relatives are widely distributed in the natural environment, including soil, animal intestines and seawater. A whole genome sequencing study uncovered its unusual features, which overall indicate that this bacterium is a member of Firmicutes despite of its high G+C content (68.7%). The genome appeared to retain fully the genes for primary metabolism, except for carbonic anhydrase. We discovered that carbon dioxide is a marked inducer of the mono-growth of S. thermophilum, and speculated that this is due to a lack of carbonic anhydrase. The lines of evidence suggest that S. thermophilum requires additional conditions for full growth, including not only the supply of an unknown positive factor but also the elimination of oxygen and self-growth inhibitory substances. We conclude that the role of the cognate Bacillus is to establish a complex environment suitable for the growth of S. thermophilum, which is achieved by supplying and removing multiple factors. Understandings of this type of mutualism should provide new insight into microbial physiology as well as the issue of uncultivability.

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The constituents of cape aloe were investigated after a preliminary screening of the growth-inhibiting effect on Ehrlich ascites tumor cells (EATC) of several extracts of this plant. Ten compounds were isolated from the dichloromethane (CH₂Cl₂) extract that showed the strongest activity, and their structures were elucidated as aloe-emodin (1), p-hydroxybenzaldehyde (2), p-hydroxyacetophenone (3), pyrocatechol (4), 10-oxooctadecanoic acid (5), 10-hydroxyoctadecanoic acid (6), methyl 10-hydroxyoctadecanoate (7), 7-hydroxy-2,5-dimethylchromone (8), furoaloesone (9), and 2-acetonyl-8-(2-furoylmethyl)-7-hydroxy-5-methylchromone (10) based on MS and various NMR spectroscopic techniques. Compounds 2–7 were isolated for the first time from cape aloe. Compounds 4–7 and 10 showed a significant growth-inhibiting effect, and compound 1 exhibited a remarkable synergistic effect on compounds 8–10, which was not observed with the treatment by each compound alone on EATC. These results suggest that the strong growth-inhibiting effect of the CH₂Cl₂ extract was dependent not on one compound alone, but on the synergistic effect from the combination of compound 1 and the other compounds.

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We performed in-situ ultraviolet resonance Raman (UVRR) spectroscopy of the larval cocoon silk of the hornet, Vespa simillima xanthoptera Cameron, and compared the result with that of the silkworm, Bombix mori. The UVRR spectrum of the hornet cocoon differed markedly from that of the B. mori cocoon: peaks attributable to tyrosine (Tyr) were observed strongly, and tryptophan (Trp) peaks weakly, in the spectrum of the B. mori cocoon, whereas peaks attributable to Trp exclusively appeared in the spectrum of the hornet cocoon.

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The filamentous fungus, Lambertella corni-maris (L. corni-maris), a mycoparasite on Monilinia fructigena, produces the antibiotics, lambertellols A (1), B (2), and lambertellin (3), in a substantial amounts under acidic conditions, whereas these antibiotics were hardly detected when the fungus was cultured on a potato-sucrose (PS) medium without added acids. Our investigations also revealed that the host, M. fructigena, changed its surroundings into acidic conditions, suggesting that the acidic conditions acted as kairomones that stimulated the production of 1–3.

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An attempt was made to use a simple procedure to obtain (R)- and (S)-2-aminobutanoic acids [(R)- and (S)-1] which are non-proteinogenic α-amino acids and are useful as chiral reagents in asymmetric syntheses. Compound (RS)-1 p-toluenesulfonate [(RS)-2], which is known to exist as a conglomerate, was optically resolved by replacing crystallization with (R)- and (S)-methionine p-toluenesulfonate [(R)- and (S)-3] as optically active co-solutes. When (S)-3 was employed as the co-solute, (R)-2 was preferentially crystallized from a supersaturated solution of (RS)-2 in 1-propanol, as was (S)-2 in the presence of (R)-3. (R)- and (S)-2 recrystallized from 1-propanol were treated with triethylamine in methanol to give (R)- and (S)-1 in optically pure forms.

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After the formation of dianions of a carboxylic acid with lithium diisopropylamide, oxygen was bubbled into the solution to produce 2-hydroperoxy acid. Then the reaction mixture was acidified with a 2 N HCl solution and subsequently elevated to 50 °C to afford the aldehyde with the loss of one carbon atom. Even saturated (C₁₀–C₂₀) and unsaturated (C_18:1) carboxylic acids were converted into the odd aldehydes (C₉–C₁₉, C_17:1) in high yields. This conversion was found to be an efficient method for the preparation of carboxylic acids (Cn) to one-carbon degraded aldehydes (Cn-1) via 2-hydroperoxy acids.

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Annexins (ANXs) are a family of calcium dependent phospholipid binding proteins. Phospholipids such as phosphatidylserine are rapidly exposed on the surfaces of injured endothelial cells, activated platelets, and apoptotic cells in a large number of disorders. In this study, annexin V and XI (ANXV and ANXXI) were individually fused to the C-terminal of staphylokinase (SAK), a fibrin-selective thrombolytic protein, to form chimeras for evaluation of their in-vitro thrombolytic activities. The two chimeras were found to have plasminogen activation activity of comparable efficiency. When the chimeras were challenged under higher concentrations of plasmin for 1 h, hydrolysis of them into moieties was not seen on SDS–PAGE. In two thrombolytic assays, SAK-ANXXI was found to resolve both platelet rich plasma (PRP) clots and platelet poor plasma (PPP) clots with an efficiency similar to that of SAK. However, SAK-ANXV showed significantly reduced efficiency. With regard to anticoagulation ability, SAK-ANXXI was also found to have a stronger effect on dose-dependent extension of clotting time among the four tested proteins. The unique long N-terminal tail of ANXXI, composed of 202 residues, in contrast to the 16 residues of ANXV, probably served successfully to dispatch two moieties to function properly in a complicated microenvironment. Hence, a new option other than the most committed ANXV for the ANX based chimera without elaboration of linker construction is presented.

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The antioxidant properties of aqueous extracts from the dinophycean flagellates Gymnodinium impudicum and Alexandrium affine and the raphidophycean flagellate Chattonella ovata were examined. An electron spin resonance (ESR)-spin trapping method coupled with steady state kinetic analysis showed that all of the extracts directly scavenge superoxide, and that the superoxide scavenging potential of any of the extracts was comparable to that of L-ascorbic acid. As for hydroxyl radical scavenging, the Fenton reaction and the method of ultraviolet radiation to hydrogen peroxide were used as hydroxyl radical generation systems. All of extracts reduced the level of hydroxyl radicals in both of the systems, indicating that the extracts also directly scavenge hydroxyl radicals. Since the levels of phenolic compounds did not correlate with the antioxidant activities of the extracts, substances other than phenolic compounds also appeared to be attributable to the activities. It is of our interest that the scavenging activities of extract from G. impudicum against superoxide and hydroxyl radicals were increased by heat exposure at 100 °C and 200 °C respectively. Although the reason for the increased activities of the aqueous extract from G. impudicum is not clear, the heat-resistance of the extract from G. impudicum might make it a desirable antioxidant.

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The purpose of this study was to investigate the hepatoprotective effects of a fermented substance from Aspergillus phoenicis (FSAP) on chronic liver injuries induced by carbon tetrachloride (CCl₄) in rats. CCl₄ (20%; 0.2 ml/100 g body weight) was given twice a week for 9 weeks, and the rats received FSAP throughout the whole experimental period. Plasma ALT and AST, spleen weight, and hepatic levels of lipid peroxidation and hydroxyproline were significantly lower in the rats treated with FSAP as compared to CCl₄ only. Liver pathology in the FSAP-treated rats was also improved. Real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) analysis showed that FSAP treatment increased the expression of matrix metalloproteinase 13 and decreased the expression of methionine adenosyltransferase 2A, collagen (α1)(I), collagen (α1)(III), transforming growth factor-β1, and tissue inhibitor of metalloproteinase 1. These results clearly indicate that FSAP partially reduced the liver fibrosis in rats induced by CCl₄.

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Tumor conditioned medium (CM) has been widely used to stimulate endothelial cells to form capillary-like structures in in vitro angiogenesis models. We report herein the effect of HT1080 and A549 CM after they were mixed with microvascular endothelial cells medium-2 (EGM-2) on angiogenesis in human umbilical vein endothelial cells (HUVECs). Both HT1080 and A549 CM decreased HUVEC proliferation, to different extents. While A549 CM significantly increased capillary-like structure formation in a co-culture system, no effect of HT1080 was apparent. Inhibition of p38 mitogen-activated protein kinase (MAPK) blocked both basal and A549 CM induced capillary-like structure formation, but inhibition of extracellular signal-regulated kinases (ERK) and that of c-Jun N-terminal protein kinases (JNK) MAPK had no such effect. Activation of ERK MAPK was inhibited by both CMs, whereas p38 MAPK was inactivated by HT1080 and activated by A549 CM and a control. Neither CM had an effect on JNK MAPK. The results suggest that p38 MAPK played a critical role in capillary-like structure formation in the co-culture, partly via promotion of apoptosis in HUVECs.

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Four genes encoding alcohol dehydrogenase (Adh) isozymes in the thermotolerant yeast Kluyveromyces marxianus, a potent candidate for ethanol production at high temperatures, were investigated. Of these, KmADH3 and KmADH4 were cloned and sequenced, and their deduced amino acid sequences were compared with those of KmAdh1 and KmAdh2 and other Adhs of Kluyveromyces lactis and Saccharomyces cerevisiae. The four KmAdhs had high sequence similarity, though KmAdh3 and KmAdh4 possessed an amino-terminal extension as a mitochondrial targeting sequence, and appear to belong to the zinc-containing Adh family. These results and the results of Southern blot experiments suggest that there are at least four Adh isozymes in K. marxianus, two cytoplasmic enzymes and two mitochondrial enzymes. The expression profile revealed that KmADH genes are differently expressed depending on growth phase and carbon source, suggesting that these highly homologous Adhs play distinctive roles in cells.

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The model dicotyledon Arabidopsis thaliana has a characteristic small sub-family of phytochrome-interacting bHLH (basic Helix-Loop-Helix) factors, which are collectively designated the PIL (or PIF) (PHYTOCHROME INTERACTING FACTOR-LIKE) family proteins. In this study, we identified and characterized a set of highly homologous members (designated OsPIL11 to OsPIL16) in the model monocotyledon rice (Oryza sativa). Some of them (OsPIL11, OsPIL12, and OsPIL13) showed the ability to interact with the putative OsPRR1 (PSEUDO-RESPONSE REGULATOR 1) clock component, as far as the results of yeast two-hybrid assays were concerned. It was found that the expression of OsPIL13 is under the control of circadian rhythms (clock), while the expression of OsPIL15 is negatively regulated by light upon the onset to light exposure of etiolated seedlings. When the rice genes (OsPIL11 to OsPIL15) were over-expressed in A. thaliana, the resulting transgenic seedlings displayed anomalous morphologies with very long hypocotyls during early photomorphogenesis. These results suggest the view that the identified OsPILs are functional counterparts (or orthologs) of AtPILs, which are known to play important roles in red light-mediated (phyA and/or phyB-dependent) signal transduction pathways at immediate positions downstream of the photoreceptor in A. thaliana.

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A balsam pear (Momordica charantia L.) chitinase (Mcchit1) was purified and sequenced at the N-terminal. The genomic and cDNA coding sequences of Mcchit1 were cloned by rapid amplification of 3′ cDNA ends (3′-RACE) and the Y-shaped adaptor dependent extension (YADE) method. Sequence analysis showed that the Mcchit1 protein is a class I chitinase containing a chitin-binding domain and a catalytic domain, but no C-terminal extension. Northern blot indicated that the Mcchit1 transcription is wound-inducible. Overexpression of Mcchit1 dramatically increased intercellular and intracellular endochitinase activities, suggesting that the Mcchit1 gene encodes a secretory endochitinase. It was also found that overexpression of Mcchit1 significantly enhanced resistance to the plant pathogenic fungus Phytophthora nicotianae in transgenic N. benthamiana plants and against Verticillium wilt in transgenic cottons, indicating that the Mcchit1 gene can be a useful gene in plant engineering against fungal diseases.

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Nuclear lamins are a type of intermediate filament (IF) proteins. They have a characteristic tripartite domain structure with a α-helical rod domain flanked by non-α-helical N-terminal head and C-terminal tail domains. While the head domain has been shown to be important for the formation of head-to-tail polymers that are critical assembly intermediates for lamin IFs, essential structural elements in this domain have remained obscure. As a first step to remedy this, a series of mouse lamin A mutants in which the head domain (30 amino acid residues) was deleted stepwise from the N-terminus at intervals of 10 residues were bacterially expressed. The assembly properties in vitro of the purified recombinant proteins were explored by electron microscopy. We observed that while a lamin A mutant lacking N-terminal 10 residues formed head-to-tail polymers, a mutant lacking N-terminal 20 residues or the whole head domain (30 residues) showed significantly decreased potency to form head-to-tail polymers. These results suggest that the last 20 residues (from Arg-11 to Gln-30) of the head domain of mouse lamin A contain essential structures for the formation of head-to-tail polymers. The last 20 residues of the head domain include several conserved residues between A- and B-type lamins and also the phosphorylation site for cdc2 kinase, which affects lamin IF organization in vivo and in vitro. Our results provide clues to the molecular mechanism by which the head domain plays a crucial role in lamin polymerization.

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We found that appropriate treatment with a highly potent and long-lasting abscisic acid analog enhanced the tissue expansion of scutellum during early seedling development of rice, accompanied by increases of protein and starch accumulation in the tissue. A comparative display of the protein expression patterns in the abscisic acid analog-treated and non-treated tissues on two dimensional gel electrophoretogram indicated that approximately 30% of the scutellar proteins were induced by abscisic acid. The abscisic acid-induced proteins included sucrose metabolizing, glycolytic, and ATP-producing enzymes. Most of these enzyme proteins also increased during the seedling growth. In addition, the expression of some isoforms of UDP-glucose pyrophosphorylase, 3-phosphoglycerate kinase, and mitochondrial ATP synthase beta chain was stimulated in the scutellum, with suppressed expression of α-amylase. We concluded that abscisic acid directly and indirectly stimulates the expression of numerous proteins, including carbohydrate metabolic enzymes, in scutellar tissues.

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In Arabidopsis thaliana, each member of a large family of AS2/LOB (ASYMMETRIC LEAVES 2/LATERAL ORGAN BOUNDARIES) genes encodes a plant specific protein. They are highly homologous to one other. A mutational lesion in the representative AS2 gene results in the development of anomalous asymmetric leaves, implying that these family members commonly play some roles in plant development. In this study, we found that ectopic overexpression of ASL9 (ASYMMETRIC LEAVES 2 LIKE 9) in transgenic plants displayed a markedly anomalous architecture during the development of adult plants. Then we found that among AS2/LOB family members, ASL9 is distinct from the others in that it is exclusively regulated by the plant hormone cytokinin in a manner dependent on His-Asp phosphorelay signal transduction. We further found that when supplied externally in a medium, cytokinin specifically affected the growth properties of ASL9-ox seedlings. Taken together, the results of this study suggest that the cytokinin-induced ASL9 gene is implicated in regulation of the development of Arabidopsis thaliana.

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The pH dependence of the reaction of various renins was investigated using sheep angiotensinogen as a substrate. Human renin showed two separate peaks, but rat and mouse Ren1 renins showed one peak with a shoulder. A comparison of the predicted subsite residues of human renin with those of rat and mouse Ren1 renins revealed that Arg82, Ser84, Thr85, Ala229, and Thr312 are unique in the human sequence. We examined the possible importance of these residues in the unique pH profile of the human renin reaction by replacing these residues with the corresponding residues of rat renin. The replacement of Ser84 of human renin with Gly changed the pH dependence of the reaction to one peak, similarly to rat and mouse Ren1 renins. Other mutant human renins kept two separate peaks, similarly to wild-type human renin. These results indicate that Ser84 of human renin contributes to the biphasic pH dependence of the renin-angiotensinogen reaction.

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During an investigation of plant cell cultures that might be useful in the treatment of renal disorders, we established a vigorously-growing E-4 callus culture of Eritrichium sericeum that produced large amounts of caffeic acid metabolites, (−)-rabdosiin (1.8% dry wt) and rosmarinic acid (4.6% dry wt). Elicitation of the calli by methyl jasmonate induced a 38% increase in total polyphenol production. The most efficient method of eliciting (−)-rabdosiin biosynthesis was through the treatment of E-4 calli with cuprum glycerate, which induced an increase in (−)-rabdosiin production of as much as 4.1% dry wt. Oral administration of E-4 callus biomass (100 mg/kg/d for 30 d) to rats with induced Masugi-nephritis caused an increase in diuresis and lowered creatinine excretion and proteinuria levels as compared with Masugi-nephritis untreated rats. While all of the Masugi-nephritis untreated rats began to suffer, near a quarter of the E-4 treated rats remained in good health. This result indicates that the E-4 culture has the potential to alleviate the symptoms associated with nephritis.

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The ability to form amyloid fibrils from a wide range of proteins would open up the opportunity to augment studies of the molecular basis of amyloid fibril formation. We investigated 36 different conditions with respect to four model proteins to evaluate their ability to form amyloid fibrils. In a 5% ethanol solution at pH 2 at 57 °C, hen egg white lysozyme, bovine β-lactoglobulin, and bovine trypsinogen formed mature-type fibrils, while only histone H2A formed immature-type fibrils. Under these conditions, 25 of the 38 proteins formed amyloid fibrils. In addition, three additional proteins formed fibrils in a solution containing 5% trifluoroethanol instead of 5% ethanol. In summary, a total 28 proteins formed amyloid fibrils. Under these extreme conditions, chemical fragmentation was observed. Destabilization of the native structure, strengthening of hydrogen bonds, and chemical fragmentation are thought to play important roles in the formation of amyloid fibrils.

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To examine the possibility of module interaction in the thermal unfolding of different modular architectures, four truncated proteins were constructed from Clostridium stercorarium Xyn10B: a family 10 catalytic module (CM10), a polypeptide compound of one family 22 carbohydrate-binding module (CBM22-2) and the catalytic module (CBM22-CM10), two family 22 CBMs and the catalytic module (2CBM22-CM10), and only two family 22 CBMs (2CBM22). Thermal unfolding of four proteins were observed by differential scanning calorimetry. CM10 was unfolded reversibly and denatured as one component. The unfolding of protein CBM22-CM10 comprising CBM22-2 connected with CM10 was irreversible, and can be assumed to be one-component denaturation. Protein 2CBM22, with two CBM22s in tandem, unfolded as two independent modules. However, 2CBM22-CM10, with two CBM22s, unfolded as two and not the expected three separate components. These findings constitute the first reported case in which differences in thermal unfolding units and mechanisms were derived from differences in the modular architectures of proteins.

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The production of insulin-like growth factor-binding protein-1 (IGFBP-1) in HepG2 was increased by cadmium cation (Cd²⁺) at 3 μM, but not by other divalent cations. The mRNA level of IGFBP-1 was also increased by the administration of 3 μM of Cd²⁺. These results suggest that Cd²⁺ impacts the gene expression of IGFBP-1, which leads to production of IGFBP-1.

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A treX in the trehalose biosynthesis gene cluster of Sulfolobus solfataricus ATCC 35092 has been reported to produce TreX, which hydrolyzes the α-1,6-branch portion of amylopectin and glycogen. TreX exhibited 4-α-D-glucan transferase activity, catalyzing the transfer of α-1,4-glucan oligosaccharides from one molecule to another in the case of linear maltooligosaccharides (G3–G7), and it produced cyclic glucans from amylopectin and amylose like 4-α-glucanotransferase. These results suggest that TreX is a novel isoamylase possessing the properties of 4-α-glucanotransferase.

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β-Glucan was recently shown to have the ability to enhance and stimulate the immune system in humans, but little is known about its the anti-inflammatory effects. We investigated the effect of β-glucan on the production of tumor necrosis factor-alpha (TNF-α), a major pro-inflammatory mediator, in lipopolysaccharide (LPS)-stimulated BV2 microglial cells. β-Glucan decreased the production and expression of TNF-α. In addition, it blocked LPS-stimulated activation of nuclear factor kappa B (NF-κB). Hence β-glucan might suppress LPS-stimulated TNF-α production by inhibiting NF-κB in BV2 microglial cells.

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The novel flavonoid, leucocyanidin-3-O-β-D-glucoside, possessing a 4,2″-glycosidic linkage was isolated from green mature acerola (Malpighia emarginata DC.) puree and given the trivial name “aceronidin.” To examine the functions of aceronidin, its antioxidative activity and both its α-glucosidase and α-amylase inhibition activities, as a potential inhibitor of the sugar catabolic enzyme, were evaluated against those of taxifolin, catechin, isoquercitrin and quercitrin which each have a similar structure. The 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical quenching activity of aceronidin was stronger than that of α-tocopherol and comparable to that of flavonoids. In the yeast α-glucosidase inhibitory assay, aceronidin showed significantly greater inhibition than the other flavonoids tested. In the human salivary α-amylase inhibitory assay, aceronidin showed inhibition activity. Taken together, these results indicate aceronidin to be a potent antioxidant that may be valuable as an inhibitor of sugar catabolic enzymes.

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We investigated whether chylomicron formation is involved in the dietary phosphatidylcholine (PC)-induced increase in triglyceride (TG) absorption using an inhibitor of chylomicron formation, pluronic L-81 (L-81). In rats, cannulas were implanted into the duodenum (exps. 1 and 2) and the mesenteric lymph duct (exp. 1), and an emulsified lipid solution containing the test lipids (soybean oil, SO or soybean oil plus phosphatidylcholine, LE) with or without L-81 was infused through a duodenal cannula at a rate 3 ml/h for 2 h, and followed by infusion of a glucose–NaCl solution for 2 h. Mesenteric lymph was collected for 4 h (exp. 1). In exp. 2, the mucosa and contents of the small intestine were collected at 20, 40, or 90 min after the start of duodenal infusion of the test lipid to evaluate accumulation of lipids incorporated into the mucosa in the rats without a lymph cannula. In exp. 1, lymphatic TG outputs rapidly increased with infusion of both test lipids without L-81, but L-81 abolished these increases. TG accumulated in the small intestinal mucosa with L-81 treatment in a time-dependent manner, but the levels of accumulation were similar between the SO and LE groups (exp. 2). There were no differences in the amounts of lipid remaining in the small intestinal lumen between the L-81-treated SO and LE groups. These results indicate that uptake of lipid into the mucosal cells was not increased by LE. We conclude that the formation of chylomicron is responsible for increases in the promotive effect of a high level of dietary PC on the lymphatic absorption of TG.

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Previously, we found that an intraperitoneally administered chemotactic peptide, N-formyl-Met-Leu-Phe (fMLP), and MMK-1, a selective agonist of formyl peptide receptor-like 1 (FPRL1) receptor, the low affinity subtype of the fMLP receptor, prevented the alopecia in neonatal rats induced by the anticancer agent etoposide. The anti-alopecia effect of fMLP was not inhibited at all by Boc-FLFLF, a selective antagonist of formylpeptide receptor (FPR), which is the high affinity subtype of the fMLP receptor, but it was partly inhibited by Trp-Arg-Trp-Trp-Trp-Trp-NH₂ (WRW⁴), an antagonist of FPRL1 receptor. On the other hand, the anti-alopecia effect of MMK-1 was completely abolished by WRW⁴. The anti-alopecia effects of fMLP and MMK-1 were also inhibited by Lys-D-Pro-Thr (K(D)PT) and pyrrolidine dithiocarbamate, which are inhibitors of interleukin-1 (IL-1) and nuclear factor-κB (NF-κB) respectively. Hence, we suggest that the anti-alopecia mechanisms of intraperitoneally administered fMLP and MMK-1 include activation of NF-κB via IL-1 release downstream of the FPRL1 receptor homolog in rats.

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Hyperhomocysteinemia is associated with an increase in the incidence of vascular diseases, including retinal vascular diseases. We examined the effects of high plasma levels of homocysteine on retinal glial cells and vascular endothelial growth factor (VEGF) expression. Male Sprague-Dawley rats were fed either a 3.0 g/kg homocystine diet or a control diet for 2 week. The homocystine-diet group had higher plasma levels of homocysteine and thiobarbituric acid reactive substances (TBARSs) and lower plasma levels of folate, retinol, α-tocopherol, and retinal expression of CuZn superoxide dismutase (SOD) than the controls. The rats fed the homocystine-diet showed an increase in vimentin, glial fibrillary acidic protein (GFAP), and VEGF immunoreactivity in the retina as compared to the controls. The increase in vimentin immunoreactivity in the hyperhomocysteinemic rats was correlated with changes in GFAP immunoreactivity in astrocytes within the ganglion cell layer. We found for the first time that short-term hyperhomocysteinemia-induced oxidative stress activates retinal glial cells and increases VEGF expression in the retina.

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Acetate has been found to have an inhibitory effect on the activity of carbohydrate-responsive element-binding protein (ChREBP) in cultured hepatocytes, this being a transcription factor that regulates several genes required for the conversion of glucose to fatty acids in the liver. The aim of this study was to investigate whether an oral administration of acetate would contribute to reducing lypogenic genes and protecting against obesity. We orally injected 5.2 mg/kg BW of acetate to obesity-linked type 2 diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats. The treatment with acetate showed a marked reduction in lipid accumulation in the adipose tissue, protection against accumulation of fat in the liver, and improved glucose tolerance. An analysis by Northern blotting revealed that the transcripts of several lipogenic genes in the liver of OLETF rats were decreased by the acetate treatment. On the basis of those results, it was indicated that acetate was a potential compound to improve obesity and obesity-linked type 2 diabetes.

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Fulvic acid (FA) was extracted and purified from Canadian Sphagnum peat (CP-FA) and characterized by using an element analysis meter, Fourier transform infrared (FT-IR) spectroscopy, electron spin resonance (ESR) spectroscopy, and ¹³C-nuclear magnetic resonance (¹³C-NMR) spectroscopy. To investigate the antiallergic effect of CP-FA, we incubated rat basophilic leukemia (RBL-2H3) cells with 0.001–10.0 μg/ml of CP-FA and determined the β-hexosaminidase release inhibition at different response stages. The intracellular calcium [Ca²⁺]_i level was also determined by using Fluo 3-AM, a calcium-specific fluorescent probe, and the cytotoxicity of CP-FA was determined by the 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay. The results revealed that RBL-2H3 cells incubated for 48 h with 0.001–10.0 μg/ml of CP-FA did not show any decreased viability. CP-FA inhibited the β-hexosaminidase release by IgE-sensitized, antigen-stimulated RBL-2H3 cells at the antigen-antibody binding stage and the antibody-receptor binding stage. CP-FA also inhibited histamine release from A23187 plus PMA- or compound 48/80-stimulated KU812 cells. Furthermore, there was a decrease in the intracellular [Ca²⁺]_i level in IgE-sensitized cells incubated with CP-FA and stimulated with antigen. Our results show that CP-FA may be useful for the treatment or prevention of allergic diseases.

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Cancer protection associated with the consumption of olive products is well established, but not for leukemia. The protective effects of olive (Olea europaea L.) leaves were investigated by incubating human promyelocytic leukemia HL-60 cells with olive leaf extracts (OLEs) from seven principal Tunisian olive varieties, namely, Chemchali, Chemlali, Chétoui, Gerboui, Sayali, Zalmati and Zarrazi. The results showed significant growth inhibition of HL-60 cells incubated for 48 h with a 100-fold dilution of each OLE which had been obtained by incubating 10 g of dried leaves in 100 ml of 70% ethanol for one week with subsequent ultrafiltration. DNA fragmentation was observed in the cells incubated for 19 h with a 100-fold dilution of the Chemchali, Chemlali and Zalmati extracts. The results of a nitroblue tetrazolium (NBT) assay revealed NBT reduction, a differentiation marker, by the OLE-treated cells after an overnight incubation. The Gerboui extract showed the highest NBT reduction ability at more than 90%. An HPLC analysis revealed the presence of apigenin 7-glucoside in the extract, which was found in subsequent experiments to be responsible for the Gerboui extract-mediated cell differentiation.

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We determined the effects of yolk water-soluble protein (YSP) on bone formation in pre-osteoblastic MC3T3-E1 cells. YSP (50–5,000 μg/ml) increased cell proliferation and collagen content. Alkaline phosphatase (ALP) activity was also increased by YSP treatment. After enhancement of ALP activity, significant augmentation of calcification was observed. These results suggest that YSP is a promising agent for the prevention and treatment of bone loss.

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A polysaccharide-rich substance isolated from black currant, named cassis polysaccharide (CAPS), was partially digested with β-galactosidase from Aspergillus oryzae and its immunostimulatory activity was investigated. The in vitro cytokine-inducing effect of CAPS on RAW264 cells was gradually decreased along with lowering of the average MW of CAPS. In vivo, partially digested CAPS with a mean MW of approximately 20,000 showed the most potent antitumor activity against Ehrlich carcinoma in mice.

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Jejunal GLUT5 is elevated with triiodothyronine (T₃) during weaning of rats. A perfusion of fructose into the small intestine of T₃-injected rats at 21 d induced expression of the GLUT5 gene, but one into that of vehicle-injected rats did not. These results suggest that T₃ and fructose coordinately enhance jejunal expression of the GLUT5 gene in rats during weaning period.

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We examined the effects of red potato flakes (RPF) on serum antioxidant potential and hepatic mRNA in rats. The serum thiobarbituric acid-reactive substances concentration and hepatic superoxide dismutase mRNA level in rats fed RPF were significantly lower and higher respectively than those in control rats. These results suggest that RPF might improve the antioxidant system by enhancing hepatic SOD mRNA.

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Four kinds of polychlorinated biphenyl (PCB)-degrading Rhodococcus sp. (TA421, TA431, HA99, and K37) have been isolated from termite ecosystem and under alkaline condition. The bph gene cluster involved in the degradation of PCB/biphenyl has been analyzed in strain TA421. This gene cluster was highly homologous to bph gene clusters in R. globerulus P6 and Rhodococcus sp. RHA1. In this study, we cloned and analyzed the bph gene cluster essential to PCB/biphenyl degradation from R. rhodochrous K37. The order of the genes and the sequence were different in K37 than in P6, RHA1, and TA421. The bphC8_K37 gene was more homologous to the meta-cleavage enzyme involved in phenanthrene metabolism than bphC genes involved in biphenyl metabolism. Two other Rhodococcus strains (HA99 and TA431) had PCB/biphenyl degradation gene clusters similar to that in K37. These findings suggest that these bph gene clusters evolved separately from the well-known bph gene clusters of PCB/biphenyl degraders.

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The influence of carbon sources on bacterial community structure in the gut of the wood-feeding higher termite Nasutitermes takasagoensis was investigated. 16S rRNA gene sequencing and terminal-restriction fragment length polymorphism (T-RFLP) analyses revealed that the bacterial community structure changed markedly depending on feed components at the phylum level. Spirochaetes was predominant in the clone libraries from wood- and wood powder-fed termites, whereas Bacteroidetes was the largest group in the libraries from xylan-, cellobiose-, and glucose-fed termites, and Firmicutes was predominant in the library from xylose-fed termites. In addition, clones belonging to the phylum Termite Group I (TG1) were found in the library from xylose-fed termites. Our results indicate that the symbiotic relationship between termite and gut microorganisms is not very strong or stable over a short time, and that termite gut microbial community structures vary depending on components of the feeds.

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The ability of fungi used in the preparation of fermented soybean foods to metabolize the soy isoflavones daidzein and genistein was investigated. A total of 21 fungal strains from dou-chi, miso, sake, soy sauce, and sufu were screened. The genera of the tested fungi included Actinomucor, Aspergillus, Candida, Debaryomyces, Monascus, Mucor, Rhizopus, Saccharomyces, and Zygosaccharomyces. The results were that all tested Aspergillus strains from these soybean foods, including five A. oryzae strains, one A. sojae strain, and one A. tamarii strain, metabolized both daidzein and genistein. In contrast, no other tested fungi from the fermented soybean foods metabolized either daidzein or genistein. The metabolites of daidzein and genistein by Aspergillus strains were identified as 8-hydroxydaidzein and 8-hydroxygenistein, respectively, based on their mass, ¹H-, and ¹³C-NMR spectra.

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We focused on the effects of a mutation of xylose reductase from Pichia stipitis (PsXR) on xylose-to-ethanol fermentation using recombinant Saccharomyces cerevisiae transformed with PsXR and PsXDH (xylitol dehydrogenase from P. stipitis) genes. Based on inherent NADH-preferring XR and several site-directed mutagenetic studies using other aldo-keto reductase enzymes, we designed several single PsXR mutants. K270R showing decreased NADPH-preferring activity without a change in NADH-preferring activity was found to be a potent mutant. Strain Y-K270R transformed with K270R PsXR and wild-type PsXDH showed a 31% decrease in unfavorable xylitol excretion with 5.1% increased ethanol production as compared to the control in the fermentation of 15 g l⁻¹ xylose and 5 g l⁻¹ glucose.

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